Datasets:

hippocrates
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PubMed_Summ_test

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Split (3)

train · 120k rows

id stringlengths 11 16	query stringlengths 279 826k	answer stringlengths 174 4.34k
PubmedSumm2000	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: in august 2013 , a 9-year - old girl who was returning to france after a 1-month stay with her family in casablanca , morocco , was admitted to lille university hospital in lille . symptoms started in casablanca 3 days before hospital admission and included fever , severe diarrhea ( > 10 liquid defecations / day ) , vomiting , and abdominal pain in the hypogastric area and in the right and left lower quadrants associated with bilateral dorsal pain , anorexia , and weakness . blood count showed 13,850/mm leukocytes ( 75.4% neutrophils , 15.9% lymphocytes , 8.5% monocytes ) . c - reactive protein level was increased at 266 mg / l ( low risk : < 1.0mg / l ; average risk : 1.03.0 mg / l ; high risk > 3.0 mg / l ) . traveler s gastroenteritis was diagnosed , and symptomatic treatment with acetaminophen , phloroglucinol glucoside , and acetorphan was prescribed . however , abdominal pain increased , and total food intolerance occurred in the following hours . an abdominal ultrasound was performed , revealing appendicitis with suppuration in the recto - uterine pouch and a reflex ileus . parasitologic examination of fecal matter revealed only abundant blastocystis vacuolar forms , with > 5 parasites per field . we further confirmed absence of cryptosporidium spp . using glycerin assay and real - time pcr . yeasts and a multimicrobial flora were present in the fecal material , but other infectious agents such as salmonella spp . , shigella spp . similarly , multimicrobial flora , but no pathogenic bacteria , were detected in the peritoneal liquid and recto - uterine pouch ( table ) . histopathologic observation revealed acute suppurative appendicitis with ulcerations extending deep into the muscularis , which was covered with a suppurative and fibrinous exudate . we observed infiltration by numerous neutrophils , eosinophils , plasma cells , and lymphocytes through all layers and into the serous membrane ( figure , panel a ) . micrographs showing histopathologic examination of appendix samples from a child who had peritonitis , casablanca , morocco , 2013 . a ) ulceration ( u ) covered with suppurative and fibrinous exudates ( se ) ( hematoxylin - eosin stain ) . b ) blastocystis parasites ( arrows ) in the lumen ( l ) , and at the surface of the epithelium ( e ) ( hematoxylin - eosin stain ) . c ) blastocystis parasites ( arrows ) in the lumen , at the surface of the epithelium and in the lamina propria ( lp ) of the mucosa ( immunofluorescence labeling with anti - blastocystis paraflorb antibody ) . after hematoxylin - eosin staining and immunofluorescence labeling by using the anti - blastocystis paraflor b monoclonal antibody ( boulder diagnostics , marlborough , ma , usa ) , we detected parasitic forms in the lumen and in the lamina propria of the mucosa ( figure , panels b , c ) . we used real - time pcr for blastocystis parasite detection as described ( 4 ) , using dna extracted from stools , appendix , peritoneal liquid , and the recto - uterine pouch , which all tested positive . we subsequently performed small subunit rdna ( ssu rdna ) amplification , then cloned the pcr product and sequenced 5 clones from all the dna samples to subtype ( st ) blastocystis isolates and detect mixed infections ( 5 ) . the ssu rdna gene sequences obtained in this study have been deposited in genbank under accession nos . the child completely recovered after an appendectomy , removal of a stercolith from the appendix lumen , and treatment with tinidazole , 20 mg / kg / d , and ceftriaxone , 50 mg / kg / d for 10 days , together with gentamicin , 5 mg / kg / d for 5 days . although tinidazole is not the first line medication for treatment of blastocystis infection , the child recovered completely and showed total clearance of parasites at day 73 : using microscopy and real - time pcr on fecal samples , we found negative results for blastocystis . data obtained from the child s mother revealed simultaneous cases of gastroenteritis in 26 family members : 13 adults , 3498 years of age , and 13 children , 18 months15 years of age , who lived in the same building at the residential mohammadi area of casablanca . adults had mild or moderate diarrhea but symptoms were more severe in children , who all had abundant diarrhea , vomiting , and weight loss . repatriation in france of an 18-month - old baby was considered , but his condition improved . unfortunately , no explorations were performed , therfore the diagnosis could not be microbiologically documented . reports of blastocystis infection associated with diarrhea and clinical symptoms in immunocompetent and immunocompromised patients have increased during the past 2 decades ( 2 ) . tissue invasion of blastocystis parasites in the appendix ( 6 ) or in the colon mucosa ( 3 ) , associated with acute or chronic inflammation , has been reported . however , controversy still exists over whether this parasite is commensal or pathogenic ; this case further supports its invasive and inflammatory potential . previous reports regarding the presence of blastocystis parasites in 4 of 100 appendix specimens from patients with acute appendicitis ( 7 ) , and of pseudoappendicular illness , which led to appendectomies in children with intestinal infection caused by this parasite ( 8) , suggest that blastocystis infection could be associated with appendicitis . nevertheless , the actual role of blastocystis in the pathogenesis of appendicitis remains inconclusive . in this report , the presence of a stercolith , which can be found in 50%80% of appendicitis cases , suggested mechanical obstruction of the appendix s lumen , which is the main etiology of appendicitis . here , we report dissemination of blastocystis into the lumen , the mucosa , and the recto - uterine pouch exudate , associated with appendicular acute inflammation , and no other infectious agent was detected . these observations , together with the well - documented acute or chronic inflammation occurring in humans or animals with blastocystis infections ( 3,9 ) , likely support the contribution of this infection to the inflammatory process . indeed , the presence of pathogenic strains among st3 has been confirmed through experimental infections in rats ( 9 ) . additionally , a substantial inflammatory reaction and an increased propagation of human colorectal cancer cells exposed to blastocystis st3 antigens has been demonstrated in vitro ( 10 ) . for humans , the pathogenicity of different sts is unclear and remains a debatable issue . st1 isolates were found to be more prevalent among symptomatic patients in lebanon ( 5 ) , but st3 was found to be the only st that showed pathogenic potential in malaysian patients when compared with st1 and st2 ( 11 ) . st3 was also found to be significantly associated with diarrhea in libya ( p = 0.008 ) ( 12 ) . for this case , the fact that only st3 was detected in all analyzed samples , whereas a mixed infection with st2 and st3 was found in the child s stools , further supports the high invasive potential of st3 . st3 is the most common st in europe , but , in african countries , its frequency varies from 17.8% in libya ( 12 ) to 61.9% in egypt ( 13 ) . in morocco , a 28.7% prevalence of blastocystosis has been reported , but data concerning the st distribution of the parasite are not available ( 14 ) . furthermore , although blastocystis infection could not be confirmed among the child s relatives , the simultaneous occurrence of gastroenteritis cases in the same family and the absence of other infectious agents in the child s stools suggest a potential outbreak of blastocystis infection . blastocystis parasites could have spread within the child s family , as previously reported in italy , where 2 adopted children originating from india and the cte divoire transmitted blastocystis parasites to their adoptive parents and grandmother ( 15 ) . possible acquisition of this parasite from a common source such as contaminated water could also explain family transmission in this report . altogether , these data highlight 1 ) the need for both systematic parasitologic examinations of stools in patients with invasive infections who are traveling from countries with high blastocystis prevalence and 2 ) the need for routine provision of imidazoles for empiric treatment of peritonitis . Output:	despite increasing reports that blastocystis infection is associated with digestive symptoms , its pathogenicity remains controversial . we report appendicular peritonitis in a 9-year - old girl returning to france from morocco . only blastocystis parasites were detected in stools , appendix , peritoneal liquid , and recto - uterine pouch . simultaneous gastroenteritis in 26 members of the child s family suggested an outbreak .
PubmedSumm2001	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: terra preta soils were likely formed between 500 and 2500 years ago by pre - columbian populations . these soils are found in the amazonian region distinguished by the presence of ceramic artifacts , animal bones and high amounts of charred organic materials , which are commonly referred as black carbon ( bc ) . ade soils are considered as model soils because their high chemical fertility ( high content levels of organic c , p , ca , mg , zn , and mn ) and stable organic matter . also , ade soils are characterized by their distinct microbial communities , with a higher diversity and richness when compared to the surrounding soils . the aim of this study was to investigate the taxonomic and functional diversity of two ade sites based on metagenomic approach . soils were collected in the hatahara site located in the municipality of iranduba ( 0316494s 6012340w ) , in the amazonas state , brazil . dna extraction from 250 mg of soil was carried out using powersoil dna isolation kit ( mobio laboratories , carlsbad , ca , usa ) , according to the manufacturer 's protocol . dna quality and concentration were measured by agarose gel electrophoresis and nanodrop 1000 spectrophotometer ( thermo scientific , waltham , ma , usa ) . soil dna samples were used to prepare paired - end libraries ( 2 300 bp ) using the miseq reagent kit v.2 ( 500 cycles ; illumina , san diego , ca , usa ) , for further shotgun metagenomic sequencing in a miseq personal sequencing system ( illumina , san diego , ca , usa ) . a total of 3,716,966 paired - end and non - paired forward sequences ( average length of 241 bp ) were joined using the pear software . metagenomic sequences were annotated using the mg - rast automatic annotation server ( . ) . exploratory data analysis was conducted in stamp software . the most abundant phyla detected were proteobacteria ( 40 2% ) , actinobacteria ( 18 1% ) , firmicutes ( 5 0.3% ) and acidobacteria ( 4 0.5% ) . others phyla detected were planctomycetes ( 3.5 0.4% ) , chloroflexi ( 2.5 0.3% ) , cyanobacteria ( 2.5 0.1% ) , bacteroidetes ( 2 0.2% ) and verrucomicrobia ( 2 0.2% ) . the archaea diversity was represented by the phyla crenarchaeota , euryarchaeota , korarchaeota , nanoarchaeota and thaumarchaeota , with approximately 1.5 0.5% of total abundance . protein annotations were grouped into functional categories and the most abundant functions were related to carbohydrates ( 15 0.5% ) , clustering - based subsystems ( 14 0.2% ) , aminoacids ( 11 0.16% ) , miscellaneous ( 7 0.03% ) and protein metabolism ( 6 0.16% ) . the most abundant category level 3 is associated with serine - glyoxylate cycle ( 1.8 0.5% ) . we hypothesize that these functions are related to methylotrophs and methane reducing on ade soils , but further studies are necessary to validate this inference . our dataset reveals a great taxonomic and functional microbial potential to be explored by bioprospecting of new enzymes or identifying unknown microorganisms . terra preta soils were likely formed between 500 and 2500 years ago by pre - columbian populations . these soils are found in the amazonian region distinguished by the presence of ceramic artifacts , animal bones and high amounts of charred organic materials , which are commonly referred as black carbon ( bc ) . ade soils are considered as model soils because their high chemical fertility ( high content levels of organic c , p , ca , mg , zn , and mn ) and stable organic matter . also , ade soils are characterized by their distinct microbial communities , with a higher diversity and richness when compared to the surrounding soils . the aim of this study was to investigate the taxonomic and functional diversity of two ade sites based on metagenomic approach . soils were collected in the hatahara site located in the municipality of iranduba ( 0316494s 6012340w ) , in the amazonas state , brazil . dna extraction from 250 mg of soil was carried out using powersoil dna isolation kit ( mobio laboratories , carlsbad , ca , usa ) , according to the manufacturer 's protocol . dna quality and concentration were measured by agarose gel electrophoresis and nanodrop 1000 spectrophotometer ( thermo scientific , waltham , ma , usa ) . soil dna samples were used to prepare paired - end libraries ( 2 300 bp ) using the miseq reagent kit v.2 ( 500 cycles ; illumina , san diego , ca , usa ) , for further shotgun metagenomic sequencing in a miseq personal sequencing system ( illumina , san diego , ca , usa ) . a total of 3,716,966 paired - end and non - paired forward sequences ( average length of 241 bp ) were joined using the pear software . metagenomic sequences were annotated using the mg - rast automatic annotation server ( . ) . exploratory data analysis was conducted in stamp software . the most abundant phyla detected were proteobacteria ( 40 2% ) , actinobacteria ( 18 1% ) , firmicutes ( 5 0.3% ) and acidobacteria ( 4 0.5% ) . others phyla detected were planctomycetes ( 3.5 0.4% ) , chloroflexi ( 2.5 0.3% ) , cyanobacteria ( 2.5 0.1% ) , bacteroidetes ( 2 0.2% ) and verrucomicrobia ( 2 0.2% ) . the archaea diversity was represented by the phyla crenarchaeota , euryarchaeota , korarchaeota , nanoarchaeota and thaumarchaeota , with approximately 1.5 0.5% of total abundance . protein annotations were grouped into functional categories and the most abundant functions were related to carbohydrates ( 15 0.5% ) , clustering - based subsystems ( 14 0.2% ) , aminoacids ( 11 0.16% ) , miscellaneous ( 7 0.03% ) and protein metabolism ( 6 0.16% ) . the most abundant category level 3 is associated with serine - glyoxylate cycle ( 1.8 0.5% ) . we hypothesize that these functions are related to methylotrophs and methane reducing on ade soils , but further studies are necessary to validate this inference . our dataset reveals a great taxonomic and functional microbial potential to be explored by bioprospecting of new enzymes or identifying unknown microorganisms . Output:	the anthropogenic amazon dark earth soil is considered one of the world 's most fertile soils . these soils differs from conventional amazon soils because its higher organic content concentration . here we describe the metagenome sequencing of microbial communities of two sites of anthropogenic amazon dark earth soils from amazon rainforest , brazil . the raw sequence data are stored under short read accession number : prjna344917 .
PubmedSumm2002	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: smoking affects vascular and hormonal systems and is also involved in the development of atherosclerosis , thrombogenesis and vascular occlusion . chronic smoking adversely influences the prognosis of nephropathies . in primary hypertension , urinary albumin excretion as an index of renal damage the rate of progression to renal failure amongst smokers is twice in both type i and type ii diabetes . this study was intended for non - diabetic normotensive subjects to find out the effect of smoking on renal functions such as microalbuminuria , creatinine clearance and urinary albumin creatinine ratio ( acr ) . this was a community - based prospective cross - sectional cohort study conducted on 120 non - diabetic normotensive subjects as per the inclusion criteria . it was approved by the institutional ethics committee and conducted in accordance with the guidelines of the indian council of medical research , international conference on harmonization - good clinical practices instructions of drug controller general of india new delhi and institutional sop with a proper informed consent . inclusion criteria were age 30 - 70 years , normotensive ( 139/89 mmhg ) , non - diabetic ( fasting plasma glucose 125 mg / dl ) , non - obese ( body mass index [ bmi ] < 30 kg / m ) , no family history of premature vascular disease , normal total cholesterol level ( < 200 mg / dl ) , normal renal function ( urea 40 mg / dl and creatinine 1 mg / dl ) , clinically well and not on any regular cardiovascular medication and given informed consent . exclusion criteria were age less than 30 and above 70 years , diabetics or using insulin or oral hypoglycaemic agents , hypertensive or using antihypertensive medication , hyperlipidemic or using lipid lowering drugs , obese ( bmi 30 kg / m ) , abnormal renal function ( urea and creatinine ) , urinary tract infection , significant renal disease or using diuretics , angiotensin converting enzyme inhibitors , alcohol consumption or other significant drugs , fever , vigorous physical activity , menstruating / pregnant women and not willing to give consent . smoker was defined as anyone who had smoked at least 20 bidi / day for 5 years ( 5 pack - years ) or equivalent . smokers were divided into four sub - groups : very light smoker ( 5 - 9 pack - years ) , light smoker ( 10 - 14 pack - years ) , moderate smoker ( 15 - 19 pack - years ) and heavy smoker ( > 20 pack - years ) . out of 120 non - diabetic normotensive subjects , 80 ( 66.66% ) were smokers and 40 ( 33.33% ) were age matched non - smokers as control . the baseline physical characteristic and investigation were compared in these two groups using the statistical tests of significance ( p = ns ) [ table 1 ] . baseline subjects characteristics and their comparison in both groups overnight fasting blood sugar , blood urea , serum creatinine and lipid profile were measured and first morning void ( timed ) quantitative mid - stream urine was sample taken for screening of urinary albumin and urinary creatinine concentrations [ table 1 ] . laboratory methods used included ; clinitek micro albumin reagent strips ( urinary albumin and acr ) , jaffe colorimetric method ( serum creatinine concentration ) , glucose enzymatic colorimetric ( glucose oxidase / peroxidase ) method ( plasma glucose ) , enzymatic colorimetric cholesterol ( cholesterol oxidase / peroxidase aminophenazone ) method ( serum lipid profile ) , urine analyzer uriscan optima ii + combistix sg , korea ( urinary leukocyte and erythrocyte ) . all calculations were performed with the statistical package for the social sciences ( spss ) version 9.0 ( spss , chicago , usa ) software by chi - square analysis or analysis of variance . out of 120 non - diabetic normotensive subjects , 80 ( 66.66% ) were smoker and 40 ( 33.33% ) were non - smoker . 83 subjects ( 69.16% ) were male and 37 subjects ( 30.83% ) were female . the mean age in smoker group was 48.68 11.82 years and in non - smoker group was 46.10 10.77 years , ranged from 30 to 70 years ( p = 0.247 ) [ table 1 ] . the two groups were comparable in terms of all parameters except high - density lipoprotein ( hdl ) level ( p = 0.031 ) [ table 1 ] . smokers had higher mean urinary albumin level ( 52.84 mg / l ) than non - smokers ( 19.25 mg / l ) ( p < 0.0001 ) [ figure 1 ] . among smokers ( n = 80 ) , microalbuminuria was directly related to the amount of smoking ( pack - years ) [ figure 2 ] . seventy three smokers ( 91.25% ) and nine non - smokers ( 22.5% ) had urinary albumin level > 20 mg / l ( microalbuminuria ) . seven smokers ( 8.75% ) and 31 non - smokers ( 77.5% ) had urinary albumin level < 20 mg / l [ figure 3 ] . comparison of mean urinary albumin level between smoker ( n = 80 ) and non - smoker ( n = 40 ) relationship between quantity of smoking and mean urinary albumin in smoker comparison of smoker ( n = 80 ) and non - smoker ( n = 40 ) subjects for microalbuminuria smoker ( n = 80 ) had higher mean urinary acr ( 93.98 g / mg ) than non - smoker ( n = 40 , 18.99 g / mg ) ( p < 0.001 ) [ figure 4 ] . among smokers ( n = 80 ) , urinary acr level was directly related to the amount of smoking ( pack - years ) [ figure 5 ] . sixty four smokers ( 80% ) and two non - smokers ( 5% ) had urinary acr level > 30 g / mg . only 16 smokers ( 20% ) and 38 non - smokers ( 95% ) had urinary acr < 30 g / mg [ figure 6 ] . comparison of mean urinary albumin creatinine ratio level between smoker ( n = 80 ) and non - smoker ( n = 40 ) relationship between quantity of smoking and urinary albumin creatinine ratio in smoker comparison of smoker ( n = 80 ) and non - smoker ( n = 40 ) subjects for high urinary albumin creatinine ratio male and female smokers had no statistically significant distribution of urinary albumin , urinary creatinine and urinary acr ( p = ns ) . this study shows non - diabetic normotensive smoker had higher mean urinary albumin level , which is directly related to the amount of smoking ( pack - years ) among smokers . also , more smokers had microalbuminuria , which signify smokers have 4-fold higher prevalence of microalbuminuria than non - smokers . the heart outcome prevention evaluation study documented that smoking was an independent determinant of microalbuminuria in all participants , i.e. , non - diabetic and diabetic patients with a high cardiovascular risk profile . prevend study showed statistically significant difference in urinary albumin excretion in non - smokers and smokers . a recent study found that patients with hypertension and left ventricular hypertrophy smoking > 20 cigarettes / day had a 1.6-fold higher prevalence of microalbuminuria and a 3.7-fold higher prevalence of macroalbuminuria than never - smokers . several studies documented that smoking is an independent predictor of ( micro ) albuminuria in otherwise healthy hypertensive subjects . the prevalence of microalbuminuria is almost double in smoking than non - smoking lean patients with the primary hypertension . one of the underlying mechanisms by which smoking induces albuminuria and abnormalities in renal function is through advanced glycation end products ( ageps ) . ageps are cross - linking moieties formed from the reaction of reducing sugars and the amino groups of plasma proteins , lipids and nucleic acids . it is known that ageps are responsible for enhanced vascular permeability and that they accelerate vasculopathy of end - stage diabetic renal disease . , have shown that both aqueous extracts of tobacco and cigarette smoke contain glycotoxins , highly reactive glycation products that can rapidly induce agep formation on proteins in vitro and in vivo . it is reasonable to expect that the ageps formed by the reaction of glycotoxins from cigarette smoke with serum and tissue proteins will have the same effect on the systemic and renal vasculature as mentioned . another mechanism , based on the patho - physiological effect of smoking induced renal damage , is insulin resistance . several investigators have described smoking to be causally related to insulin resistance in non - diabetic subjects . insulin resistance has been known to be related to both albuminuria and abnormalities in renal function . both mechanisms act through endothelial dysfunction that is by inducing an imbalance between the contracting and relaxing substances produced by the endothelium . the plasma concentration of endothelin 14 has shown to be increased in smokers as compared to non - smokers , also indirect evidence available for a disturbance of endothelin , prostacyclin or nitric oxide release on stimulation in smokers . this study shows non - diabetic normotensive smoker had significantly higher mean urinary acr which is directly related to the amount of smoking ( pack - years ) among smokers . amongst non - smokers , females have low muscle mass compared with males ; therefore , this sex specific difference in acr is due to decreased urinary creatinine excretion in females . warram et al . , also proposed sex - specific acr cut points for microalbuminuria as 17 and 25 g / mg for men and women , respectively . a smoker had serum creatinine and creatinine clearance comparable with non - smoker , similar to prevend study . among smokers , both male and female had comparable urinary albumin , urinary creatinine and urinary acr ( p = ns ) , which signifies that the effect of smoking on renal function is not affected by gender . in this study , subjects having hypercholesterolemia ( total cholesterol > 200 mg / dl ) were excluded due to their known influence on renal function . smoker had significantly lower hdl than non - smokers , similar to several studies , which shows hdl is lower in smokers than in non - smokers . however , limitations of this study are a small number of subjects , single center data and screening with one timed urine sample , it is in comparison with one of the many ongoing prevention of renal and vascular end stage disease ( prevend ) group studies ( prevend ) , running in the city of groningen , the netherlands ( n = 7476 ) with the mean age of smoker ( 47 12 years who smoke < 20 cigarette / day , 46 10 years who smoke > 20 cigarette / day ) and non - smoker ( 47 13 years ) . this area of research needs further attention of physicians and nephrologists , looking to the highly prevalent smoking addiction in indian community as an independent risk factor and its impact on renal function , therefore ; more such studies can be planned at various centers . non - diabetic normotensive smokers have significantly higher level of urinary albumin and urinary acr than non - smoker , which is directly proportional to quantity of smoking . similarly , smokers have a higher prevalence of 4-fold for microalbuminuria and 16-fold for increased urinary acr than non - smokers . smoking significantly reduces the hdl level ; however , no significant effect on serum creatinine and creatinine clearance . Output:	smoking is associated with an excessive morbidity and mortality from a variety of diseases . the aim of this study was to find out the effects of smoking on renal function study in non - diabetic , normotensive subjects . a community - based , prospective , cross - sectional cohort study was conducted on 120 subjects ; 80 ( 66.66% ) were smokers and 40 ( 33.33% ) age matched non - smokers ; with age range of 30 to 70 years . measurement of fasting sugar , urea , creatinine , lipids and one time screening of urinary albumin and urinary creatinine was done . smokers had significantly higher urinary albumin and albumin creatinine ratio ( acr ) ( 52.84 46.42 mg / l , 93.98 78.68 g / mg ) than non - smokers ( 19.25 7.77 mg / l , 18.99 6.65 g / mg ) , respectively ( p = < 0.001 , p = < 0.001 ) . microalbuminuria and urinary acr level were directly related to the amount of smoking ( pack - years ) . among smokers , 73 ( 91.25% ) had microalbuminuria ( > 20 mg / l ) and 64 ( 80% ) had increased urinary acr ( > 30 g / mg ) . smoker had significantly lower high - density lipoprotein level ( 36.66 10.28 mg / dl ) compared to non - smokers ( 41.22 11.72 mg / dl ) ( p = 0.031 ) . urea , creatinine , creatinine clearance , total cholesterol , low density lipoprotein , triglyceride levels were comparable ( p = ns ) . in conclusion , smokers have a 4-fold higher prevalence of microalbuminuria than non - smokers .
PubmedSumm2003	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: ace inhibitors and arbs , the most potent antihypertensive and renoprotective pharmaceutical agents also recognized for their cardioprotective activity , have been widely applied to a number of illnesses . these agents function by interruption of the production or action of angiotensin ii , which is the main end - product of the renin clinical consequences include ( a ) reduction of glomerular capillary pressure , ( b ) reduction of systolic blood pressure and proteinuria and ( c ) an antiproliferative effect , all of which are considered major actions of these drugs . their side effects , which have been extensively studied in animals and humans , are classified into three categories : ( a ) those related to pharmacologic action , ( b ) those related to chemical structure and ( c ) those involving hypoperfusion of fetal vasculature , including that of renal tissue . the first category is likely related to women exposed to a pharmaceutical toxic effect during pregnancy and this effect may influence the expression of genes related to the renin side effects related to hypoperfusion of fetal vasculature ( or ace inhibitors arbs fetopathy ) include renal insufficiency , tubular dysplasia , fetal anuria with oligohydramnios , hypocalvaria , pulmonary hypoplasia and intrauterine death . since the discovery of these agents in 1965 , many studies have reported the effects of ace inhibitors on the fetus and maternal exposure , especially during the second and third trimesters of pregnancy [ 47 ] . subsequently , arbs were manufactured ; these powerful antihypertensive drugs are highly selective and specific to the renin angiotensin system and were developed with the aim of having less adverse effects on the fetus . however , a number of side effects similar to those described in fetuses and newborns of women treated during pregnancy with ace inhibitors have been reported . the present study describes a pregnant woman who was hypertensive for the last 3 years and treated during the entire pregnancy period with olmesartan medoxomil , the seventh of manufactured arb agents . following severe oligohydramnios , diagnosed at 33 gestational weeks , she underwent a caesarean section and gave birth to a female baby . in the first 48 postnatal hours the mother was a 40-year - old primigravida with a 3-year history of essential hypertension . she was treated with olmesartan medoxomil ( olmetec munich , germany ) , a single daily dose of 20 mg , and her hypertension was well controlled with this regimen . her conception was assisted by ivf . at 33 weeks of gestation , a severe degree of oligohydramnios was diagnosed by ultrasonographic evaluation . a premature female baby was delivered , with a weight of 1.980 g , length 45 cm and occipital frontal circumference 30.5 cm . shortly after the birth , the newborn presented respiratory distress syndrome , and was intubated and ventilated with oxygen supplementation ; the newborn received two doses of sulfactant . oligoanuria was also observed on the first and second postnatal days . on the third day post - birth , renal ultrasonography revealed two kidneys of normal size with increased echogenicity , while on the fifth postnatal day , the plasma creatinine level was 3.1 mg / dl . the neonate was transferred from the maternity hospital to the neonatal intensive care unit at the children 's hospital for further assessment and management . on admission , the newborn suffered from breathing difficulty and acute renal failure and was on ncpap 21% o2 . on several occasions , dolichocephaly with cranial and facial asymmetries , light dysplasia of the external ear , equestrian figures of the feet , wide anterior fontanel and dilatation of celebral ventricles were also detected . all of these facial characteristics were likely closely related to the severe degree of oligohydramnios . on brain ultrasonography , ihv of first degree on both sides and increased periventricular echogenicity were recorded , as well as a small asymmetry of lateralis ventricle of brain l > r . repeated renal ultrasonography confirmed the previously reported findings of increased echogenicity on two otherwise normal kidneys . muscular tone and neonatal reflexes were normal for the age of gestation , but wide anterior fontanel and sutures were detected . laboratory findings on admission were as follows : hb 11.5 g / dl , ht 35% , wbc 13.3 10/l ( pmn46% , l32% , m13% ) , plt 223 10 l , glucose 80 mg / dl , urea 201 mg / dl , creatinine 2.8 mg / dl , uric acid 12.9 mg / dl , urine spot for fena 13% , sgot 16 u / l , sgpt 17 u / l , -gt 56 u / l , alp 114 u / l , ca 9.7 mg / dl , p 7.4 mg / dl , mg 2.8 mg / dl , k 3.9 mmol / l , na 134 mmol / l , cl 88 mmol / l , ua 12.6 mg / dl , total bilirubin 3.1 mg / dl , direct 0.93 mg / dl , cpk 64 iu / l , ldh 342 iu / l , total protein 4.4 g / dl , albumin 2.8 g / dl , crp 0.43 mg / l , plasma aldosterone 872 ng / dl ( normal 19141 ng / dl ) and renin 507 pg / ml ( normal 40 220 pg / ml ) . during hospitalization , several challenges cpap was continued for 3 days post - admission followed by diffuse o2 for the next 24 h. following a conservative approach , meticulous fluid and electrolyte adjustment resulted in a gradual improvement of renal function so that by day 19 after birth , the plasma creatinine level was 0.9% . plasma creatinine levels during the first 2 weeks of postnatal period and at 1 year of life are displayed in figure 1 . the acute renal failure event and severe degree of oligohydramnios were both attributed to the mother 's antihypertensive therapy and are related to the temporarily reduced fetal gfr . one month post - admission , the baby was discharged from the unit in good condition with urea 25 mg / dl , plasma creatinine 0.8 mg / dl and renal trasonographic findings compatible with normal sized kidneys and normal parenchyma echogenicity . at the latest follow - up examination , the baby ( 1 year old ) was well developed and normotensive ( bp 105/60 mmhg ) . renal function tests showed 35 mg / dl and 0.5 mg / dl plasma urea and creatinine levels , respectively . the renin angiotensin system is very active during fetal organogenesis and plays an important role , particularly in renal development . as early as 1980 , well - documented animal studies showed that treatment with ace inhibitors during pregnancy causes fetal anomalies or fetal loss due to severe hypoperfusion . one year later , the first case report on a human fetus exposed to ace inhibitors was published , and many studies have been reported since then . in 1992 , the food and drug administration ( fda ) strongly emphasized a warning , especially to the obstetrical community , concerning the use of these agents during pregnancy . in 1995 , it was estimated that 50% of neonates born to mothers exposed to ace inhibitors would be affected by these treatments , while another 25% of the births would be fatal . arbs were later developed as an alternative treatment , aimed to have less adverse effects on the fetus . both ace inhibitors and arbs , however , are very powerful antihypertensive agents with a similar clinical outcome , and most of the effects in the maternal kidneys have been determined to be due to the dilatation of the efferent arterioles of the glomerulus . it results in hypoperfusion and a decrease in the glomerular filtration rate , which likely affects the development of the fetus . moreover , these pharmaceutics are excreted by the kidneys and can cross human placenta to be excreted into the amniotic fluid and presumably be swallowed by the fetus . therefore , fetal hypoperfusion and the consequent anomalies involving the kidneys are not only indirectly related to the mother 's hypotensive status , but also directly related to the drugs in the fetal circulation . additionally , it is possible that exposure to a pharmaceutical toxic agent could also influence the expression of renin angiotensin system genes . a number of reports have shown that administration of arbs in early pregnancy , or at least during the first trimester , is related to 6% of the major abnormalities [ 1416 ] . to our knowledge , 11 reports have been published describing side effects in the fetuses of mothers exposed to arbs during their second and third trimesters of pregnancy . nearly all pregnant women treated by these agents gave birth to babies either suffering from fetal abnormalities or born dead . it is of interest that valsartan and losartan were used in four and three of the reported cases , respectively , and all cases of losartan treatments died [ 3,4,9,1719 ] . however , another study reported that three normal babies were born to mothers treated by arbs during their first trimester of pregnancy . the most recently developed arb is olmesartan olmetic , and its use during pregnancy , at a dose of 10 mg / day , has only been reported once . according to the authors of this previous study , following oligohydramnios diagnosis at 29 gestation weeks , olmesartan olmetic regimen was withdrawn and rehydration along with a single dose of furosemide was administered to the pregnant woman . the result of this early intrauterine intervention was a healthy baby , born without renal or other abnormalities . it was therefore postulated that fetus renal dysfunction , following olmesartan olmetic maternal exposure , is probably reversible . our reported findings are , to some extent , consistent with this paper . in our case , the affected newborn was also exposed during his entire fetal life to olmesartan medoxomil at a dose of 20 mg / day . despite oligohydramnios diagnosis apart from various characteristics , mainly craniofacial directly related to oligohydramnios , the neonate suffered from oligoanuric type acute renal failure during the first weeks of life due to olmesartan medoxomil exposure . renal ultrasonographic findings of increased echogenicity on two otherwise normal kidneys are consistent with acute renal damage . the baby responded very well to the conservative management , and within first 2 weeks of life , recovery of renal function was observed . thus , one possibility is that fetal renal toxicity following maternal exposure to olmesartan medoxomil is dose related . another possible explanation is that the adverse reaction of arbs may be due to genetic polymorphisms in drug - inhibiting enzymes or receptor blockers , which would alter drug concentration , thus affecting drug response . in summary , here we reported a case of a premature female baby born with oligoanuric type of acute renal failure following the mother 's exposure to olmesartan medoxomil . we observed a gradual recovery of renal function during the first 2 weeks of life , and therefore , considered that renal impairment was transient . at the latest follow - up 1 year later , renal function was normal , arguing against the presence of irreversible renal damage . whether this specific antihypertensive agent , olmesartan medoxomil , administered during pregnancy at the dose of 20 mg daily is generally safe with no side effects for the fetus is not yet clear . Output:	angiotensin - converting enzyme ( ace ) inhibitors and angiotensin ii ( at ii ) receptor blockers ( arbs ) are widely used antihypertensives with well - recognized renoprotective and cardioprotective effects . although treatment with these agents generally does not result in adverse metabolic consequences , their use during human pregnancy has been associated with negative reactions . here we report a premature baby with a history of oligohydramnios and maternal exposure to the arb olmesartan medoxomil who was transferred to our institution with acute renal failure . conservative treatment with diuretics and meticulous management of fluids and electrolytes resulted in an improvement in renal function in the patient . we conclude that olmesartan medoxomil may cause reversible renal failure in premature neonates .
PubmedSumm2004	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: numerical and structural centrosome abnormalities are hallmarks of various cancers and have been implicated in the generation of multipolar mitoses and chromosomal instability . centrosomes function as major microtubule organizing centers in animal and human cells and thus contribute importantly to proper spindle formation and function during mitosis . normally , the single centrosome of a g1 cell duplicates precisely once prior to mitosis in a process that is intimately linked to the cell division cycle . accurate control of centrosome duplication is critical for symmetric mitotic spindle formation and thereby contributes to the maintenance of genome integrity . centrosome alterations in tumor cells are believed to lead to multipolar mitosis , asymmetric chromosome segregation and aneuploidy . for most cancers , however , the association between centrosome aberrations and malignant progression is merely based on correlation and a causative role of centrosome abnormalities in establishing and maintaining numerical chromosome instability as defined by dynamic changes in chromosome copy numbers has not been universally established . here , we studied centrosome abnormalities and chromosomal instability in an abdominal lymph node biopsy sample from a 19-years - old male patient with recurrent burkitt 's lymphoma ( the use of patient material in this study was approved by harvard medical school office of research subject protection ; hms irb docket # e012302 - 1 ) . routine hematoxylin & eosin ( h&e ) staining of formalin - fixed , paraffin - embedded tissue revealed the histomorphology of malignant burkitt 's lymphoma ( fig . burkitt 's lymphoma is a highly aggressive b cell neoplasia with a characteristic t(8;14 ) chromosomal translocation involving the c - myc locus on chromosome 8 and typically the immunoglobulin heavy chain locus on chromosome 14 . gtg - banded metaphase cells were obtained from unstimulated 24 hour cultures and the t(8;14 ) chromosomal translocation was detected in all fifteen metaphases analyzed ( fig . ( a ) h&e staining of a lymph node biopsy specimen from a patient with burkitt 's lymphoma . arrowhead indicates loss of one copy of chromosome 17 ( see text for details ) . slides were then boiled in 10 mm citrate buffer ( ph 6.0 ) for 30 min in a microwave oven followed by digestion with digest - all-3 pepsin solution ( zymed ) for 5 min at 37c . slides were blocked with 10% normal donkey serum ( jackson immunoresearch ) for 15 min at room temperature followed by incubation with a mouse monoclonal anti--tubulin antibody ( gtu-88 ; sigma ) at a 1:1000 dilution overnight at 4c . after washing in pbs , sections were incubated with a donkey anti - mouse rhodamine red conjugated secondary antibody ( jackson immunoresearch ) at a 1:100 dilution in pbs for 2 hours at 37c . cells were finally washed in pbs , counterstained with dapi ( vector ) , and analyzed by epifluorescence microscopy . since normal centrosome duplication gives rise to two centrosomes per cell , cells with more than two -tubulin dots were considered abnormal . in addition , structural centrosome abnormalities as characterized by deviations from the typical dot - like staining pattern were assessed . numerical and/or structural centrosome abnormalities were detected in approximately 30% to 50% of cells ( fig . accurate detection of mitotic spindle poles , however , was obscured due to tissue sectioning which precluded a quantitative evaluation of centrosome - associated mitotic abnormalities . quantitation of more than one - hundred mitoses revealed a normal configuration in 95% of cells ( 76% normal metaphases and 19% normal anaphases fig . arrowheads indicate abnormal centrosomes ( arrowheads and inserts correspond counterclockwise ) . both numerical abnormalities ( top right insert ) and structural abnormalities with excessive -tubulin positive material are found ( top left and lower left inserts ) . ( d ) quantitation of lymphoma cells undergoing metaphase or anaphase with a normal polarity ( open bars ) as opposed to an abnormal polarity ( black bars ) detected in h&e stained sections . ( e ) fluorescence in situ hybridization ( fish ) analysis for chromosome 11 confirms diploidy of the malignant lymphoma . whether the observed centrosome abnormalities were associated with ongoing numerical chromosome instability , chromosome 11 copy numbers were analyzed by fluorescence in situ hybridization ( fish ) . chromosome 11 copy number changes are among the five most frequently found numerical chromosome abnormalities in burkitt 's lymphoma karyotypes retrieved from the cgap database . for interphase fish , a spectrum green - labeled chromosome 11 -satellite probe ( d11z1 ) was used according to the manufacturer 's protocol ( vysis ) . none out of more than one - thousand cells analyzed showed more than two copies of chromosome 11 ( fig . consistent with this observation , cytogenetic analysis revealed a single predominant pseudo - diploid karyotype : 46 , xy , t(8;14)(q24;q32),-17 , der(19)t(7;19)(q11.2;q13 ) , + mar ( fig . this karyotype is in line with previous studies that showed no evidence for gross aneuploidy in most burkitt 's lymphomas . the absence of ongoing numerical chromosome instability was not due to impaired proliferative activity as evidenced by the high mitotic index ( fig . the tumor suppressor protein p53 is inactivated in up to 40% of burkitt 's lymphoma and loss of one copy of chromosome 17 , which harbors the p53 gene , is found in the case presented here . in order to test whether the remaining p53 allele might be mutated and since most mutant p53 proteins are stabilized , we performed p53 immunofluorescence staining of the burkitt 's lymphoma in comparison to normal lymph node tissue ( fig . tissue sections were processed as described above and incubated with a mouse monoclonal anti - p53 antibody ( do-7 , novocastra ) at a 1:100 dilution overnight at 4c followed by a rhodamine red - conjugated secondary donkey anti - mouse antibody at a 1:100 dilution for 2 hours at 37c . in contrast to normal tissue , immunofluorescence analysis of the burkitt 's lymphoma revealed strong p53 positivity suggesting that the remaining p53 allele has undergone mutation ( fig . expression of p53 was assessed by immunofluorescence staining ( do-7 monoclonal antibody ) using sections from a normal lymph node ( upper panels ) as negative control . , our results show that centrosome abnormalities do not necessarily correlate with ongoing numerical chromosome instability and that even the presence of multipolar metaphases is not sufficient to predict whether a cell or a tumor is chromosomally unstable . this indicates that numerical chromosome stability can be maintained in a tumor despite high - level centrosome abnormalities . in conclusion , the impact of centrosome abnormalities on genomic destabilization and malignant progression needs to be analyzed in the context of the cellular background in which these alterations occur . the question why a tumor cell with centrosome abnormalities does not necessarily acquire numerical chromosome imbalances may have several answers . in general , two models that lead to centrosome abnormalities with distinct functional consequences can be envisioned . according to the first model , numerical centrosome abnormalities can arise as a consequence of abortive mitotic events and impaired cytokinesis [ 14 - 16 ] . under such conditions , centrosome abnormalities are unlikely to cause expansion of a population of genomically unstable tumor cells because such cells are unable to complete mitosis and to give rise to viable progeny . in striking contrast , studies in breast cancer and primary human cell populations expressing the human papillomavirus type 16 ( hpv-16 ) e7 oncoprotein provided evidence that abnormal centrosome duplication can precede genomic destabilization and arise in morphologically normal cells with functional p53 . such cells have no inherent cell division defects , and thus , they are more likely to undergo abnormal mitosis even though additional checkpoint mechanisms may need to be subverted in order to give rise to chromosomally unstable daughter cells . in support of the notion that abnormal centrosome numbers and multipolar mitoses do not necessarily cause genomic instability , it has been shown that in hpv e7-expressing cells there are dramatic quantitative differences between multipolar metaphase cells and cells with a multipolar ana- or telophase configuration . a similar finding is presented here where multipolar anaphases were undetectable despite approximately 5% of multipolar metaphase cells ( fig . the molecular basis for a hypothetical checkpoint that halts mitotic progression in the presence of multiple spindle poles has not yet been established . although p53 has been implicated in a post - mitotic checkpoint following spindle disruption , results presented here and reports by others suggest that checkpoint mechanisms other than p53 are involved in suppressing aneuploidy . this does not rule out that p53 inactivation contributes indirectly to genomic destabilization by interfering with apoptotic cell death and thus allowing chromosomally unstable cells to remain in the proliferative pool . another mechanism that may limit chromosome missegregation in the presence of multiple spindle poles involves coalescence of centrosomes whereby a bipolar spindle is formed by clusters of multiple centrosomes . overexpression of c - myc in vitro has been shown to result in structural chromosomal aberrations and dna damage . structural chromosome abnormalities in addition to the t(8;14 ) translocation are frequently detected in burkitt 's lymphoma including the case presented here . these findings , together with the observed loss of one copy of chromosome 17 , may indicate that chromosomal instability might have been present at earlier stages of lymphomagenesis . since overexpression of c - myc can cause a g2/m arrest and polyploidization , accumulation of centrosomes in cells with impaired mitotic progression likely accounts for the majority of cells with centrosome abnormalities . whether the c - myc oncogene can directly contribute to the formation of multiple centrosomes in otherwise normal cells as it has been shown for the hpv-16 e7 oncoprotein awaits further clarification our results show that numerical and/or structural centrosome abnormalities are not a universal surrogate marker for chromosomal instability . further studies are warranted to investigate whether other cancers show a similar lack of correlation between centrosome abnormalities and numerical chromosome instability . moreover , our study suggests that cellular mechanisms other than p53 inactivation may interfere with progression through mitosis in the presence of centrosome - related cell division defects . further investigations addressing these problems will help to lead to a more definitive model of the role of centrosome abnormalities in cancer development and progression . dapi , 4',6'-diamidino-2-phenylindole ; edta , ethylenediaminetetraacetate ; gtg , g - bands obtained with trypsin and giemsa ; h&e , hematoxylin & eosin ; pbs , phosphate - buffered saline . author 1 ( s. d. ) coordinated the study and performed immunofluorescence analysis of centrosomes and fish . author 2 ( b. h. l. ) participated in the histological diagnosis of the lymphoma and also in the collection of tumor material and clinical information , and provided the hematoxylin & eosin image . author 3 ( p. d. c. ) performed the cytogenetic analysis and diagnosis of the malignant lymphoma and provided the image of the karyotype . author 4 ( k. m. ) participated in the design and interpretation of the study . supported by a research grant from astrazeneca to k. m. s. d. was supported by a research fellowship of the deutsche forschungsgemeinschaft . Output:	numerical and structural centrosome abnormalities are detected in various human malignancies and have been implicated in the formation of multipolar mitoses , chromosome missegregation , and chromosomal instability . despite this association between centrosome abnormalities and cancerous growth , a causative role of centrosome aberrations in generating chromosomal instability and aneuploidy has not been universally established . we report here excessive numerical and structural centrosome abnormalities in a malignant burkitt 's lymphoma harboring the characteristic t(8;14 ) chromosomal translocation . using conventional karyotyping and fluorescence in situ hybridization ( fish ) , we detected no signs of ongoing numerical chromosome instability , although the tumor displayed sporadic multipolar metaphases . these findings demonstrate that centrosome abnormalities are not a universal surrogate marker for chromosomal instability in malignant tumors . moreover , our results suggest a model in which additional cellular alterations may be required to promote centrosome - related mitotic defects in tumor cells .
PubmedSumm2005	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: today , growing public concern regarding caloric and fat intake has raised the demand for low- or reduced - fat foods [ 1 , 2 ] . yogurt , a traditional dairy product produced by bacterial fermentation of milk with a starter culture containing streptococcus salivarius ssp . bulgaricus , has been very popular for its reduced fat content , special texture , flavor , and tang , as well as its nutritional and health benefits beyond those of milk [ 35 ] . in china , the yogurt market constitutes a segment with great potential for expansion . in 2003 , the consumption of yogurt and its related drinks was estimated to account for 11% of the total dairy production of china . although yogurt and its related products are favored by many people in china , various adulterations in milk and yogurt products have aroused great public concern about the quality and safety of yogurt . since 2007 , a series of scandals involving the adulteration and contamination of dairy products with melamine and other compounds such as cyanuric acid , ammeline , and ammelide have brought serious challenges to chinese food quality supervision departments [ 712 ] . to deal with the crisis , numerous targeted analysis methods were developed to detect melamine and its analogues in dairy products , including liquid chromatography [ 13 , 14 ] , gas chromatography , laser raman spectrometry , immunoassay , electrochemical analysis , flow injection analysis , and infrared spectroscopy [ 2022 ] among others [ 2325 ] . however , new reports on adulterations with illegal ingredients other than melamine and its analogues are popping up from time to time . a recent hot issue is the yogurt adulterations with nonmilk proteins , including vegetable protein powder , edible gelatin , and even industrial gelatin . obviously , if we just rely on targeted analysis methods , the adulterations would be out of control and the analysis would be trapped in a cycle of adulteration , targeted analysis , and new adulterations , and so on . therefore , untargeted detection methods are required to enable the screening of dairy products for a range of known and unknown adulterants . untargeted detection of adulterations is a typical problem of chemometrics class modelling techniques [ 28 , 29 ] , which aim to answer the question of whether a new sample should be accepted or rejected by a sought - for class ( e.g. , pure and authentic yogurt ) . a class model proceeds as follows : ( 1 ) representative objects of the target class are collected and their characteristic signals are measured , ( 2 ) a class model is trained based on the measured signals to describe the distribution of authentic objects , and ( 3 ) a new object is analyzed and predicted by the class model . firstly , to ensure the sensitivity of the class model , a training set of the representative authentic objects should be collected to include most if not all of the important variations of authentic products . second , the specificity of the class model should be validated by predicting future objects . as a rapid analysis method , near - infrared ( nir ) spectroscopy has been widely used in food quality control for some advantages over traditional chemical analysis methods [ 3134 ] : ( 1 ) no or limited sample preparation , ( 2 ) less analysis time and cost , ( 3 ) the potential for nondestructive and online analysis , and ( 4 ) the ability to simultaneously characterize multiple chemical components . therefore , the objective of this paper was to develop a rapid method for protein adulteration identification of chinese yogurt by diffuse nir spectroscopy and chemometrics class modelling techniques . a recently proposed one - class partial least squares model [ 35 , 36 ] was used to model authentic yogurt samples . fresh milk samples were collected from local dairies and were heated to about 80c to kill any undesirable bacteria . the milk was then cooled to 40c and a starter bacteria culture was added , and the temperature was maintained for 12 hours to allow fermentation in a fermentor . the prescribed starter culture ( hecan , changzhou , china ) includes streptococcus salivarius ssp . the fermentation was terminated by keeping the yogurt samples at 4c and adulterating , and nir measurements were performed in 12 hours . adulterated yogurt samples were prepared by blending the previous pure samples randomly with different levels of adulterant solutions , including edible gelatin ( pucheng , hangzhou , china ) , industrial gelatin ( hengtong , foshan , china ) , and soy protein powder ( jichuan , hangzhou , china ) . the thicknesses of pure and adulterated yogurt objects were kept to be approximately equal by adding pure water , which is the common practice of protein adulteration . the information concerning the 60 pure and 197 adulterated yogurt objects is listed in table 1 . the nir diffuse reflectance spectra of pure and adulterated yogurt samples were measured in the spectral range from 4000 to 12000 cm on a bruker tensor37 ftir spectrometer ( bruker optics , ettlingen , germany ) using opus software . all spectra were measured in a quartz cup with a pbs detector and an internal gold background as the reference . sixty - four scans were performed for each object and more scans did not improve the signal quality significantly . the spectrum of pure water was obtained by averaging the five repeated measurements of water membranes on the internal gold background . to ensure the specificity of class models to detect potential adulterants , unnecessary variations in the target - class samples should be removed or reduced . because water has strong absorbance in nir range , the water variations in milk and during production , transportation , storage , and processing of yogurt can cause great signal variations in pure yogurt samples , which would make a class model wrongly accept more adulterated yogurt objects . therefore , in order to remove the influence of water variations , the spectra of both training and prediction objects were orthogonally projected ( op ) onto the complement space of water spectrum as follows : ( 1)xnew=(iss+)xraw , where xraw and xnew are the vectors of raw and preprocessed spectra of an object and s is the spectrum of pure water , respectively . standard normal variate was originally designed to reduce scattering effects in the spectra but was also proved to be effective in correcting the interference caused by variations and reducing spectral backgrounds . therefore , snv transformation was also used as a preprocessing option . representative training and prediction sets are required to train and validate a class model ; in this paper , the duplex algorithm was used to divide the analyzed objects into a training set and a test set . duplex algorithm proceeds as follows : ( 1 ) the two samples with largest euclidean distance were selected and put in the training set , ( 2 ) the two objects with largest distance among the remaining samples were put in the test set , and ( 3 ) steps ( 1 ) and ( 2 ) are repeated until one has obtained as many test objects as predefined . by alternatively selecting the furthest samples for the training set and test set , duplex can obtain two data sets almost with equal distributions in the experimental space . ocpls balances the explained variances and compactness of the target class by projecting each training object onto the class average . with a training matrix x ( n by p ) including p features of n objects from the target class , ocpls works in the framework of partial least squares ( pls ) regression as follows : ( 2)1=xbpls+e , where each element of the response vector 1 ( n by 1 ) is 1 , bpls ( p by 1 ) is the pls regression coefficient vector , and e ( n by 1 ) denotes regression errors . it should be noted that the training data x should not be column centered ; otherwise , all the predictors would be orthogonal to the response vector 1 . the model errors e can be assumed to have a normal distribution and its standard deviation can be seen as a measure of the target - class dispersion . the standard deviation of e can be estimated as the prediction errors of monte carlo cross - validation ( mccv ) . given a significance level , , the 1- confidence interval of the predicted response value ( yun ) for accepting a new sample can be estimated as ( 3)(1^ez1/2^,1^e+z1/2^ ) , where z1/2 is the critical value of standard normal distribution , ^e is the mean of e , and ^ is the standard deviation of model error estimated by mccv . to evaluate the performance of class models , sensitivity ( sens ) and specificity ( spec ) of prediction were used as follows : ( 4)sens = tptp+fn , spec = tntn+fp , where tp , fn , tn , and fp denote the numbers of true positives , false negatives , true negatives , and false positives , respectively . the raw nir spectra of pure yogurt and water are shown in figure 1 . figure 2 demonstrates the raw spectra of yogurt objects adulterated with different levels of edible gelatin , industrial gelatin , and soy protein powder . in each subplot of figure 2 , a shift was added to differentiate doping levels . as demonstrated in figures 1 and 2 , the spectra of pure water and pure and adulterated yogurt samples have almost same absorbance bands in the range of 400012000 cm , which can be attributed to the high contents and the strong absorbance of water . because of peak overlapping , the raw spectra have a very poor resolution , and the spectra variations caused by other chemical components were seriously masked by the absorbance bands of water . therefore , proper spectra preprocessing is required to remove the unnecessary spectral variations due to water and highlight the spectral variations caused by adulterants . all the raw spectra were also projected onto the orthogonal complement space of water spectrum . then , the duplex algorithm was used to split the 60 pure yogurt samples into a training set of 40 samples and a test set of 20 samples . all the 197 adulterated yogurt samples were used as negative test samples to test the specificity of the class model . therefore , the training set had 40 positive ( pure ) yogurt objects and the test set included 20 positive and 197 negative ( adulterated ) objects . based on the raw , snv - transformed , and op spectra , ocpls models were developed and the model performance was evaluated by sensitivity and specificity obtained from the prediction of the test objects . mccv with 10% left - out samples was used to determine the number of latent variables and the repeat number was 100 . the predicted residual sum of squares ( press ) values by mccv were examined and the number of components was determined to obtain a low press value , and more components can not reduce the press value significantly . as seen from table 2 , both snv and op spectra preprocessing can improve the specificity of class models by reducing the unwanted variations caused by water absorbance and backgrounds . the best model was based on op spectra with sensitivity of 0.900 and specificity of 0.949 , respectively . the prediction results obtained with op spectra were demonstrated in figure 5 . in figure 5 , the adulterated yogurt samples were arranged according to an ascending doping level . with both op- and snv - preprocessed spectra , the wrongly accepted samples ( false positives ) had a doping concentration of 0.5% for edible gelatin , 1% for industrial gelatin , and 1% for soy protein powder , respectively . moreover , the distance of an adulterated object to the critical value increases with doping level , indicating that the identification of higher doping levels , namely , 1% edible gelatin , 2% industrial gelatin , and 2% soy protein powder , can be safely detected . nir spectroscopy combined with a recently proposed chemometrics class modelling method , ocpls , has shown much potential for developing untargeted detection for protein adulteration in chinese yogurt samples . in the current experimental conditions , adulterations with edible gelatin ( 1% ) , industrial gelatin ( 2% ) , and soy protein powder ( 2% ) can be detected by ocpls with proper preprocessing of spectra . the results demonstrate that orthogonally projection of raw spectra onto the spectra of pure water is useful to remove unwanted variations . if the within - class variations are controlled effectively , the model specificity can be improved and lower levels of adulterants can be detected . Output:	untargeted detection of protein adulteration in chinese yogurt was performed using near - infrared ( nir ) spectroscopy and chemometrics class modelling techniques . sixty yogurt samples were prepared with pure and fresh milk from local market , and 197 adulterated yogurt samples were prepared by blending the pure yogurt objects with different levels of edible gelatin , industrial gelatin , and soy protein powder , which have been frequently used for yogurt adulteration . a recently proposed one - class partial least squares ( ocpls ) model was used to model the nir spectra of pure yogurt objects and analyze those of future objects . to improve the raw spectra , orthogonal projection ( op ) of raw spectra onto the spectrum of pure water and standard normal variate ( snv ) transformation were used to remove unwanted spectral variations . the best model was obtained with op preprocessing with sensitivity of 0.900 and specificity of 0.949 . moreover , adulterations of yogurt with 1% ( w / w ) edible gelatin , 2% ( w / w ) industrial gelatin , and 2% ( w / w ) soy protein powder can be safely detected by the proposed method . this study demonstrates the potential of combining nir spectroscopy and ocpls as an untargeted detection tool for protein adulteration in yogurt .
PubmedSumm2006	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: drug - induced liver injury ( dili ) is defined as a liver injury due to xenobiotics , herbs , or medications that leads to either liver dysfunction or abnormal liver serology , in the setting of no other identifiable cause . unlike idiosyncratic drug induced liver injury ( idili ) , dili seems to be dose - dependent , predictable , and reproducible . dili has been associated with more than 1,000 medications and is the most common cause of acute liver failure in the united states , accounting for approximately 50% of fulminant cases.15,6 while retrospective studies have shown acetaminophen to be the most common cause of dili , studies in liver transplant patients have demonstrated antibiotics and immunosuppressive agents , such as tacrolimus and azathioprine , as the top causes.7,8 bjornsson et al . performed a prospective study and found that amoxicillin - clavulanate was the most commonly implicated antibiotic . the incidence was approximately 19 cases per 100,000.9 in addition to the typical suspects stated above , herbal and dietary supplements have been a rising cause of dili , now accounting for up to 9% of all cases.10 in fact , one study based in china , which spanned eight years , found that nearly half of all cases were secondary to medicinal herbs.11 while dili is most commonly due to only one drug , a prospective study demonstrated that nearly 10% of cases were attributable to more than one medication.9 dili has several clinical manifestations ranging from asymptomatic elevations in liver enzymes to fulminant liver failure . hepatic , or cytochrome p450 , metabolism of a drug has been strongly linked to dili , and one study demonstrated that those medications that rely on more than 50% hepatic metabolism have a greater risk for alanine transferase ( alt ) elevation , liver failure , and mortality.12 other proposed mechanisms for dili include autoimmunity and hypersensitivity.2,13 despite the possible mechanisms , treatment options all follow the same principles . the most important , and first , step is early diagnosis followed by discontinuation of the offending agent.2 the most common presenting symptom in most cases of dili is jaundice , although many cases are asymptomatic or have elevated enzymes incidentally detected.11 however , dili can present as cholestatic , hepatocellular , and mixed ; and proper diagnosis will direct the treatment of a particular case . cholestatic liver injury is defined by predominant elevations of alkaline phosphatase ( alp ) , whereas hepatocellular liver injury is defined as predominant elevations of aspartate transferase ( ast ) and alt , and mixed has elevations in both alp and transaminases.14 it is important to note that alt is liver - specific , whereas elevations in ast may be associated with damage to skeletal or cardiac muscle or in conditions such as myocardial infarction and rhabdomyolysis . hepatocellular injury remains the most common presentation and is correlated with a worse outcome.15,16 table 1 provides specific definitions for both cholestatic and hepatocellular injury , which may aid in the identification of the patient 's clinical diagnosis . once liver injury has been established , the next step is to determine the underlying cause . risk factors for dili have been difficult to determine with many studies because of conflicting results in regard to sex and age - related occurrences . however , a recent study found that older age and female sex was associated with a cholestatic type injury , while younger age and male sex was associated with hepatocellular type injury . genetic polymorphisms have been consistently found to be a risk factor for dili but are not typically tested for in patients prior to receiving a medication.17,18 finally , liver transplant patients where the reason for transplant was primary sclerosing cholangitis are at greater risk for developing dili.8 chronic liver disease is a risk factor but only for certain medications , including methotrexate , isoniazid , and hiv antiretroviral therapy.2 when diagnosing dili , several established criteria can be used , none of which is considered to be a gold standard . hy 's law is specific but not sensitive , whereas the roussel uclaf causality assessment method ( rucam ) is more sensitive but more difficult to administer in its truest form as it is complicated and requires rechallenging patients once they have recovered.1921 one modification is the digestive disease week - japan ( ddw - j ) scale , which includes lymphocyte stimulation testing , although this test has not been validated.22 yet another modification is the clinical diagnostic scale ( cds ) or maria and victorino ( m&v ) scale , which is more simple to administer but less predictive in patients who have had a prolonged period of time between drug use and development of symptoms or in those patients who have developed chronic liver injury.23,24 when examining these scales , common features , which make them specific and sensitive tests , include temporal relationship , exclusion of other causes , and prior reports of hepatoxicity of the suspected medication . while specific criteria may be used , at a minimum the above three commonalities should be investigated . for reference , wang et al found in china that the majority of cases occurred between 5 and 90 days of the initiation of the drug.11 as an aid for the clinician , table 2 provides medications and their typical liver injury presentation . once a specific drug has been identified as the cause of dili , it must be discontinued.2 following withdrawal of the suspected agent , therapy is largely supportive with a few notable exceptions for acetaminophen and amanita mushroom poisoning . all patients ' laboratory values , including ast , alt , alp , bilirubin , and international normalized ratio ( inr ) and mental statuses should be monitored for changes . it is important to note that measuring only ast and alt is insufficient when monitoring liver function in dili . damage to hepatocytes causes elevation of ast and alt levels , and once sufficient damage has occurred , there are less hepatocytes and ast and alt levels begin to decrease.14 therefore , bilirubin and inr must also be monitored . with discontinuation of the drug , most cases resolve without further sequelae , with one prospective study reporting a median duration from diagnosis to normalization of laboratory values of 64 days.9 lee et al . performed an eight year prospective , double - blind , placebo control trial of n - acetylcysteine ( nac ) for patients with acute liver failure not secondary to acetaminophen overdose . while overall survival was similar in the populations ( 70% vs. 66% ) , transplant - free survival was significantly better for those who received nac ( 40% vs. 27%).25 at this time , nac therapy can and should be considered for patients who are presenting with acute liver failure . in addition , a small uncontrolled study performed by wree et al . compared steroid pulse therapy with steroid step down therapy , both in combination with ursodeoxycholic acid , in the treatment of drug - induced liver injury , including patients with hepatocellular and/or cholestatic injury . both therapies demonstrated a decrease in time to normalization of bilirubin , ast , and alt values.26 however , since this was an uncontrolled study , it is uncertain if the observed improvement in their patient population actually reflected the natural history of dili . treatment for acetaminophen toxicity largely consists of nac therapy . studies have conflicted between whether oral or intravenous ( iv ) therapy should be given . yarema et al . compared iv to oral administration and found that hepatoxicity occurred less often in patients who received iv therapy within 12 hours of ingestion.27 however , a recent study performed using a simulation system demonstrated that for those patients presenting within 24 hours of acetaminophen ingestion , the oral protocol is superior to the 21-hour intravenous protocol in persevering hepatocytes . these authors argued that yarema 's study contained two differing treatment groups.28 regardless , nac therapy should be started if there is an elevation in ast , a detectable acetaminophen level , or if the level is above the treatment line on the rumack - matthew nomogram.29 a computer simulation suggested that international normalized ratio ( inr ) did not undertreat patients but did lead to overtreatment , whereas treating until alt peaked did not undertreat and rarely overtreated patients.28 current recommendations are to treat according to the protocol and to recheck ast and the acetaminophen level . at this point , ast should be less than 100 iu / l and acetaminophen level should be less than 10 mcg / ml . if not , therapy should be continued using the normalization of inr as a marker of resolution.30 amanita mushroom ingestion can lead to liver injury via the amatoxin , which inhibits rna polymerase ii and leads to hepatocyte necrosis . this is followed by symptomatic improvement but an elevation in ast and alt followed by the development of jaundice . since the hepatic phase is preceded by a gastrointestinal phase where dehydration and metabolic derangements may have developed , it is important to treat any dehydration and electrolyte abnormalities.31 while it has no proven efficacy for long - term survival , repeated activated charcoal administration is often recommended , which will prevent reabsorption of the amatoxin . while amatoxin may cause metabolic acidosis on its own , activated charcoal , which contains propylene glycol , can also cause a high anion gap metabolic acidosis . while clinicians should be aware of this potential complication , it should not preclude treatment with activated charcoal.32 silibinin is universally accepted as a treatment modality for amatoxin poisoning because it inhibits the transfer of amanitin into hepatocytes . it should be administered within 48 hours of mushroom ingestion . the current recommended dose is 20 to 50 mg / kg / day iv , which should be continued for 4896 hours . the recommended dose is 1,000,000 iu / kg for first day and 500,000 iu / kg for next two days via continuous iv administration . finally , if started early , the molecular adsorbent recirculating system may be considered as it does improve liver function , although no specific studies have been done with amanita mushroom poisoning.31 patients who present with fever , rash , and eosinophilia should be considered for a diagnosis of drug - induced autoimmune hepatitis . if the dili is severe , corticosteroid therapy should be considered as studies have demonstrated normalization of biochemical tests within six months.33 finally , patients who present with a cholestatic picture may complain of intense pruritus . treatment options for these patients include emollients , hydroxyzine , diphenhydramine , bile acid resins , and rifampicin.34 extracorporeal systems have progressed through advances in genetically produced cell lines , stem cell - derived functional hepatocytes , immortalized human hepatocytes as well as improved techniques and methods for preserving the hepatocytes . the system , composed of a hollow - fiber cartridge lined with porcine hepatocytes , was used in a multi - center trial showing survival benefit to those treated.35 advances in bioartificial livers continue with current trials being conducted by vital therapies and hepa wash gmbh ( clinicaltrials.gov ) . these devices provide the potential for significant benefit to patients with dili , subacute fulminate failure , and fulminate failure ; and we are encouraged for the use of this technology in the future treatment of these patients . the king 's college criteria have been developed for both paracetamol and non - paracetamol causes in order to determine when a patient should be initially referred for transplantation . the criteria include a prothrombin time ( pt ) over 100 seconds or at least three of the following : pt over 50 seconds , bilirubin > 300 micromol / liter , age below 10 or over 40 , an interval between jaundice and encephalopathy greater than seven days , or drug toxicity.37 other criteria that may be used include clinchy 's criteria and escudie 's criteria . patients with fulminant liver failure , defined as the presentation of hepatic encephalopathy within eight weeks of the development of symptoms related to liver disease , should be referred for transplant . in the united states , according to united network for organ sharing ( unos ) , a status 1a listing may be obtained if the patient has a life expectancy of seven days or less , does not have a pre - existing liver disease , and is in the intensive care unit requiring either ventilator assistance or dialysis or with an inr greater than two.38 however , it is important to keep in mind the contraindications for transplant , which include significant comorbidities and active malignancy . each patient should be considered on a case - by - case basis , and the guidelines should be consulted when considering a patient for transplant . although it has a varied presentation and multiple possible drug causes , treatment for all cases requires discontinuation of the offending agent . if a patient has ingested acetaminophen or amanita mushrooms , appropriate therapy should be administered . all patients can now be considered for nac therapy and should be monitored for normalization of biochemical tests . Output:	drug - induced liver injury ( dili ) has been linked to more than 1,000 medications and remains the most common cause of acute liver failure in the united states . here , we review the most current literature regarding treatment and make recommendations for the management of this relatively common disease . since treatment of dili remains largely elusive , recent studies have attempted to define new management strategies for these difficult patients . early diagnosis and withdrawal of the suspected medication is the mainstay of treatment of dili . for acetaminophen and amanita mushroom poisoning , there are specific therapies in use . finally , there are other possible management modalities for dili , including corticosteroids and ursodeoxycholic acid .
PubmedSumm2007	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: balanced general anesthesia comprises amnesia , analgesia , hypnosis , muscle relaxation , and obtundation of reflexes . to achieve all these components , we use different combinations of drugs . however muscle relaxants may confound our evaluation of depth of anesthesia due to lack of motor activity and may make the patient aware of the intraoperative events . bispectral index ( bis ) is a widely used quantitative parameter for evaluating depth of anesthesia and sedation . it is a continuous noninvasive electroencephalographic method that has been proposed to monitor the hypnotic state during sedation and anesthesia . the alpha-2 receptor agonist dexmedetomidine has potent sedative properties and has received food and drug administration approval for use in the intensive care unit ( icu ) for sedation . dexmedetomidine has analgesic sparing properties and is not associated with respiratory depression in therapeutic doses .. it has been used as an anesthetic sparing agent but not as a sole anesthetic agent . we have proposed to evaluate the effect of dexmedetomidine on depth of anesthesia and compared it with that of propofol by using hemodynamic variables and bis values , with the aim of studying the feasibility of dexmedetomidine as a sole anesthetic agent in maintaining depth of anesthesia . approval for the study was obtained from our institution 's ethics committee and written informed consent was obtained from the patients . sixty male and female patients , 18 - 65 years of age , asa physical status of i , scheduled for laparotomy under general anesthesia were considered for enrolment randomly into two groups ( 30 patients in each ) by a computer - generated randomization scheme . patients having heart block , hypertension , diabetes mellitus , renal dysfunction , psychiatric illness , or patients taking antipsychotic drugs or sedatives were excluded from the study . baseline hemodynamic variables were noted by attaching monitors [ noninvasive blood pressure , heart rate ( hr ) , oxygen saturation , end - tidal carbon dioxide , electrocardiogram , and temperature ] . group a patients received bolus dose of propofol ( 1 mg / kg for 10 min ) followed by an infusion of propofol ( 50 mcg / kg / min ) and group - b received bolus dose of dexmedetomidine ( 1 mcg / kg for 10 min ) followed by an infusion of dexmedetomidine ( 0.5 mcg / kg / h ) . after preoxygenation for 3 min , anesthesia was induced with intravenous ( i.v ) thiopentone sodium ( 3 - 5 mg / kg ) . sedation and anesthesia was maintained with infusion of study drugs ( propofol or dexmedetomidine ) . depth of anesthesia was evaluated by monitoring intraoperative hemodynamic values ( hr , blood pressure ) and bis value . sensors were placed as follows : one at the center of forehead approximately 2 inches above the bridge of nose ; second one directly above forehead , and third one on temple between the corner of the eye and hairline . parameters [ age , gender , weight , mean arterial pressure ( map ) , hr , bis value , sedation score ] were compared between the two groups by unpaired t test . all data with p these two groups were comparable in respect to demographic variables ( age , weight , gender , asa ps i , type of surgery , and duration of anesthesia ) [ table 1 and figure 1 ] . demographic data of two groups the two groups were also comparable with respect to their baseline map and hr . postintubation rise was less in group b. subsequent map and hr were also less in group b. but the difference between the two groups was statistically significant only for map at 20 min after infusion ( p = 0.039 ) [ figures 2 and 3 ] . trend of mean arterial pressure in two groups trend of heart rate in two groups baseline bis value was comparable between the two groups . bis value decreased after induction in both the groups but patients getting dexmedetomidine infusion showed statistically significant lower values of bis when compared with propofol ( p < 0.0001 ) [ table 2 and figure 4 ] . intraoperative bis value trend of bispectral index in the two groups sedation score was noted in the immediate postoperative period by ramsay sedation scale . it was seen that patients recieving dexmedetomidine were more sedated in the postoperative period , but it did not impair ventilation though the difference between the two groups was statistically significant ( p < 0.0001 ) [ table 3 and figure 5 ] . none of the patients in both the groups was able to recall any intraoperative events . general anesthesia has several components , namely , amnesia , hypnosis , analgesia , immobility , and blunting of autonomic reflexes . often in operating rooms anesthetic induction has been referred to as going to sleep . but all the components of general anesthesia may not be achieved as muscle relaxants may confound our evaluation of depth of anesthesia due to lack of motor activity and may make patients aware of the intraoperative events . for achieving adequate depth of anesthesia , use of inhalational or intravenous anesthetic agents in hypnotic doses however , it has been observed that certain anesthetic drugs in subhypnotic concentrations produce amnesia , ( eg , propofol , midazolam ) due to engrams ( physical alterations in neural tissues thought to be substrate for memory ) impaired by these drugs . in these cases of episodic amnesia , memory is lost before consolidation . in our study , we have similarly used subhypnotic equisedative doses of propofol and dexmedetomidine and observed episodic amnesia in these cases . similar dosing pattern of bolus and infusion were used by previous authors who used a higher loading dose of dexmedetomidine . in the current study , a minimum dose of both the drugs was chosen that ensured amnesia but caused least side effects such as hypotension , bradycardia , or arrhythmia alpha-2 agonist dexmedetomidine is a sedative and analgesic agent , which has been used for icu sedation upto 24 h after surgery . dexmedetomidine provides hemodynamic stability and appears to have no clinically important adverse effect on respiration . compared with propofol and midazolam , dexmedetomidine was as effective in maintaining adequate sedation for prolonged mechanical ventilation . dexmedetomidine has also been used for sedation for short surgical procedures and as an adjunct resulting in reduced intraoperative requirements of anesthetics . it has been used as sole anesthetic agent only in a few occasions , whereas propofol has been used to maintain the depth of anesthesia routinely . the results of our study validate the use of dexmedetomidine as a maintenance anesthetic agent . loading dose in this study has been reduced as maximum adverse effects were observed immediately after administration of the loading dose . target bis is delayed when loading dose is reduced but in this study target bis ( 40 - 65 ) was achieved with the addition of an induction agent . dexmedetomidine even in lower loading dose produced better amnesia for procedural sedation compared with remifentanil . ramsay and his co - workers reported the use of dexmedetomidine at a higher dose for intubation in difficult airway scenarios without any induction agent or neuromuscular blocker and observed hypotension , bradycardia , and upper airway obstruction . in the current study , along with study drug , induction agent and neuromuscular blockers were used to facilitate endotracheal intubation and much less dose could be used in contrast to the cases reported by ramsay . bis is a continuous noninvasive electroencephalographic method that has been proposed to monitor the hypnotic state during sedation and anesthesia . its use has been documented to be associated with lesser chance of intraoperative awareness . in a study on volunteers , yusuke and his co - workers compared bis value with a clinical scoring [ observer assessment of alertness sedation score ( oaa / s ) ] both with propofol and dexmedetomidine . they found that equivalent doses of dexmedetomidine produced lower bis value and the bis value of 46 with dexmedetomidine was comparable to bis value of 67 with propofol both of which produced oaa / s score 2 ( oaa / s 2 denotes adequate sedation ) . in the current study patients getting dexmedetomidine infusion showed statistically significant lower values of bis when compared with propofol ( bis value 60 - 65 in propofol as opposed to 50 - 55 with dexmedetomidine ) . despite the difference in bis equal depth was achieved with both the drugs with no recall in any patient as assessed by modified brice questionnaire . this may be explained by the different mechanisms of sedation by the two study drugs . dexmedetomidine produces sleep by hyperpolarization of noradrenergic locus ceruleus neurons as opposed to gaba agonism by propofol . a rise in map and hr was noted after intubation with subsequent fall of both in the two groups . postintubation rise was less in group b. subsequent map and hr were also less in group b. but the difference between the two groups was statistically significant only for map at 20 min of infusion ( p = 0.039 ) . earlier study has also shown that dexmedetomidine attenuates stress response to intubation by decreasing central sympathetic outflow . another study done in bariatric surgery found dexmedetomidine produces better control of hemodynamic parameters compared with fentanyl . sedation score was calculated in both the groups in immediate postoperative period by ramsay sedation scale . it was seen that patients receiving dexmedetomidine were more sedated during postoperative period ( p value < 0.0001 ) , but without any impairment of ventilation . it is consistent with the result of the study done by venn and grounds who also found more sedation with dexmedetomidine postoperatively but without any delay in extubation . the longer sedation with dexmedetomidine could be explained by longer elimination half - life of the drug . but incidence of delayed recovery and longer discharge time with dexmedetomidine were observed by other investigators . the observations of the present study could have been more conclusive with larger sample size , multicenter trial and measurement of plasma levels of drugs . from our study we can conclude that dexmedetomidine is comparable with propofol as maintenance anesthetic agent and it can produce better control of hemodynamic variables and bis value . so , dexmedetomidine may be used as sole anesthetic agent for maintenance of depth of anesthesia . Output:	background and aims : intravenous agents such as propofol are commonly used to maintain adequate depth of anesthesia . dexmedetomidine which has an anesthetic sparing effect is being considered for maintaining intraoperative depth of anesthesia . we hypothesized to compare the effect of dexmedetomidine on depth of anesthesia with propofol and evaluated whether dexmedetomidine can be used as sole anesthetic agent in maintaining depth of anesthesia.materials and methods : sixty patients of asa ps i , 18 - 65 years of age , scheduled for laparotomy under general anesthesia were randomly divided into two groups of 30 each . group a received propofol 1 mg / kg bolus followed by infusion ( 50 mcg / kg / min ) and group b received dexmedetomidine 1 mcg / kg bolus followed by infusion ( 0.5 mcg / kg / h ) . both the groups were administered standard general anesthesia with routine monitoring along with bispectral index ( bis ) and values were recorded at intervals of 10 min . in all patients ramsay sedation score was recorded after extubation and they were assessed for recall of intraoperative events using modified brice questionnaire.results:heart rate and mean arterial pressure were less in group b than group a. intraoperative bis values were significantly lower in group b ( p < 0.0001 ) . although sedation score was more in group b it did not prolong recovery . no recall was found in any patient.conclusion:dexmedetomidine was comparable with propofol in maintaining anesthesia and it can produce better control of hemodynamics and bis value . thus dexmedetomidine can be used as the sole maintenance anesthetic agent .
PubmedSumm2008	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: nephrotic syndrome ( ns ) is an illness consisting in leakage of proteins in urine , resulting in life threatening conditions due hypovolemia , hypercoagulation , and infection . the annual incidence of ns in children in the usa and in europe has been estimated to be 17 per 100,000 children , with a cumulative prevalence of 16 per 100,000 children [ 13 ] . nephrotic syndrome in children can be classified according to 3 three groups : secondary , congenital and infantile , and idiopathic . secondary nephrotic syndrome is defined as nephrotic syndrome associated with well - defined diseases that are inflammatory ( e.g. , lupus nephritis , acute postinfectious glomerulonephritis , iga nephropathy , henoch - schnlein purpura , etc . ) or not ( e.g. , alport syndrome , focal sclerosis due to reduced nephronic mass resulting from renal scarring , etc . ) . congenital and infantile nss are occurring before the age of one year and are mostly associated with infections ( e.g. , syphilis , toxoplasmosis , etc . ) or with mutations of genes coding for podocytes proteins and are steroid resistant . idiopathic nephrotic syndrome ( ins ) is the most frequent form of ns in children representing more than 90 percent of cases between 1 and 10 years of age and 50 percent after 10 years of age . ins is defined by the association of the clinical features of ns with renal biopsy findings of diffuse foot process effacement on electron microscopy and minimal changes ( called minimal change disease ( mcd ) ) , focal segmental glomerulosclerosis ( fsgs ) , or diffuse mesangial proliferation ( dmp ) on light microscopy . the vast majority of patients with mcd ( > 90 percent ) respond to glucocorticoid therapy whereas only 50 percent of those with dmp and 30 percent of those with fsgs are expected to do so . the latter include : age younger than six years of age , absence of hypertension , absence of hematuria , normal complement levels , and normal renal function . however , onset of nephrotic syndrome in the first year of life , particularly in the first three months of life , is more likely to be due to a gene mutation and to be resistant to glucocorticoids . it is therefore actually generally admitted that a course of glucocorticoids should be given without previous kidney biopsy when the illness has started after the age of one year whereas the upper age limit to do so is generally considered to be 10 years since only 10 percent of patients under 10 years old are steroid resistant in comparison with 20% for the totality of patients less than 18 . the ins pathophysiology has been attributed in the past mainly to structural abnormalities and a loss of anionic charges of the glomerular basal membrane ( gbm ) leading to proteinuria . actually the podocyte has become the favourite candidate for constituting the main part of the glomerular filtration barrier . the latter is highly specialized , terminally differentiated cells with cytoplasmic extensions , the so - called foot processes anchored on the gbm , forming the slit diaphragm ( sd ) which is essential in retaining proteins inside the lumen of capillary loop . genetic studies of hereditary forms of ns have led to the identification of proteins playing a crucial role in slit - diaphragm signalling , regulation of actin cytoskeleton dynamics , maintenance of podocyte integrity , and cell - matrix interactions . structural elements of the sd ( nephrin , podocin , and cd2ap ) and actin cytoskeleton ( a - actinin-4 ) control podocyte differentiation and survival , cell polarity , and cytoskeletal dynamics . podocyte and glomerular development are critically regulated by the transcription factor wt1 and phospholipase c 11 ( plc11 ) mediated signals . the calcium channel trpc6 , which localizes in membrane lipids supercomplex along podocin , regulates mechanosensation sensed at the sd , whereas the structural component of the gbm , laminin - b2 , is essential for podocyte cell - matrix interactions . podocyte integrity may also be affected by derangements in proteins involved in varied subcellular processes including the mitochondrial respiratory chain , dna restructuring and repair , and lysosomal function . finally , identification of novel genetic determinants in glomerular disease , such as high - risk haplotypes in the myh9 gene , may also explain the increased risk of some adult patients to glomerular injury . ins secondary to mutations of genes coding for podocytes proteins is typically steroid resistant . excellent reviews on this topic [ 69 ] including the following systematic approach for genetic testing by the group of antignac have been recently published . a mutation of nphs1 , encoding nephrin and responsible for the finnish - type congenital nephrotic syndrome , is found in most patients presenting with a nephrotic syndrome in the first 3 months of life . the second most frequent mutation in that age group concerns nphs2 , which encodes podocin and is particularly frequent in central europe . finally some patients less than 3 months of age present with a mutation of the wt1 ( also often associated with the denys - drash and the frasier syndromes characterized by gonads and genital abnormalities ) and/or the plce1 genes , particularly when the renal biopsy shows diffuse mesangial sclerosis . in the group of patients starting the illness between 4 and 12 months ( infantile nephrotic syndrome ) and later on ( childhood nephrotic syndrome ) , nphs2 followed by nphs1 is the first genes to be tested in nonsyndromic patients presenting srns associated with minimal glomerular changes / fsgs in the infantile or childhood period . in the remaining patients with the same histological lesions , genetic testing for wt1 mutations ( exons 8 and 9 in phenotypically female patients ) should be performed , while screening for plce1 mutations may be considered in some cases ( mainly in familial cases ) . mutations in the cd2ap , actn4 , trpc6 , and inf2 have also been found anecdotally in childhood srns ( and for review , [ 79 ] ) . a mutation of lamb2 , which encodes laminin beta 2 ( a component of the glomerular basement membrane ) , is responsible for pierson syndrome . other rare syndromal conditions are mitochondrial disorders ( gene na coding for nonprotein trna ) , nail - patella syndrome ( gene lmx1b coding for lim homeobox transcription factor 1 beta ) , schimke immunoosseous dysplasia ( smarcal1 coding for swi / snf2-related , matrix - associated , actin - dependent regulator of chromatin , subfamily a - like 1 ) , mandibuloacral dysplasia ( zmpste24 gene coding for the zinc metalloproteinase ste 24 ) , and galloway - mowat syndrome ( gene gmsi coding for gmsi ) . genetic studies are indicated when steroid resistance has been demonstrated in order to withdraw immunosuppression if a mutation is present or in case of congenital nephrotic syndrome before starting any treatment since the great majority of cases is steroid resistant and since steroid treatment can be complicated by severe infectious in neonates . the causes of ns recurrence after transplantation in fsgs cases ( 3050% ) have been recently reviewed and analyzed by the group of antignac . in contrast to patients with an immune form of ns , those with an inherited structural defect of the glomerular filtration barrier represent a subset of patients for whom the primary disease can not a priori recur . surprisingly , recurrence of proteinuria post transplantation has been reported in some patients bearing mutations in the nphs1 , nphs2 , actn4 , and wt1 genes ( for a review , ) , and the mechanism of recurrence remains unsolved for a significant proportion of these cases . the most often provided explanation for recurrence of ns is the development of antibodies against the neoantigen which is suggested by the fact that treatment with steroids , cyclophosphamide , and plasmapheresis may lead to remission ; however , the percentage of graft loss remains significant ( for a review , ) . however , this eventuality is not frequent and genetic studies remain useful before transplantation to precise the risk of posttransplant recurrence and to help for the decision of living kidney donor or not . ins that is not associated with a mutation of genes coding for podocyte 's proteins is actually thought to be the consequence of an immunological dysfunction leading to a circulating factor that modifies the permeability of the glomerular filtration barrier . in 1974 , shalhoub proposed that mcns was a disorder of lymphocyte function with increased plasma levels of a lymphocyte - derived permeability factor . this hypothesis was based on several clinical observations that suggested the involvement of the immune system in the pathogenesis of idiopathic ns , for example , the response to immunosuppressive drugs and the association with hodgkin disease and with allergy . many reports have been published on patients who developed ns after having experienced allergic reactions to inhaled allergens , with vaccinations , food , and insect stings . furthermore , the incidence of atopy was reportedly higher in patients with idiopathic ns than in healthy subjects , ranging from 17 to 40% in mcns patients compared with 1023% in age - matched control subjects . allergy is associated with an elevated production of ige by b - lymphocytes , and several investigators have reported an elevation of ige in the serum of ns patients . the role of circulating factor is particularly suggested by the following observations ( for a review , ): ( 1 ) lmmediate recurrence of proteinuria after transplantation , ( 2 ) transfer of proteinuria to fetus , ( 3 ) efficacy of plasmapheresis and immunoadsorption in reducing proteinuria , ( 4 ) transfer of proteinuria to rat after injection of serum or plasma . valuable studies are difficult to obtain since homogeneous patients groups are necessary and it is often not the case since the duration of proteinuria varies at the time of presentation and some patients may have already started treatment . hyperlipidaemia which is a common complication may activate the immune system , as shown by lenarsky et al . . van de berg and weening have recently reviewed the immunological role of t cells and cytokines in ins . frank et al . showed that cd8-positive t - cells of idiopathic ns patients are clonally expanded , which was not observed in healthy controls . zhang et al . showed high levels of nf-b ( nuclear factor b ) dna - binding activity in t - cells from untreated mcns patients during relapse compared with the mcns patients in remission while treated with immunosuppressants . kimata et al . studied the unstimulated production of cytokines by t - lymphocytes of mcns patients and found an increased production of il-13 , whereas production of il-4 was normal . an elevated expression of il-13 mrna was shown by yap et al . using a semiquantitative rt- ( reverse transcriptase- ) pcr technique . using a subtractive cdna library screening technique , zhang et al . reported differential expression of transcripts involved in the t - cell receptor - mediated complex signaling cascade and a decreased expression of il-12 receptor 2 mrna by pbmc in untreated mcns patients during relapse compared with mcns patients in remission . the latter studies reveal the involvement of t - cells in the pathogenesis of idiopathic ns and , more specifically , th2-mediated immunity . van de berg and weening have studied , by quantitative real - time pcr , the expression of il-1 , il-1ra ( il-1 receptor antagonist ) , il-2 , il-4 , il-5 , il-9 , il-10 , il-13 , tnf- , and ifn-by pbmc from patients with mcns during relapse and remission and from a control group of patients with ns primarily caused by endogenous alterations within the glomerular filter , for instance , mutations in the genes encoding nephrin and podocin . out of the cytokines studied , only the expression of il-10 and il-13 mrna was significantly upregulated in relapsing mcns patients when compared with mcns patients in remission . the latter authors and others ( for a review , ) have shown that podocytes constitutively express functional transmembrane receptor complexes for il-4 , il-10 , il-13 , and tnf-. the possible role of il-13 is also suggested by the ns rat model of lai et al . . il-13 was overexpressed in wistar rats through transfection of a mammalian expression vector cloned with the rat il-13 gene , into the quadriceps by in vivo electroporation . the il-13-transfected rats showed significant albuminuria , hypoalbuminemia , and hypercholesterolemia when compared with control rats . no significant histologic changes were seen in glomeruli of il-13-transfected rats . however , electron microscopy showed up to 80% of podocyte foot process fusion . glomerular gene expression was significantly downregulated for nephrin , podocin , cd 80 , and dystroglycan . immunofluorescence staining intensity was reduced for nephrin , podocin , and dystroglycan il-4r alpha in il-13-transfected rats compared with controls . abdel - hafez et al . suggest in review of the literature and of their own results that the relation between allergy and ins could be the stimulation by il-13 of the expression of cd 80 on podocytes . they report that urinary cd80 levels are increased in patients with mcd during relapse and return to normal after remission . they also have preliminary evidence that the source of the cd80 is the podocyte because they found , by using immunohistochemical staining , that cd80 was expressed by podocytes in kidney biopsy specimens from patients with mcd in relapse . the successful treatment of srns on native kidney or after kidney transplantation with anti - tnf antibodies strongly suggests that this cytokine participates to the pathogenesis of some types of idiopathic nephrotic syndrome [ 21 , 22 ] . it is also suggested by high levels of tnf in patients with active disease and tnf normalization with remission and by an animal model of ns that is controled by anti - tnf agents ( for a review , [ 21 , 22 ] ) . the beneficial treatment by rituximab , a monoclonal antibody directed against cd20 , in difficult ssinss suggest , a role for b cells in ins [ 2325 ] . sellier - leclerc et al underline that most arguments gathered by shalhoub in support of t cell dysfunction have a counterpart to supporting b cell dysfunction and that the contribution of b cells and the potential role of immunoglobulin chains in modifying the glomerular permeability to protein in children with steroid - sensitive nephrotic syndrome have been repeatedly reported . in addition , b cells may be involved through an unidentified antibody - independent pathway , that might be a control on t cells . aside cytokines other factors have been suspected to be involved in the pathogenesis of ins . one of them consists in radical oxygen species ( ros ) that have been largely studied by the group of ghiggeri ( for a review , ) . some experimental models of nephrotic syndrome result from substances as puromycine and adriamycin that induce oxidative stress in glomeruli . furthermore the injection of h2o2 induces proteinuria in rats and no prevents the increase of permeability to albumin induced by the production tnf alpha - induced o2- production in an isolated rat glomeruli system . recently , bertelli et al . demonstrated a 10-fold increase of ros production by resting pmn in ins compared to normal pmn . when pmns were separated from other cells , ros increased significantly in all conditions while a near normal production was restored by adding autologous cells and/or supernatants in controls , vasculitis , and postinfectious glomerulonephritis but not in ins . the second finding was that the oxidative burst by pmn was regulated highly by t lymphocytes , mainly tregs , by means of soluble factors and that this regulatory circuit was altered in ins . the group of moin saleem has intensively studied the possible pathophysiological role of hemopexin ( hx ) in ins [ 30 , 31 ] . it is predominantly produced in the liver , and it increases in the acute phase reaction to inflammation or infection . it has been suggested that in normal conditions circulating hx is inactive but under certain circumstances hx becomes activated as a serine protease . kidney sections incubated with hx have a reduction of the anionic layer and reduced sialoglycoproteins . in vivo , activated hx induced reversible proteinuria in rats parallel to podocyte foot process effacement . activated hemopexin is increased in children with minimal change nephrotic syndrome . in vitro within 30 minutes of treatment with hemopexin , actin reorganized from stress fibers to cytoplasmic aggregates and membrane ruffles in wild - type podocytes [ 30 , 31 ] . this process is nephrin dependent since it did not occur in nephrin - deficient podocytes and in cells that do not express nephrin and was inhibited by preincubation with human plasma . in addition , hemopexin led to a selective increase in the passage of albumin across monolayers of glomerular endothelial cells and to a reduction in glycocalyx . what remains to be elucidated is the primary events leading to the activation of hx . a possibility resides in the inhibition of hx inhibitors or in their leakage in urine . in the latter case , hx activation should be only a secondary event depending on the increased permeability of the glomerular filtration barrier to proteins . the group of virginia savin in us has studied and characterized the circulating factor in fsgs by analyzing the plasma of patients presenting with a posttransplant relapse ( for a review , ) . those studies are based on standard methods of biochemical purification and analyses of molecular characteristics followed by gel electrophoresis and mass spectrometry . they have used a functional assay of permeability activity with isolated rat glomeruli that shows changes in the glomerular capillary permeability to albumin after incubation with the patient plasma or serum . this assay has made it possible to perform sequential purification steps and select fraction(s ) with enhanced activity . using galactose as an effective affinity material to enrich activity of fsgs plasma they reported that cardiotrophin - like cytokine factor 1 ( clc-1 ; encoded by clcf1 ) , a member of the interleukin 6 family , is present in the enriched fraction of fsgs plasma , and that clc-1 increases glomerular palb , and its injection causes proteinuria in rats . pathophysiological consequences of nephrotic syndrome as hypovolemia , acute renal failure , edema , hypercoagulation , and infections should be treated symptomatically . the basis of treatment of ins by steroids was the early hypothesis of the implication of an immunological factor in the pathophysiology of the disease . however , recent experiments using podocytes in vitro have shown that steroids and cyclosporine , aside their effect on the immunological system , may also act directly on the podocyte to stabilize its structure [ 32 , 33 ] . recovery of a normal permeability to proteins of the glomerular filtration barrier is rendered possible by steroids in the majority of cases . the cochrane reviews that use severe criteria of quality to analyze results of clinical studies are very useful for the clinician . the treatment of the primary immunological cause of ins resides in the use of steroids . initial dosage of 60 mg / m / d prednisone or prednisolone pursued by alternate day administration to reduce side effects is a common feature . steroids protocols are mostly differing by duration , way of tapering , and definition of steroid resistance . a 2007 cochrane review has analyzed the results of the literature concerning the best initial steroid treatment . the authors conclude that in their first episode of ssns patients should be treated for at least three months with an increase in benefit for up to seven months of treatment . for a baseline risk for relapse following the first episode of 60% with two months of therapy , daily prednisone or prednisolone given for four weeks followed by alternate - day therapy for six months would reduce the number of children relapsing by 33% . however , it remains to be known if the time of administration or the cumulative steroid dosage is the most important determinant . this question is addressed partly in a double blind rct comparing the effect of the same cumulative prednisone dosage given during 3 or 5 months by a study actually going on at the university of rotterdam ( the netherlands ) . in case of frequently relapsing or steroid - dependent nephrotic syndrome , repeated and prolonged high dosage of prednisone might lead to severe side effects as growth retardation , osteoporosis , infections , diabetes , cataract , hypertension , hirsutism , and cushing aspect . in order to avoid the latter , different nonsteroid treatments have been used . the 2008 cochrane analysis of the already published results of the latter concludes that eight - week courses of cyclophosphamide and prolonged courses of cyclosporin and of levamisole reduce the risk of relapse in children with relapsing ssns compared with corticosteroids alone . they conclude also that clinically important differences in efficacy are possible and that further comparative studies are still needed . infection , infertility , and possibly cancer at long term for cyclophosphamide and hypertension , hirsutism , and chronic renal failure for cyclosporine lead the clinicians to consider alternative treatments . a preliminary study comparing cyclosporine to mycophenolate mofetyl suggested that the latter could replace cyclosporine in the majority of cases . recently , the successful use of rituximab , a monoclonal anti - cd20 antibody , has been reported to prevent relapses in difficult steroid - dependent ssins in several reports [ 2325 ] . however , the latter should be confirmed in a rct to better define efficacy , modalities ( administration during relapse or after remission induction , used alone or in combination ) , indications , and safety . presently , rituximab has to be considered as the last option , for patients who can not be managed properly despite alternate day steroids , mmf , and anticalcineurins ( because of ongoing relapses and/or side effects of drugs and/or noncompliance ) . the last cochrane review on this topic reports that when cyclosporin was compared to placebo or no treatment , there was a significant increase in the number of children who achieved complete remission . cyclosporin also significantly increased the number of children , who achieved complete or partial remission compared with iv cyclophosphamide . the analysis of results of treatment of srins is complicated by the heterogeneity of steroid resistance definition according to protocols and countries . another issue concerns the lack of studies considering the presence of mutations of podocytes proteins to explain different responses to treatment . it seems indeed important to distinguish the latter from srins possibly due to a circulating factor since any immunological treatment is most probably ineffective in the former . when no mutation is detected , a circulating factor might be suspected , particularly when a recurrence is observed after kidney transplantation . in that case , removal of the latter by plasma exchange can be effective but recurrence is often observed when treatment is interrupted . the future treatment strategy will imply the determination of the latter factor and the use of specific treatment to antagonize its effect . based on this principle , rcts studying the effect of galactose and anti - tnf ab have already been initiated by the font study group . another approach used by the latter group is to antagonize the effect of tgf , a cytokine playing a major role in fibrotic processes . Output:	idiopathic nephrotic syndrome ( ins ) is the most frequent form of ns in children . ins is defined by the association of the clinical features of ns with renal biopsy findings of minimal changes , focal segmental glomerulosclerosis ( fsgs ) , or mesangial proliferation ( mp ) on light microscopy and effacement of foot processes on electron microscopy . actually the podocyte has become the favourite candidate for constituting the main part of the glomerular filtration barrier . most cases are steroid sensitive ( ssins ) . fifty percents of the latter recur frequently and necessitate a prevention of relapses by nonsteroid drugs . on the contrary to ssins , steroid resistant nephrotic syndrome ( srins ) leads often to end - stage renal failure . thirty to forty percents of the latter are associated with mutations of genes coding for podocyte proteins . the rest is due to one or several different circulating factors . new strategies are in development to antagonize the effect of the latter .
PubmedSumm2009	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the authenticity and true status of tobacco use , especially in the form of smoking among the patient clientele is always a matter of concern for their physicians . assessing a true status of smoking is important for physicians in regards to initiation , maintenance and successful completion of smoking cessations programs . in addition , it can be helpful for appropriate antenatal care and fetal wellbeing 's monitoring in pregnant patients who may hesitate to truly state their habits , use and abuse of tobacco smoking . moreover , knowledge of true status of tobacco smoking can guide the anesthesiologists in the preoperative assessment of tobacco smokers , wherein the perioperative pulmonary complications can be directly related to the presence , duration and grading of tobacco smoking abuse . finally , the true status of tobacco use can help the administrators to effectively implement the smoking cessation and health promotion programs among their students , their employees as well as general population in question . exhaled carbon monoxide analyzers have been used for the true status assessment of tobacco smoking . besides the utility in tobacco smoking abuse , extremely high levels of detected carbon monoxide by these analyzers give an additional indication for potential carbon monoxide poisoning that can be secondary to undetected home environments or occupational environments related exposures to carbon monoxide . even though , awareness toward hazards of smoking is increasing among common masses , individuals find it difficult to quit their years old habit of smoking . the use of smoke check meter for analysis of exhaled breath carbon monoxide is the need of hour so that such cases can be repeatedly motivated through counseling sessions and cross checked by smoke check meter until the time the individual is able to quit smoking for his / her health benefits . the purpose of the current study was to assess the authenticity of self - reported habit of tobacco smoking among a population sample of male respondents in rural india . after institutional review board approval and written informed consent , 205 respondents who had presented as personal caregivers for patients at an academic university hospital in rural india , were invited to enroll in this prospective correlation study . after taking informed consent , principal investigator who is a community medicine physician , asked these respondents to complete oral questionnaires that assessed their status of tobacco smoking ( if any ) as well as duration of tobacco smoking ( in years ) , type of tobacco smoking ( bidi that is a traditional indian cigarette vs. standard cigarette ) and frequency of tobacco smoking ( number of bidis / cigarettes per day ) . other recorded parameters were respondents age and their occupational history to rule out potential contributions of environmental exposure to carbon monoxide in occupations known to have high risk of carbon monoxide exposure and poisoning . subsequently , the respondents were explained the method of carbon monoxide breath analysis by smoke check , cardinal health , chatham , kent , uk , as shown in figure 1 that was provided free - of - cost to the academic university hospital by cipla , mumbai , india , for research purposes . smoke check meter used in the current study ( cardinal health , chatham , kent , uk ) smoke check meter was allowed to reach room temperature prior to use . mouthpiece adapter with disposable mouthpieces that have unidirectional valve were used to prevent cross - contamination between the respondents . once the unit was ready , the respondent was asked to hold breath for 20 s with smoke check buzzing at the end of countdown from 20 to 0 . this 20-s delay was to allow the equilibration of respondents alveolar carbon monoxide levels with circulating carboxyhemoglobin levels in blood . if the respondents were not able to hold their breath for 20 s , they were allowed to exhale in the meter earliest at the end of 12 s when the auto - zero function of the smoke check had been completed . after the countdown ( usual duration 20-s , but minimum 12-s ) , the respondents were asked to seal the mouthpiece with their lips followed by slow , but complete exhalation so that smoke check could read their exhaled carbon monoxide levels . interpretations of smoke check meter readings ( cardinal health , chatham , kent , uk ) after the completion of data collection , the responses to oral questions were statistically analyzed by investigator ( blinded to either data acquisitions ) for the presence of authenticity of tobacco smoking among the respondents correlation as indicated by the objective and confirmatory test results of smoke check . chi - square test and fisher exact test with contingency tables was applied for comparing proportions . probability statistics with likelihood ratios ( lrs ) ( with 95% confidence limits ) were analyzed with the help of online vassarstats ( richard lowry , 1998 - 2014 , united states ) . there was incomplete data collection in 30 respondents primarily due to inability to appropriately perform the breath analysis tests . there was no significant difference in the respondents age across the various smoke check color indicators groups as shown in table 2 . the smoke check color indicators were significantly different ( p < 0.0001 ) in the respondents who were diagnosed smokers per oral questionnaires versus diagnosed nonsmokers per oral questionnaires as shown in table 3 . there was no significant effect of the potential occupational exposure to carbon monoxide among smokers ( p = 0.76 ) as well as nonsmokers ( p = 0.079 ) ; however , the p value was much lower among nonsmokers than among smokers suggesting that potential occupational exposure to carbon monoxide may have affected our results ( without reaching the level of statistical significance ) primarily for the respondents who were nonsmokers as shown in table 4 . the duration of tobacco abuse was not compared as majority of respondents did not remember their actual durations of abuse . after excluding the four respondents who smoked both bidis and cigarettes , the smoke check color indicators groups for smokers were compared for the amount / frequency of tobacco smoking . it was determined that the gradually increasing number of bidis / cigarettes consumed per day reflected in the higher grades ( green to amber to red ) of smoke check color indicators that was statistically significant ( p = 0.04 for bidis consumers vs. p = 0.0006 for cigarettes consumers ) as shown in table 5 . the low number of smokers in red with alarm indicator groups and ambiguity of the number of bidis / cigarettes consumed by these respondents precluded their inclusion in the final statistical analysis as shown in table 5 . the probability statistics of authenticity of oral questionnaires for assessing smoking status showed that self - reporting was only 75% sensitive and 76% specific with 80% positive predictive value and 70% negative predictive value when compared with the confirmatory smoke check meter results as shown in table 6 . this means that respondents who were nonsmokers per oral questionnaires either under - reported their smoking status with maximum under - reporting level 30% ( 1-negative predictive value ) or were potentially exposed to high carbon monoxide at their occupations or homes . in our respondents pool , there were only a total of 12 respondents who were exposed to potentially high carbon monoxide levels at their occupations ( painter , driver , mill worker , police inspector , welding employee , sewer worker , mechanic , and fabrication worker ) but this did not confound our results significantly as shown in table 4 suggesting under - reporting by the respondents about their tobacco smoking status as the primary underlying cause for our study results . although , the lrs were not very far removed from equivocal number 1 ( the best results for lrs are > 5 and < 0.2 for positive test lr+ and negative test lr respectively ) , still the lrs for the current study were good enough ( lr+ = 3.16 ; lr = 0.33 ) , which are interpreted as following : comparison of respondents ages included in the study correlation between the smoke check readings and oral questionnaires of all respondents effect of coexistent potential occupational exposure to carbon monoxide and its effects on the smoke check readings quantitative bidi or cigarette smoking among smokers and its correlation with smoke check readings ( four respondents excluded who were combo smokers ) probability of oral questionnaire 's reliability as screening tool for cigarette smoking when the respondent had said i am smoker , it was 3.16 times more likely to have nongreen indicator versus green indicator on confirmatory smoke check meterwhen the respondent had said i am nonsmoker , it was 0.33 times more likely to have nongreen indicator versus green indicator on confirmatory smoke check meter . when the respondent had said i am smoker , it was 3.16 times more likely to have nongreen indicator versus green indicator on confirmatory smoke check meter when the respondent had said i am nonsmoker , it was 0.33 times more likely to have nongreen indicator versus green indicator on confirmatory smoke check meter . policies for health promotion at large and smoking cessation programs at individual levels requires authentic self - reporting and recognition of tobacco abuse as personal and major risk factor that is related to many preventable or modifiable diseases including but not limited to cardiac diseases , pulmonary disorders , and cancers . the objectivity of exhaled breath carbon monoxide analyzers provides this opportunity to both tobacco abusers as well as their treating physicians . these treating physicians encompass various patient caregiving fields including but not limited to community medicine , public health , anesthesiology , primary care , family medicine , internal medicine , obstetrics , occupational medicine , and adolescent health . these monitors additionally instill awareness among the sworn - nonsmokers , wherein the high levels of exhaled breath carbon monoxide will be related to unintentional exposures to carbon monoxide at their homes and/or occupations . even though , our study did not investigate with the respondents about their reasons for under - reporting , the potential reasons for under - reporting are very light smoking , already trying to quit , concerns about social desirability , fear of repercussions , and simple obstinacy to lie . although the exhaled breath carbon monoxide meters are considered reliable for assessing tobacco smoking , the only primary controversy related to them is the cut - off point for judging smoking status . like most meters , smoke check that we used for our study has a cut - off of 6 parts per million of carbon monoxide as the cut - off point for the diagnosis ; however some authors have used cut - off of 10 parts per million . in india , especially in rural areas , people prefer to smoke bidis and this reflects in our study results too with 68% respondents being bidi - smokers and 32% respondents being cigarette smokers . even though , it had been reported that bidi - smoking create more carbon monoxide levels in the smokers , our results in table 5 suggest that when compared to bidi consumption , lower numbers of cigarette consumption reach the equivalent levels of smoke check color indicators ; however , these lower numbers did not reach levels of significance secondary to insufficient power of the analysis and small sample size . the question about time since last bidi - cigarette smoke was not asked from the respondents that might have explained some smokers showing green indicator on smoke check . due to half - life of carboxyhemoglobin ( about 5 h during sleep and much less after physical exercise ) , sometimes it has been recommended to perform presmoking breath analysis followed by postsmoking breath analysis within 10 min after last smoke . as all the readings for exhaled breath analysis were performed in the prenoon hours , the comparative analyses among the respondents were not confounded . although it has been recommended to perform exhaled breath analysis during afternoons to avoid the misleading low levels detected in the early morning checks for carbon monoxide that would have fallen due to its half - life during the sleep , high levels of carbon monoxide detected in morning checks for carbon monoxide indicate strong and high tobacco smoking dependence . hence , our method of prenoon checks for carbon monoxide may have contributed to some of the false positives ( smoker per questionnaire , but green indicator per smoke check meter ) . the confrontation question about the reason of under - reporting was not asked from the respondents who showed nongreen indicator on smoke check even though they had said that they were nonsmokers . moreover , this posttest investigation would have delineated some respondents potential exposure to carbon monoxide in their home or occupation environments if they would have adamantly denied tobacco smoking in the posttest questioning . moreover , since this equipment is very cost - effective , its use at a larger scale , that is , at primary health care level will go a long way in assisting the indian tobacco control initiative cotpa act 2003 and bring about a tobacco free society . in summary , true status of tobacco smoking needs to be assessed at various stages of patient caregiving , and exhaled breath carbon monoxide analysis can be an easy tool to this aim with easy provisions and acceptability among the patient populations that we as physicians cater to in rural india . Output:	background : the authenticity and true status of tobacco use , especially in the form of smoking among the patient clientele is always a matter of concern for their physicians.objectives:the purpose of this study was to assess the authenticity of self - reported habit of tobacco smoking among a population sample of male respondents in rural india . methods : respondents were asked to complete oral questionnaires that assessed their status of tobacco smoking ( if any ) as well as duration of tobacco smoking , type of tobacco smoking , and frequency of tobacco smoking . subsequently , exhaled breath carbon monoxide analysis was performed to detect their amounts of exhaled carbon monoxide.results:in 175 respondents , the smoke check color indicators were significantly different ( p < 0.0001 ) in the respondents who were diagnosed smokers per oral questionnaires ( n = 92 ) versus diagnosed nonsmokers per oral questionnaires ( n = 83 ) . the probability statistics of authenticity of oral questionnaires for assessing smoking status showed that self - reporting was only 75% sensitive and 76% specific with 80% positive predictive value and 70% negative predictive value.conclusion:true status of tobacco smoking with exhaled breath carbon monoxide analysis can be an easy clinical maneuver with community health screening and health promotion implications among patient populations in rural india .
PubmedSumm2010	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: iran , which is located in southwest asia , located in an epidemiologic transition and faces the double burden of diseases . breast cancer is the most common cancer in iranian women ; meanwhile the iranian patients are relatively young . given that iran has a female population of about 38 million , this corresponds to a total number of 8500 new cases of breast cancer annually . breast cancer has more frequently occurred in wealthy countries due to a higher prevalence of bc risk factors , such as aged population , older age at first pregnancy , nulliparity , inappropriate breastfeeding , high - calorie intake , sedentary occupation and steroid hormonal consumption . on the other hand , bc survival would be lower in less affluent countries , and in women with low income , or educational level . based on the report from the ministry of health and medical education ( mohme ) ; cancer is the third cause of death in iran after cardiovascular heart disease , then the injuries [ 3 , 4 ] . experimental data has strongly suggested that estrogens have a role in the development and growth of breast cancer . tumor formation may also result from excessive hormonal stimulation of an organ in which normal growth and function would be under endocrine control . the response of an organ to the proliferative effects of a hormone might be a progression from normal growth , to hyperplasia , then to neoplasia . indirect evidence of this sequence has included the increased risk of breast cancer associated with early menarche , late first full - term pregnancy , and late menopause as well as the reduced risk associated with early menopause . immune surveillance has described the process whereby precancerous and malignant cells have recognized by the immune system as damaged , and then would be consequently targeted for elimination . natural killer ( nk ) cells and cytotoxic t lymphocytes ( ctls ) both have played important roles in the innate and adaptive immunity against intracellular pathogens and tumor cells . granule exocytosis is the main pathway for immune elimination of virus - infected cells and tumor cells by ctls and nk cells . after target - cell recognition , release of the cytotoxic granule contents in to the immunological synapse , have formed between the killer cell and its target induces apoptosis . the perforin / granzyme ( gzm ) pathway is a major mechanism for these cytotoxic lymphocytes to kill their targets . granzyme h ( gzm h ) has regarded as an orphan granzyme with unknown biologic functions in immune defense cells . gzm h , which expands the cell death - inducing repertoire of innate immune system , was a functional cytotoxic serine protease of nk cell granule . several studies have recently shown gzmh to be constitutively and highly expressed in human nk cells and to possess chymotrypsin - like ( chymase ) enzymatic activity . human gzmh triggers rapid cell death of target tumor cells that is characteristic of dna fragmentation and mitochondrial damage and generation of reactive oxygen species [ 8 , 12 ] . the steroid hormones , estradiol , plays an important role in the progression of breast cancer , and a majority of the human breast cancers start out as estrogen dependent and express the estrogen receptor ( er ) . the biological effects of estrogen have mediated by its binding to one of the structurally and functionally distinct ers ( er and er ) . the action of progesterone has mediated through its intracellular cognate receptor , the progesterone receptor ( pr ) , which functions as a transcription factor that regulates gene expression . mutation or aberrant expression of the coregulators might thus affect the normal function of the pr , and hence disrupt the normal development of the mammary gland , which might lead to breast cancer . 30 breast cancer patients who were under study at cancer research center , shahid beheshti university of medical sciences ( crc , sbums ) , and 30 control women in premenopausal status , have participated in this study . a total four ml blood has drawn from patients and control women with a 21-gauge needle by syringe . radioimmunoassay ( ria ) has applied for measurement of serum estrogen ( bio source , kip0629 ) . a sandwich enzyme - linked immunosorbent assay ( elisa ) has applied for measurement of granzyme h in serum ( cusabio , gzmh - e17366h ) . immunohistochemistry ( ihc ) for assaying estrogen and progesterone receptors ( novocastra , product code : rtu - cb11 , rtu - pcr-312 ) result have expressed as meansd of experiments . student 's t - test has used for comparison the means between two groups , logistic regression for bc risk . to assess correlation between granzymeh and estrogen , correlation bivariate test mean of serum levels of granzyme h patients was 71.612.2 and 96.1613.4 in control group ( p<0.0001 ) . mean of serum estrogen in patients group was 107.523.8 and in control group was 90.514.8 ( p<0.003 ) ( table1 ) . there was a weak inverse correlation between gzmh and estrogen levels in patient group ( r=-0.3 p<0.02 ) . spearman correlation analysis between estrogen , er , and pr has indicated a weak positive non - significant correlation in patient group . they are secreted towards a neoplastic target cell subsequent to perforin - dependent delivery to target cell cytosol where they have caused apoptosis in target cell ( tumor ) . according to our finding , experimental data have strongly suggested that estrogens play a great role in development and growth of breast cancer . mean levels of serum estrogen in patients group were statistically higher than controls group ( p<0.003 ) . there was a weak correlation between gzmh and estrogen levels in patient group ( r=-0.3 ) . according to study of jiang x et al . , estrogen has induced increasing levels of granzyme b inhibitor , and caused blocking cell death induced by natural killer cells . so our study has implicated existence of suppressor or problem for producing of gzmh in patients group , but it has seemed to be , that contrary to granzyme b. according to our finding levels of estrogen could nt affect on making positive er , pr . some factors like hormone consumption could increase the risk of bc . logistic regression analysis between hormone consumption with breast cancer , have indicated significant statistically relationship ( p<0.01 , or=32.6 , ci : 1.8 - 57.3 ) ( table 2 ) . epidemiologic data have provided strong evidence for an association between plasma estrogens and breast cancer risk . the 1996 , large meta - analysis of the relationship between oral contraceptive use and breast cancer risk , have shown that a history of recent oral contraceptive use , rather than duration of use , was a better predictor of breast cancer risk . these data have based primarily on older high - dose and moderate - dose oral contraceptive pills and not the recently introduced low - dose pills . it is , therefore , likely that the small increase in breast cancer risk associated with the early formulations of oral contraceptives would diminish with the new low - dose pills . human gzmh triggers rapid cell death of target tumor cells that is characteristic of dna fragmentation and mitochondrial damage and generation of reactive oxygen species .we have observed that decreasing level of granzyme h has increased the risk of breast cancer ( p<0.009,or=0.8,ci:0.8 - 0.9 ) ( table 2 ) . but our study has shown that bmi , number of delivery or breastfeeding , and age at monarch could nt affect on risk of bc . early age at menarche has been found to be one of the important determinants in etiology of breast cancer . many epidemiologic studies have suggested that as younger a woman s age at monarch , would be the higher her risk of breast cancer . since monarch at a young age has associated with earlier onset of regular menstrual cycles , early exposure to hormonal milieu associated with regular ovulatory menstrual cycles might be an important etiologic factor . one case - control study has conducted in brazil , has shown that breastfeeding could not have a protective effect against breast cancer . some researchers have believed that the number of children would not make sense for avoiding breast cancers . but many other studies have revealed that null parity is a great risk for breast cancer and the number of children is a protective factor against breast cancer . another study , which has conducted in nigeria , has shown that parity and breastfeeding are protective against breast cancer . parity and lactation period have reduced the lifetime number of ovulatory cycles , reducing the risk of breast cancer particularly in young mothers . this also possibly has reduced estrogen and increased prolactin production , which might decrease women s cumulative exposure to estrogen , thereby inhibiting the initiation or growth of breast cancer cells . a lower risk of breast cancer in women with high body mass index has reported among premenopausal woman by london et al . ( 1996 ) whereas some studies have found no association . in our study family history could nt have significant effect on bc but in patients group percentage of family history is higher than controls group . in any case , many epidemiologic studies have suggested that a family history of breast cancer increases a woman s risk of developing breast cancer and the extent of risk varies according to the nature of the family history , type of relative affected , age at which relative developed breast cancer the number of relatives affected and might vary according to the age of the individual . there are not any similar studies in iranian populations so further study has suggested to reach a better conclusion . our study has implicated existence of suppressor or problem for producing of gzmh in patients group and levels of estrogen could nt effect on making positive er , pr . our study has implicated existence of suppressor or problem for producing of gzmh in patients group and levels of estrogen could nt effect on making positive er , pr . Output:	backgroundbreast cancer ( bc ) is the most common cancer in iranian women , meanwhile the iranian patients are relatively young . granzyme h ( gzmh ) is a functional cytotoxic serine protease of nk cell granules , which expands the cell death - inducing repertoire of innate immune system . gzmh is constitutively and highly expressed in human nk cells , in order to possess chymotrypsin - like ( chymase ) enzymatic activity . the purpose of this study was to determine gzmh level , in bc and healthy women.methods30 breast cancer patients , and 30 control women in premenopausal status , have participated in this study . gzmh , estrogen levels , and er , pr have been measured in cancer and healthy women subsequently , as using elisa , radioimmunoassay , and immunohistochemistry methods.resultsmean gzmh value was lower in bc than healthy women ( p<0.0001).conclusionour study has implicated existence of suppressor or problem for producing of gzmh in patients group and levels of estrogen could n't effect on making positive er , pr .
PubmedSumm2011	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: patients with internal derangement ( i d ) and osteoarthritis ( oa ) of the temporomandibular joint ( tmj ) primarily undergo serial non - surgical treatfment , including physiotherapy , occlusal splint therapy , and medication , to relieve functional tmj pain.1,2 however , surgical procedures are sometimes required if the symptoms do not improve after nonsurgical treatment and/or are caused by degenerative changes in the condyle.3,4,5,6 however , even after symptom resolution , morphological changes of the condyle persist in these patients . these degenerative changes infrequently lead to severe occlusal deformities such as open bite and facial deformities . despite the necessity of orthodontic and/or orthognathic treatment to correct the deformities in these patients , this case report describes the successful orthodontic treatment of a japanese woman aged 31 years and 11 months who developed open bite due to bilateral condylar degeneration during the treatment for i d and oa of tmj . a japanese woman aged 31 years and 11 months was referred to our department by an oral surgeon for the correction of open bite . she had undergone orthodontic treatment that required bilateral maxillary first premolar extraction between the age of 9 and 15 years . during this period , she experienced bilateral clicking in tmj . at the age of 28 years and 2 months , she developed trismus and pain in the tmj region and visited the department of oral and maxillofacial surgery at our hospital . the oral surgeon diagnosed i d and oa of tmj and prescribed physiotherapy and occlusal splint therapy to relieve the pain . no morphological changes in the condyles were observed at that time ( figure 1a ) . in addition , osteophyte - like degenerative changes appeared in the left condyle ( figure 2 ) . consequently , visually guided tmj irrigation and open tmj surgery were performed , following which her symptoms resolved . however , bilateral degeneration of the condyles during treatment resulted in open bite with clockwise rotation of the mandible . the open bite and condyles were monitored for a year to rule out deleterious changes ( figure 1b ) . subsequently , the patient was referred to the department of orthodontics for the correction of open bite . facial photographs showed a convex profile and mentalis muscle strain on lip closure ( figure 3 ) . intraoral examination revealed open bite between the right and left second premolars ( figures 3 and 4 ) . the overbite and overjet were -3.5 mm and + 1.2 mm , respectively , and the maxillary and mandibular midlines coincided with the facial midline in the frontal view . a class ii molar relationship was observed on both sides , with moderate crowding in the mandibular anterior region . a panoramic radiograph indicated the presence of all four third molars , root resorption in the maxillary incisors , and considerable resorption of the bilateral mandibular condyles ( figures 1b and 5a ) . lateral cephalometric analysis showed the following values : anb angle , 4.5 ; fma , 46.7 ; u1-sn , 95.8 ; l1-mp , 83.7 ( table 1 , figure 5b ) . on the basis of the above findings , the patient was diagnosed with open bite caused by morphological changes in the bilateral condyles . for the definitions for cephalometric measurements in the main text , the following objectives were established : correction of open bite , elimination of mandibular anterior crowding , establishment of a class i molar relationship on both sides , and improvement in the facial profile . to achieve the treatment objectives , particularly open bite correction , we suggested two treatment plans . the first included le fort i osteotomy with upward movement of the maxillary posterior segment to induce counterclockwise rotation of the mandible . the second included orthodontic intrusion of the maxillary and mandibular molars using mini - screw implants to induce counterclockwise rotation of the mandible . the first plan required a surgical procedure under general anesthesia , which imposes heavy psychological and physical burdens on the patient , while the second could facilitate achievement of the treatment objective with minimum intervention , similar to the results of surgical treatment , although the post - treatment stability of the intruded teeth was uncertain . both plans necessitated mandibular second premolar and all third molar extractions to achieve the other objectives . after discussion with the patient , the second plan was selected because of safety and minimum patient burden . in preparation for molar intrusion , transpalatal and lingual arch appliances were placed in the maxillary and mandibular arches , respectively . then , sectional arch wires from the second premolar to the second molar on both sides were placed in the maxillary and mandibular arches . under local anesthesia , mini - screw implants were bilaterally embedded into the buccal side of the alveolar bone between the second premolar and first molar and between the first and second molars in the maxilla and between the second premolar and first molar in the mandible . four weeks after implantation , molar intrusion was initiated by the attachment of elastic chains from the mini - screw implant to the sectional arch wire . eleven months later , the overbite increased from -3.5 mm to 0.5 mm ( figure 6 ) . then , the mandibular second premolars were extracted and comprehensive orthodontic treatment was initiated to eliminate anterior crowding and establish a class i molar relationship by distalization of the maxillary arch ( figure 7 ) . orthodontic treatment was completed at the age of 36 years and 1 month , with the use of the begg retainer in the maxillary arch and the hawley retainer in the mandibular arch for retention . the overbite and overjet had increased from -3.5 mm to 3.5 mm and from 1.2 mm to 2.0 mm , respectively . favorable occlusion with a class i molar relationship and no crowding was established ( figures 8 and 9 ) . a panoramic radiograph showed no signs of progressive root resorption in the maxillary anterior teeth . however , the roots of the mandibular molars showed slight resorption , while root parallelism showed further scope of improvement ( figure 10a ) . lateral cephalometric analysis showed a decrease in the mandibular plane angle from 46.7 to 44.0. the height of the maxillary first molar had decreased by 2.9 mm , while that of the mandibular molar had decreased by 1.2 mm . the facial height had also decreased ( n - me : 133.2 mm to 127.7 mm , ans - me : 78.5 mm to 72.6 mm ; table 1 , figures 10b and 11 ) . the maxillary and mandibular incisors showed palatal and lingual inclination , respectively ( u1-sn : 95.8 to 91.8 , impa : 83.7 to 77.7 ) . superimposition of the pre- and post - treatment lateral cephalograms clearly demonstrated counterclockwise rotation of the mandible and lingual movement of the maxillary and mandibular incisors ( figure 11 ) . computed tomography revealed an unchanged condylar morphology , while t2-weighted magnetic resonance imaging revealed an improvement ( data not shown ) . after two years of retention , her facial profile and occlusion were well maintained , and the patient was satisfied with the treatment outcomes ( table 1 , figure 12 ) . i d and oa are tmj disorders characterized by limited mouth opening with functional tmj pain , and these pathologies occasionally induce morphological changes in the condyles.6,7,9 although the present patient was also diagnosed with i d and oa and was treated at the department of oral and maxillofacial surgery , she unfortunately experienced considerable condylar degeneration , which resulted in open bite . several treatment options are available for the management of this type of open bite : maxillary surgery with le fort i osteotomy , orthodontic and orthognathic surgical procedures , and condylar replacement with a costochondral graft.8,9,10,11,12,13,14,15 these three procedures generally induce counterclockwise rotation of the mandible for open bite correction , although stability of the treatment outcomes remains debatable . recently , several reports on the orthodontic treatment of open bite through molar intrusion were published.16,17,18 molar intrusion with mini - screw implants enables counterclockwise rotation of the mandible , which is similar to the results of the surgical procedures , and carries the added advantage of minimum intervention for miniscrew placement , as opposed to that for surgical procedures under general anesthesia . for our patient , we opted for orthodontic treatment with molar intrusion to correct open bite caused by condylar degeneration . the maxillary and mandibular molars were intruded without incisor extrusion , inducing counterclockwise rotation of the mandible and resulting in open bite correction without any tmj symptoms . however , slight gingival recession occurred in the mandibular anterior region , believed to be a consequence of excessive lingual movement of the mandibular anterior teeth . several reports described that approximately 20% to 30% relapse occurred in a short period of time during retention.17,18,19 in our patient , however , almost no relapse was observed 2 years after treatment ( table 1 , figure 12 ) . this may be due to the peculiarity of this type of open bite , which was caused by condylar resorption , unlike a non - pathological open bite . open bite correction through molar intrusion facilitated restoration of the original vertical dimensions , consequently preserving the functional adaptation of the circumoral musculature . further observation of the overbite and condyles will be necessary to maintain a stable occlusion . the findings from this case suggest that orthodontic intrusion of the molars is a safer and less stressful alternative for the management of open bite due to bilateral condylar degeneration secondary to i d and oa of tmj . furthermore , it improves the patient 's masticatory function , esthetics , and quality of life . Output:	this case report describes the orthodontic treatment performed for open bite caused by internal derangement ( i d ) and osteoarthritis ( oa ) of the temporomandibular joint ( tmj ) . a japanese woman , aged 31 years and 11 months , referred to our department by an oral surgeon had an open bite with clockwise rotation of the mandible and degeneration of the condyle . the overbite was corrected through intrusion of the maxillary and mandibular molars using mini - screw implants to induce counterclockwise rotation of the mandible . then , the mandibular second premolars were extracted and comprehensive orthodontic treatment was performed to establish a class i molar relationship with distalization of the maxillary arch and to eliminate anterior crowding . following treatment , her facial profile improved and a functional and stable occlusion was achieved without recurrence of the tmj symptoms . these results suggest that orthodontic intrusion of the molars is one of the safer and less stressful alternatives for the management of open bite due to degeneration of the condyles caused by i d and oa of tmj .
PubmedSumm2012	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the asymptomatic elevation of liver enzymes is a common problem , and , if it goes undetected , it can damage the liver over time ( 1 ) . in a survey of 1959 blood donors in iran , 5.1% had asymptomatic elevated liver enzymes , and the common diagnoses for half of those affected were nonalcoholic steatohepatitis ( nash ) ( 88% ) , hepatitis c ( 7.7% ) , and alcohol and drug - related liver injury ( 1.9% ) ( 2 ) . another study of about 2000 people in golestan province in northern iran found that the prevalence of persistently - elevated liver enzymes was 3.1% with an unknown etiology in about 80% , hepatitis b in 9.3% , hepatitis c in 6.2% , alcoholic liver disease in 4.6% , and fatty liver disease in 2% ( 3 ) . hgv rna and hgv antibody ( anti - e2 ) seropositivity in blood donors is about 14% and 314% , respectively ( 4 ) . the mean prevalence of hgv in blood donors is 4.8% and is different in diverse world regions , e.g. , 4.5% in caucasians , 3.4% in asians , and 17.2% in negros ( 5 ) . it seems that iran has the least prevalence of hgv , i.e. , about 1% , among other countries in the middle east , e.g. , turkey ( 4.1% ) , kuwait ( 24.6% ) , jordan ( 9.8% ) , and saudi arabia ( 2% ) ( 710 ) . seventy - five percent of infected patients have normal liver enzymes with no symptoms of hepatic disease ; however , it can cause acute and chronic hepatitis ( 11 ) . in a study of 41 acute and 67 chronic hepatitis patients with unknown etiology in china , hgv rna was detected in six patients ( 14.6% ) and 12 patients ( 17.9% ) , respectively ( 12 ) . a similar study of 55 acute unknown hepatitis patients indicated that 29.1% had hgv rna seropositivity and very severe clinical syndrome ( 13 ) . multiple studies have expressed doubts about the relationship between hgv infection and hepatic damage ( 1416 ) . thus , our aim was to investigate the association between hgv infection and unknown chronic hepatitis . the aim of this research was to evaluate the association of hepatitis g with unknown chronic hepatitis . the specific objectives of the study were : to determine the frequency of hgv antibodies in the case group and the control groupto determine the difference in the hgv seropositivity between the two groups to determine the frequency of hgv antibodies in the case group and the control group to determine the difference in the hgv seropositivity between the two groups the asymptomatic elevation of liver enzymes is a common problem , and , if it goes undetected , it can damage the liver over time ( 1 ) . in a survey of 1959 blood donors in iran , 5.1% had asymptomatic elevated liver enzymes , and the common diagnoses for half of those affected were nonalcoholic steatohepatitis ( nash ) ( 88% ) , hepatitis c ( 7.7% ) , and alcohol and drug - related liver injury ( 1.9% ) ( 2 ) . another study of about 2000 people in golestan province in northern iran found that the prevalence of persistently - elevated liver enzymes was 3.1% with an unknown etiology in about 80% , hepatitis b in 9.3% , hepatitis c in 6.2% , alcoholic liver disease in 4.6% , and fatty liver disease in 2% ( 3 ) . hgv rna and hgv antibody ( anti - e2 ) seropositivity in blood donors is about 14% and 314% , respectively ( 4 ) . the mean prevalence of hgv in blood donors is 4.8% and is different in diverse world regions , e.g. , 4.5% in caucasians , 3.4% in asians , and 17.2% in negros ( 5 ) . it seems that iran has the least prevalence of hgv , i.e. , about 1% , among other countries in the middle east , e.g. , turkey ( 4.1% ) , kuwait ( 24.6% ) , jordan ( 9.8% ) , and saudi arabia ( 2% ) ( 710 ) . seventy - five percent of infected patients have normal liver enzymes with no symptoms of hepatic disease ; however , it can cause acute and chronic hepatitis ( 11 ) . in a study of 41 acute and 67 chronic hepatitis patients with unknown etiology in china , hgv rna was detected in six patients ( 14.6% ) and 12 patients ( 17.9% ) , respectively ( 12 ) . a similar study of 55 acute unknown hepatitis patients indicated that 29.1% had hgv rna seropositivity and very severe clinical syndrome ( 13 ) . multiple studies have expressed doubts about the relationship between hgv infection and hepatic damage ( 1416 ) . thus , our aim was to investigate the association between hgv infection and unknown chronic hepatitis . the aim of this research was to evaluate the association of hepatitis g with unknown chronic hepatitis . the specific objectives of the study were : to determine the frequency of hgv antibodies in the case group and the control groupto determine the difference in the hgv seropositivity between the two groups to determine the frequency of hgv antibodies in the case group and the control group to determine the difference in the hgv seropositivity between the two groups a case - control study was conducted at the ebne - sina army hospital in hamadan , iran , from september 2011 through october 2012 . the cases were 35 military staff with unknown chronic hepatitis , and the controls were 59 patients with normal serum alt levels who were selected from the orthopedic and gynecologic clinics at the same hospital . chronic hepatitis was defined as serum alt40 iu / l for more than six months ( 17 ) . a checklist containing demographic data , alt levels , and hgv antibody states was designed to collect the data ( table 1 ) . one hundred and seventy - three subjects with serum alt 40 iu / l were evaluated for : viral hepatitis b and c by the detection of hb antigens and hcv antibodies using the elisa method with 5-ml samples of serum.non-alcoholic fatty liver disease ( nafld ) using abdominal sonography by an expert radiologist.history of alcohol or hepatotoxic drug consumption or recent infectious processes , such as cold , fever , sore throat , myalgia , arthralgia , diarrhea , abdominal pain , and vomiting ( 2 , 3 , 17 ) . viral hepatitis b and c by the detection of hb antigens and hcv antibodies using the elisa method with 5-ml samples of serum . non - alcoholic fatty liver disease ( nafld ) using abdominal sonography by an expert radiologist . history of alcohol or hepatotoxic drug consumption or recent infectious processes , such as cold , fever , sore throat , myalgia , arthralgia , diarrhea , abdominal pain , and vomiting ( 2 , 3 , 17 ) . any patients who had any of the three conditions described above were excluded from the study . in this way , 115 subjects were selected for the study , and they were followed for 69 months without any intervention , and 35 patients with permanent alt 40 iu / l were selected as the case group . the control group was comprised of patients with normal serum alt levels who were referred to the orthopedic and gynecologic clinics at the same hospital . five - milliliter serum samples were taken from the members of both the case and control groups and sent to a private laboratory to be assessed for hgv infection . the laboratory used the cortez 4 generation kit ( manufactured in the u.s . ) , and hgv anti - e2 was detected by elisa with 99.8% sensitivity . this research was approved by the ethics committee at the aja university of medical sciences . participation in the study was voluntary , and the physicians explained the purpose and procedures of the study to the participants and asked each one to sign a consent form . for the sake of confidentiality and privacy , spss version 18 ( spss inc . , chicago , il , u.s . ) was used to analyze the data . fisher s exact test , the student s t - test , and multivariate logistic regression were used for data analysis . a case - control study was conducted at the ebne - sina army hospital in hamadan , iran , from september 2011 through october 2012 . the cases were 35 military staff with unknown chronic hepatitis , and the controls were 59 patients with normal serum alt levels who were selected from the orthopedic and gynecologic clinics at the same hospital . chronic hepatitis was defined as serum alt40 iu / l for more than six months ( 17 ) . a checklist containing demographic data , alt levels , and hgv antibody states was designed to collect the data ( table 1 ) . one hundred and seventy - three subjects with serum alt 40 iu / l were evaluated for : viral hepatitis b and c by the detection of hb antigens and hcv antibodies using the elisa method with 5-ml samples of serum.non-alcoholic fatty liver disease ( nafld ) using abdominal sonography by an expert radiologist.history of alcohol or hepatotoxic drug consumption or recent infectious processes , such as cold , fever , sore throat , myalgia , arthralgia , diarrhea , abdominal pain , and vomiting ( 2 , 3 , 17 ) . viral hepatitis b and c by the detection of hb antigens and hcv antibodies using the elisa method with 5-ml samples of serum . non - alcoholic fatty liver disease ( nafld ) using abdominal sonography by an expert radiologist . history of alcohol or hepatotoxic drug consumption or recent infectious processes , such as cold , fever , sore throat , myalgia , arthralgia , diarrhea , abdominal pain , and vomiting ( 2 , 3 , 17 ) . any patients who had any of the three conditions described above were excluded from the study . in this way , 115 subjects were selected for the study , and they were followed for 69 months without any intervention , and 35 patients with permanent alt 40 iu / l were selected as the case group . the control group was comprised of patients with normal serum alt levels who were referred to the orthopedic and gynecologic clinics at the same hospital . five - milliliter serum samples were taken from the members of both the case and control groups and sent to a private laboratory to be assessed for hgv infection . the laboratory used the cortez 4 generation kit ( manufactured in the u.s . ) , and hgv anti - e2 was detected by elisa with 99.8% sensitivity . this research was approved by the ethics committee at the aja university of medical sciences . participation in the study was voluntary , and the physicians explained the purpose and procedures of the study to the participants and asked each one to sign a consent form . for the sake of confidentiality and privacy , fisher s exact test , the student s t - test , and multivariate logistic regression were used for data analysis . the mean ages of the members of the case group and the control group were 34.98.5 and 4012.3 years , respectively . all of patients in the case group were males , and the male - to - female ratio in the control group was 2.28 . there was a significant difference between the ages of the patients in the case and control groups , with the former having a significantly lower mean age than the latter based on the student s t - test ( p=0.021 ) ( table 1 ) . among all of the people in the case group and the control group , there was just one person who tested hgv anti - e2 positive in the case group ( 2.9% ) ; all of the people in the control group tested negative . thus , fisher s exact test indicated that there was no significant statistical difference of hgv infection between case group and the control group ( p=0.372 ) . in multivariate logistic regression analysis , a significant relationship was found between unknown chronic hepatitis and being male ( or=14.9 ; 95% ci=1.9117.6 , p=0.010 ) , but there was no such relationship for age ( p=0.446 ) or hgv infection ( p=0.269 ) ( table 2 ) . in our study , the frequency of hgv antibodies was not different between the patients with unknown chronic hepatitis and the healthy subjects , confirming that the hgv infection did not injure the liver . our results were in agreement with the results of a similar study conducted in italy on 50 unknown chronic hepatitis patients and 50 healthy persons ( 18 ) . some studies have concluded that it was doubtful that hgv was responsible for chronic hepatitis ( 19 , 20 ) , and several studies have shown the correlation of hgv with hematologic disorders , such as non - hodgkin s lymphoma ( 2123 ) , aplastic anemia ( 24 , 25 ) and chronic renal failure ( 2629 ) . in our study , chronic hepatitis was more frequent in males due to the selection of the cases from active military patients participating in a military health monitoring program . it seems that co - infection with other viral forms of hepatitis , such as hbv and hcv , can lead to hepatic damage whereas the hgv infection alone does not . one survey of 64 japanese children infected with hcv with co - infection with hgv showed no clinical , virological , or pathological differences between those with hcv / hgv co - infection and hcv alone ( 30 ) . in another study of 130 patients with chronic hepatitis b and 173 patients with chronic hepatitis c , the prevalence of hgv rna was about 8% and 17% , respectively , and , while hgv co - infection did not worsen the clinical symptoms of disease , it accelerated the progression to chronic hepatitis and increased the risk of hepatocellular carcinoma ( 31 ) . regarding the limitation of the study , we did not study patients with unknown chronic hepatitis for auto - immune hepatitis , which might have a role in liver inflammation and chronic hepatitis in these patients . this study showed no association between a history of hgv infection and chronic hepatitis in the patients with unknown chronic hepatitis . however , we believe that hgv rna measurement in the patients with unknown chronic hepatitis may help identify any ongoing inflammation . further studies are recommended to evaluate hgv rna seropositivity in patients with unknown chronic hepatitis . Output:	background : hepatitis g virus ( hgv ) is a hepatotrope virus with unknown importance . the genome of the virus has been detected in patients with acute or chronic non - a - e hepatitis , cirrhosis , and hepatocellular carcinoma . the aim of this study was to determine the association between hepatitis g and unknown chronic hepatitis.methods:this case - control study was performed in ebne - sina military hospital in hamadan , iran . the cases were 35 military staff with unknown chronic hepatitis . the control group consisted of 59 healthy subjects who had normal levels of serum alanine aminoteransferase ( alt ) . the data were analyzed by spss , version18 , using fisher s exact test , the student s t - test , and multivariate logistic regression analysis.results:only one patient in the case group ( 2.9% ) tested positive for hgv antibodies , and no one was infected in the control group . there was no association between hgv infection and unknown chronic hepatitis in our study ( p=0.37 ) . a significant association was found between the male gender and unknown chronic hepatitis ( or=14.9 , p=0.01).conclusion : no association between hgv infection and unknown chronic hepatitis was found in our study , so it was not necessary to evaluate these patients for hgv infection .
PubmedSumm2013	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: increased morbidity from diabetes , hypertension , cardiovascular disease , and inflammatory disorders is common in developing and developed countries [ 2 , 3 ] . more than one - quarter of health care costs related to obesity and associated disorders . promoting weight loss , decreasing body fat , maintenance of negative energy balance , reducing energy intake , and increasing energy expenditure through physical activity reduce body weight . the interest in natural herb supplements such as gymnema sylvestre , green tea , sphaeranthus indicus , garcinia mangostana , dolichos biflorus , and piper betle leaves for treatment of obesity was rapidly growing because of their minimal side effects . the spicy varieties of capsicum were investigated for their effects on weight loss and thermogenesis . commonly called chili peppers , or simply chilies , capsicum is used as a food additive in many regions due to its pungency , aroma , and color . it was safely consumed in large amounts in many countries , especially in mexico and korea , where per capita consumption reaches 15 grams per day . more moderate consumption up to 5 grams daily per capita of capsicum was reported in thailand and india . in u.s . , consumption of all peppers increased , from an average of 6.94 kg per person in 2005 to 8.66 kg per person in 2012 and consumption of bell peppers grew from 4.17 kg to 5.03 kg , while chili pepper consumption grew from 2.77 kg to 3.36 kg . the daily consumption of all peppers was approximately 24 g / person and chili pepper was 9 g / person . capsicum ( capsicum annuum l. or capsicum frutescens l. ) and paprika ( capsicum annuum l. ) were generally recognized as safe ( gras ) for their intended use in food ( 21 cfr 182.10 ; 582.10 ; 21 cfr 182.20 ; 582.20 ; 21 cfr 73.340 ) . the most abundant forms of capsaicinoids found in hot red peppers are capsaicin ( trans-8-methyl - n - vanillyl-6-nonenamide ) , dihydrocapsaicin , and nordihydrocapsaicin . about 70% of the burning sensation experienced from consumption of chili red peppers capsaicinoids ( caps ) are detected rapidly using gas chromatography or supercritical fluid extraction and supercritical fluid chromatography ( sfe / sfc ) . approximately 3 mg of capsaicinoids is present within 1 g of dried red pepper . evidences showed that capsaicin has effects on digestive tract , regulating homeostasis and other deleterious effects . capsicum and capsaicinoids improve metabolism and hormone function , stabilize blood glucose [ 9 , 10 ] , reduce insulin and leptin resistance and endothelial function , inhibiting ldl - cholesterol oxidation , and prevent cancer due to antioxidant activity [ 1416 ] . spicy food consumption showed highly consistent inverse associations with total mortality among both men and women after adjustment with potential risk factors . the adjusted hazard ratios for death were 0.90 ( 95% confidence interval 0.84 to 0.96 ) , 0.86 ( 0.80 to 0.92 ) , and 0.86 ( 0.82 to 0.90 ) for those who ate spicy food 1 or 2 , 3 to 5 , and 6 or 7 days a week , respectively . people consuming spicy foods 6 or 7 days a week showed a 14% relative risk reduction in total mortality . inverse associations were also observed for deaths due to cancer , ischemic heart diseases , and respiratory diseases . capsaicinoids ( caps ) , potential ingredient to support body weight , reduce ad libitum energy intake [ 1719 ] , increase thermogenesis and energy expenditure [ 7 , 9 , 2022 ] , and increase lipolysis [ 7 , 9 , 20 , 21 , 2326 ] . since studies have used a variety of treatments ( chilies and dietary supplements ) within the research design , the results for a given outcome measure the ingested treatment , dose , and product quality . however , assuming the overall effects as noted above , epidemiological data seem to support an association between consumption of caps containing foods and a lower incidence of obesity and associated disorders [ 15 , 27 ] . caps are major naturally occurring pungent principles in red - hot pepper ( figure 1 ) . the current study was a pilot study designed to study the safety and tolerability of capsaicinoids from capsicum extract fruit in a beadlet form , administered to a group of 12 healthy overweight female subjects to escalating dosages of caps from capsicum extract fruit ( i.e. , 210 mg capsaicinoids ) . daily diary data was maintained by subjects to show general tolerability and record adverse events . the study is designed to test different doses of caps product ( 2 mg caps , 4 mg caps , 6 mg caps , 8 mg caps , and 10 mg caps ) on tolerability , safety , and anthropometric and metabolic measures in overweight healthy subjects . the study was a single center , dose - finding , open - label , and adaptive study design with twelve ( 12 ) overweight women taking escalating doses of the study product , each dose for six ( 6 ) weeks . an ascending dose protocol evaluated a total dose of up to 10 mg daily given in different doses ( 2 mg , 4 mg , 6 mg , 8 mg , and 10 mg of caps ) . safety and tolerability are assessed by recording vital signs , electrocardiograms , clinical chemistry parameters , urinalysis , and adverse events . institutional review board ( irb ) approval was received ( copernicus group , irb , cary , nc ) . all participants completed a detailed medical history , prior and concomitant medications , and physical activity questionnaire . healthy female subjects between 25 and 55 years of age and with body mass index ( bmi ) between 25 and 34.9 kg / m were included . subjects were excluded based on regular ingestion of chili peppers ( raw or powdered form ) , black pepper , or ginger or foods known to contain chili peppers , black pepper , or ginger more than 3 times per week . subjects were excluded for any prescribed medications for chronic diseases such as inflammatory disorders , metabolic disorders , diabetes , cardiovascular disease , cancer , hiv / aids , and obesity . subjects with recent history of alcoholism ( within 12 months ) or drug abuse were excluded . study design and number of visits of participants are reported in figure 2 . medical history , review of concomitant medications , vital signs , demographic assessments , anthropometric measurements , physical examination , and laboratory assessments including urine collection for pregnancy and blood draws for cbc and cmp were assessed . subjects returned to the site every week and tolerability reviews were conducted using open - ended questions . subjects were given 2 mg caps ( 100 mg capsimax ) , 4 mg caps ( 200 mg capsimax ) , 6 mg caps ( 300 mg capsimax ) , 8 mg caps ( 400 mg capsimax ) , and 10 mg caps ( 500 mg capsimax ) during each visit . vital signs , medical history , and tolerability survey were recorded for subjects at each visit . blood samples were analyzed for safety markers such as complete blood count ( cbc ) and comprehensive metabolic panel ( white blood cells , red blood cells , hemoglobin ( hb ) , hematocrit , mean platelet volume ( mpv ) , mean corpuscular volume ( mcv ) , red cell distribution width ( rdw ) , platelet count , mean corpuscular hemoglobin , neutrophil , lymphocyte , monocytes , eosinophil , basophil , total protein , albumin , globulin , ratio of albumin and globulin , calcium , chloride , sodium potassium , and anion gap ) and liver , lung , and kidney function tests ( serum glutamic - oxaloacetic transaminase ( sgot ) , serum glutamic - pyruvic transaminase ( sgpt ) , alkaline phosphatase , bilirubin , carbon dioxide , blood urea nitrogen ( bun ) , and creatinine blood urea nitrogen ) . frequency and intensity of adverse event ( ae ) and serious aes were recorded based on the diaries and interviews during each visit . capsimax , a proprietary product , consists of capsaicinoids ( caps ) obtained from dried red fruits of capsicum annuum l. the capsicum extract was standardized into a beadlet form with food grade carbohydrates useful for food applications . capsimax is a faint , pinkish - white colored , free - flowing , uniform spheroidal beadlet with a spicy odor characteristic of dried ripe fruits of capsicum . the product was standardized to 2% capsaicinoids , of which 1.21.35% is capsaicin , 0.60.8% is dihydrocapsaicin , and 0.10.2% is nordihydrocapsaicin . the product has 1525% extract from capsicum , 4555% sucrose , and 3035% cellulose gum coatings . placebo and capsimax capsules ( providing 2 mg , 4 mg , 6 mg , 8 mg , and 10 mg caps ) from capsicum extract in beadlet form are provided by omniactive health technologies ltd . , a modified per - protocol analysis was performed including all subjects who had at least one poststudy product exposure visit . primary endpoints were tolerability assessments and aes and secondary end points were blood safety markers ( complete blood count ( cbc ) and comprehensive metabolic panel ( cmp ) ) . change in assessments for tolerability and safety measures is assessed by anova in a repeated measures design . this allowed for an assessment of the main effect of time , dose , and time by dose interaction . a paired sample t test was used to assess changes over time for each group . mean , standard deviation and significance and categorical variables are summarized as counts and percentages . data derived from diary entries , clinical chemistry assessments , questionnaires , and other relevant assessments at baseline and after supplementation . statistical software for social sciences ( spss version 19 ) was used to run all analysis . table 1 provides descriptive baseline characteristics of the participants . in the current study , 58% subjects were hispanics and 42% were non - hispanic subjects . overweight healthy women participated in the study ( mean bmi : 28 kg / m ) . subjects tolerability survey reports suggest that escalating doses of caps ( 210 mg caps ) had no significant changes in the skin , bowel movements , hair , digestion , urination , mouth or throat , and breathing . at week 2 , only 92% of the subjects reported no changes in their urination . four incidences of adverse events such as sprained ankle , elevated blood pressure , stomach ache , and increased urination are reported . however , in investigator evaluation , these incidences unrelated to product consumption . no significant changes were observed for vital signs , body weight , body mass index , and blood pressure over baseline ( sbp / dbp , table 2 ) . no statistically significant changes were observed in cbc compared to baseline for all markers ( table 3 ) . there were no significant changes in liver , lung , and kidney function test . repeated measures analysis of variance for all assessments revealed no significant effect of dose on tolerability assessments and safety measures . capsaicinoids ( caps ) serve as agonists of the transient receptor potential vanilloid subfamily member 1 ( trpv1 ) . trpv1 releases substance p , which activates the postsynaptic receptor of substance p , neurokinin-1 . caps are found to have weight loss properties based on their potential mechanism of action ( figure 3 ) . activation of neurokinin-1 results in an increased activation of the sympathetic nervous system , leading to the release of epinephrine ( epi ) and norepinephrine / noradrenaline ( na ) . recent systematic review and met analysis [ 30 , 31 ] reported that consumption of 2 mg capsaicinoids increases energy expenditure ( 50 kcal / day ) and would produce clinically significant levels of weight loss in 1 - 2 years . it was also observed that regular consumption reduced much abdominal adipose tissue levels and reduced appetite and energy intake as part of a weight management program . in vitro dissolution data ( unpublished ) for 500 mg capsimax beadlets is equal to 10 mg caps tested in acidic medium similar to stomach ph . in addition , caps from capsimax were released in alkaline medium , which is similar to conditions in the upper intestine . the release of caps in alkaline medium was gradual , over the period of 4 hours . at 4 hours results from this study showed that 6-week consumption of the study product in increasing doses ( 210 mg / d ) has no significant effect on change in skin color , bowel movements , hair , digestion , mouth or throat , and breathing including urination . there were no significant changes in anthropometric measures , clinical chemistries , cbc , metabolic measures , and vital signs compared to baseline measurements . overall , the results of the study indicated that caps supplemented dosages were safe and well tolerated . the current data supports that higher dose of caps was safe and tolerable for human consumption . the results reinforce animal and human studies on the acute effects of caps on metabolic measures such as white blood cells ( wbcs ) , neutrophils , eosinophils , basophils , monocytes , lymphocytes , t lymphocytes , b lymphocytes , and nk cells . capsaicinoids decreased the levels of acquired immunity cells and increased the number of total wbcs and neutrophils without changing the number of monocytes , eosinophils , or basophils . this indicates that intake of caps does not elicit harmful immune response . in vitro and in vivo studies the rats showed oral irritation when fed with normal capsicum extract during pilot experiments , because no irritation was noted with caps ( capsimax beadlets ) . caps and caps plus formulations did not show any adverse effects on body weights or on organs as evaluated by necropsy and histopathological examination . the caps ( capsimax ) and caps containing formulations were tolerable in experimental animals and without any behavior changes . in mutagenesis test , no real increase in revertant colony numbers in any of the five tester strains was observed following caps treatment at any concentration , regardless of metabolic activation ( s9 mix ) . there was also no tendency for higher mutation rates with increasing concentrations in the range below the border of biological relevance . the results of these investigations revealed that caps did not induce gene mutations by base pair changes or frameshifts in the genome of the strains . the results of these investigations suggest that caps were nonmutagenic in both the presence and the absence of metabolic activation . in the chromosomal aberration test , caps were tested in human peripheral blood lymphocyte cultures . caps did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used . caps were well tolerated , with no observed side effects related to gastric upset or discomfort . no difference in heart rate and systolic or diastolic blood pressure was noted as compared to placebo at up to 4 hours after dosing . the current findings suggest that daily consumption of capsaicinoids up to 10 mg was well tolerated and no significant changes in blood chemistries were observed . the limitations of the study were only up to 10 mg capsaicinoids for 7-day intervention for tolerability and safety of the product . this study is only trying to find the dose of tolerability to use in efficacy studies . recent studies suggest that capsaicinoids from capsicum extract alone and/or with other ingredients reported no adverse events and further confirmed that caps are safe [ 26 , 3945 ] . the results of these clinical trials show that 2 mg caps ( capsimax ) were safe and tolerable for human consumption . furthermore , no serious adverse effects were reported , and the regimen was well tolerated . the current findings suggest that daily consumption of 2 mg caps and up to 10 mg caps were well tolerated and no significant changes in liver and kidney functions . further long - term safety and efficacy studies are warranted . the escalating dose levels of caps from capsimax product , a highly concentrated natural capsicum fruit extract , were found tolerable and safe for human consumption . Output:	a single center , open - label , dose - finding adaptive study was conducted in twelve healthy overweight female subjects . the study was to evaluate the safety and tolerability of the capsaicinoids ( caps ) from capsicum extract in a beadlet form compared to placebo in a healthy overweight population . the investigational product capsaicinoids ( caps ) from capsicum extract in a beadlet form ( capsimax ) a proprietary encapsulated form of capsicum extract in beadlet form supplemented at 2 mg , 4 mg , 6 mg , 8 mg and 10 mg of caps . an ascending dose protocol evaluated a total dose of 10 mg daily given in five divided doses ( 2 mg , 4 mg , 6 mg , 8 mg and 10 mg of caps ) . each dose was given for a week . safety and tolerability were assessed . primary outcomes were tolerability assessments and reports of adverse events . tolerability assessments were observed on skin color and any changes in skin , bowel movement , digestion , mouth or throat , hair color or changes in hair color , urination includes frequency and burning sensations , breathing , any changes in their health . secondary outcomes were body weight , body mass index ( bmi ) , blood pressure ( sbp / dbp ) , vital signs , electrocardiograms , clinical chemistry parameters including liver function tests , lung function tests and kidney function tests and complete blood count ( cbc ) . no dose effective changes were observed . the escalating dose levels of caps in a beadlet form product found was tolerable and safe for weight management studies . tolerability assessments and safety blood markers showed no significant changes from baseline . no significant serious adverse events were reported throughout the duration of the study . further longer term studies are required to explore the tolerability of the product . this trial is registered with isrctn : # isrctn10975080 .
PubmedSumm2014	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: in the past years , bcl-2 , the mammalian homologue of ced-9 , the antiapoptotic gene initially described in caenorhabditis elegans , has emerged as a major checkpoint controlling the escape of cells from their death fate ( kroemer 1997 ) . in mammals , bcl-2 is a member of a broad family of related gene products , with various degrees of sequence identity and different functions . while bcl-2 and some members of the family ( e.g. , bcl - x ) protect the cells from apoptosis , others ( e.g. , bax ) have the opposite effect . however , while there is a general consensus on the role of bcl-2 , its mechanism of action remains elusive . on one hand , the various members of the family have been shown to dimerize and to interact with cofactors of caspases , key executors of the apoptotic machinery ( li and yuan 1999 ) ; on the other , the specific localization of the protein on the er and outer mitochondrial membrane ( lithgow et al . 1994 ) suggests that these proteins may interfere with some specific functions localized in these organelles . in particular , bcl-2 has been shown to prevent the release of cytochrome c and aif from mitochondria ( kluck et al . 1997 ) , events that are considered essential in triggering apoptosis , whereas expression of bax has opposite effects ( jurgensmeier et al . thus , the intriguing possibility has emerged that members of the bcl-2 family affect the apoptotic process by modulating ion fluxes in these organelles ( minn et al . in particular , recent work in cell clones stably expressing the oncogene suggests that bcl-2 causes an overload of ca within the er ( lam et al . . however , this effect appears in contrast with the observation that bcl-2 insertion in artificial lipid bilayers results in the formation of ionic channels of low selectivity ( minn et al . such property should in fact cause a reduction in the er ca content ( due to an increased leak ) rather than excess accumulation . in this report , we have undertaken a detailed investigation of ca homeostasis at the subcellular level using the recombinant aequorin ( aeq ) approach developed in our laboratory ( rizzuto et al . we conclude that bcl-2 overexpression affects ca handling by reducing the state of filling of the intracellular ca stores . the implications of these results for the protective action of bcl-2 on apoptosis are discussed . hela cells were grown in dme supplemented with 10% fcs in 75-cm falcon flasks . before transfection , cells were seeded onto 13-mm glass coverslips and allowed to grow to 50% confluence . at this stage , transfection with 4 g of dna ( 3 g bcl-2 + 1 g aeq ) was carried out as previously described ( rizzuto et al . for fura-2 measurements , the cells were seeded onto 24-mm coverslips and transfection was carried out with 8 g plasmid dna ( 6 g bcl-2 + 2 g mtgfp ) . for cytosolic aeq ( cytaeq ) and mitochondrial aeq ( mtaeq ) , the coverslip with the cells was incubated with 5 m coelenterazine for 12 h in dme supplemented with 1% fcs , and then transferred to the perfusion chamber . for reconstituting with high efficiency , the aeq chimeras targeted to the golgi apparatus and the er ( goaeq and eraeq , respectively ) this was obtained by incubating the cells for 1 h at 4c in krb ( krebs - ringer modified buffer : 125 mm nacl , 5 mm kcl , 1 mm na3po4 , 1 mm mgso4 , 5.5 mm glucose , and 20 mm hepes , ph 7.4 , at 37c ) supplemented with coelenterazine 5 m , the ca ionophore ionomycin , and 600 m egta . after this incubation , the cells were extensively washed with krb supplemented with 2% bsa and 1 mm egta , and then aeq reconstitution was carried out as described . agonists and other drugs were added to the same medium . in experiments with permeabilized cells , a buffer mimicking the cytosolic ionic composition ( intracellular buffer , ib : 140 mm kcl , 10 mm nacl , 1 mm k3po4 , 5.5 mm glucose , 2 mm mgso4 , 1 mm atp , 2 mm sodium succinate , 20 mm hepes , ph 7.05 , at 37c ) was employed . this solution was supplemented with either 100 m egta or a fixed [ ca ] ( buffered with 2 mm egta ) . the experiments were terminated by lysing the cells with 100 m digitonin in a hypotonic ca - rich solution ( 10 mm cacl2 in h2o ) , thus discharging the remaining aeq pool . the light signal was collected and calibrated into [ ca ] values as previously described ( brini et al . , a 13-mm round coverslip with the transfected cells was placed in a perfused , thermostated chamber located in the close proximity of a low - noise photomultiplier , with built - in amplifier discriminator . the output of the discriminator was captured by a thorn - emi photon counting board and stored in an ibm - compatible computer for further analyses . the aeq luminescence data were calibrated off - line into [ ca ] values , using a computer algorithm based on the ca response curve of wild - type and mutant aeqs , as previously described ( brini et al . 1997 ) . in some experiments using eraeq , after reaching the plateau level , an artifactual slow decrease in [ ca ] was observed . such an artifact is due to a small fraction of missorted aeq in a low [ ca ] compartment and a simple practical cure for it has been described by maechler et al . 4 , cells transfected with eraeq were used to determine the steady [ ca]er obtained by incubating the cells in low ca medium . two protocols were adopted that gave identical results . in the first , cells incubated in normal medium were depleted according to the standard protocol , but the ca concentration during the refilling was reduced to 100 m . in the second protocol , the cells were preincubated in 100 m cacl2 for 16 h , depleted , and finally refilled with an extracellular ca concentration of 100 m . the coverslip with the cells was incubated with 5 m fura-2 ( added to dme + 1% fcs ) at 37c for 30 min , and after a brief washout was then transferred to the thermostated stage of a zeiss axiovert inverted microscope equipped with a sutter filterwheel and 340/380 excitation filters . the fluorescence data were collected with a princeton instruments back - illuminated camera and expressed as emission ratios , using metafluor software ( universal imaging ) . within the microscope field , transfected and untransfected cells were identified before carrying out the fura-2 monitoring by revealing mitochondrial - targeted green fluorescent protein ( mtgfp ) fluorescence ( excitation 480 nm , emission 510 nm ) . hela cells were grown in dme supplemented with 10% fcs in 75-cm falcon flasks . before transfection , cells were seeded onto 13-mm glass coverslips and allowed to grow to 50% confluence . at this stage , transfection with 4 g of dna ( 3 g bcl-2 + 1 g aeq ) was carried out as previously described ( rizzuto et al . for fura-2 measurements , the cells were seeded onto 24-mm coverslips and transfection was carried out with 8 g plasmid dna ( 6 g bcl-2 + 2 g mtgfp ) . for cytosolic aeq ( cytaeq ) and mitochondrial aeq ( mtaeq ) , the coverslip with the cells was incubated with 5 m coelenterazine for 12 h in dme supplemented with 1% fcs , and then transferred to the perfusion chamber . for reconstituting with high efficiency , the aeq chimeras targeted to the golgi apparatus and the er ( goaeq and eraeq , respectively ) , the luminal [ ca ] of these compartments first must be reduced . this was obtained by incubating the cells for 1 h at 4c in krb ( krebs - ringer modified buffer : 125 mm nacl , 5 mm kcl , 1 mm na3po4 , 1 mm mgso4 , 5.5 mm glucose , and 20 mm hepes , ph 7.4 , at 37c ) supplemented with coelenterazine 5 m , the ca ionophore ionomycin , and 600 m egta . after this incubation , the cells were extensively washed with krb supplemented with 2% bsa and 1 mm egta , and then aeq reconstitution was carried out as described . agonists and other drugs were added to the same medium . in experiments with permeabilized cells , a buffer mimicking the cytosolic ionic composition ( intracellular buffer , ib : 140 mm kcl , 10 mm nacl , 1 mm k3po4 , 5.5 mm glucose , 2 mm mgso4 , 1 mm atp , 2 mm sodium succinate , 20 mm hepes , ph 7.05 , at 37c ) was employed . this solution was supplemented with either 100 m egta or a fixed [ ca ] ( buffered with 2 mm egta ) . the experiments were terminated by lysing the cells with 100 m digitonin in a hypotonic ca - rich solution ( 10 mm cacl2 in h2o ) , thus discharging the remaining aeq pool . the light signal was collected and calibrated into [ ca ] values as previously described ( brini et al . , a 13-mm round coverslip with the transfected cells was placed in a perfused , thermostated chamber located in the close proximity of a low - noise photomultiplier , with built - in amplifier discriminator . the output of the discriminator was captured by a thorn - emi photon counting board and stored in an ibm - compatible computer for further analyses . the aeq luminescence data were calibrated off - line into [ ca ] values , using a computer algorithm based on the ca response curve of wild - type and mutant aeqs , as previously described ( brini et al . 1997 ) . in some experiments using eraeq , after reaching the plateau level , an artifactual slow decrease in [ ca ] was observed . such an artifact is due to a small fraction of missorted aeq in a low [ ca ] compartment and a simple practical cure for it has been described by maechler et al . 4 , cells transfected with eraeq were used to determine the steady [ ca]er obtained by incubating the cells in low ca medium . two protocols were adopted that gave identical results . in the first , cells incubated in normal medium were depleted according to the standard protocol , but the ca concentration during the refilling was reduced to 100 m . in the second protocol , the cells were preincubated in 100 m cacl2 for 16 h , depleted , and finally refilled with an extracellular ca concentration of 100 m . the coverslip with the cells was incubated with 5 m fura-2 ( added to dme + 1% fcs ) at 37c for 30 min , and after a brief washout was then transferred to the thermostated stage of a zeiss axiovert inverted microscope equipped with a sutter filterwheel and 340/380 excitation filters . the fluorescence data were collected with a princeton instruments back - illuminated camera and expressed as emission ratios , using metafluor software ( universal imaging ) . within the microscope field , transfected and untransfected cells were identified before carrying out the fura-2 monitoring by revealing mitochondrial - targeted green fluorescent protein ( mtgfp ) fluorescence ( excitation 480 nm , emission 510 nm ) . to directly investigate the role of bcl-2 in ca homeostasis , we have selectively measured the [ ca ] in different cell compartments , i.e. , the cytoplasm and the organelles acting as sources ( er and golgi apparatus ) or targets ( mitochondria ) of the ca signal . 1992 ; brini et al . 1995 ; montero et al . 1995 ; pinton et al . the first is the high subcellular specificity of the probes , which allows to clearly identify the effects of bcl-2 expression on the signaling properties of the various compartments ; the second is that the ca probe can be cotransfected with bcl-2 . we and others have previously shown that in transient cotransfection , the two recombinant proteins are expressed by the same subset of cells , thus avoiding the risks associated with the use of stable clones or single cells ( brini et al . 1995 ) . the effect of bcl-2 expression on the cytosolic ca signal elicited by an agonist , atp , acting on receptors coupled , through gq proteins , to the production of inositol 1,4,5 trisphosphate ( ip3 ) was first investigated . in the experiment shown in fig . 1 , hela cells , either coexpressing bcl-2 and cytaeq ( bcl-2transfected ) or expressing only cytaeq ( controls ) were challenged with atp , which acts on a gq - coupled p2y receptor . both in control and bcl-2transfected cells , atp stimulation causes a rapid rise in cytoplasmic ca concentration ( [ ca]c ) , followed by a gradually declining sustained plateau . in bcl-2transfected cells , the [ ca ] increases evoked by stimulation with atp are significantly smaller than in controls ( peak amplitude 1.53 0.22 vs. 2.06 0.13 m , n = 10 ; fig . 1 a ) . since bcl-2 is localized in the mitochondria also , and thus could , at least in principle , enhance ca uptake in this organelle , a simple explanation for this result would be that cytosolic ca is more rapidly cleared by mitochondria , acting as an immobile buffer , placed in closed proximity to ip3-gated channels . indeed , bcl-2 overexpression does not increase , but rather reduces , the mitochondrial [ ca ] rise induced by atp ( peak amplitude 6.56 0.72 in controls vs. 4.42 0.46 m in bcl-2expressing cells ; n = 10 ) . interestingly , the reduction caused by bcl-2 overexpression appears larger in the mitochondria than in the cytosol , both in absolute terms and as percent ( 26 vs. 33% reduction , respectively ) . while not responsible for the alteration of the cytosolic ca signal , the effect of bcl-2 on mitochondrial ca homeostasis appears of major functional relevance , given that key events occurring in the mitochondrial matrix , such as the stimulation of atp production ( jouaville et al . 1999 ) and possibly the opening of the permeability transition pore , ptp ( bernardi 1999 ) , are regulated by [ ca]m . a second possible explanation for the reduction in the agonist - dependent [ ca]c ( and [ ca]m ) increases was next considered , i.e. , a decrease in the amount of ca released by the agonist - sensitive ca stores . to investigate this possibility , the [ ca ] in the lumen of the er was measured in cells cotransfected with bcl-2 and an eraeq chimera ( montero et al . in addition , since we recently have demonstrated that the golgi apparatus shares many of the ca homeostatic properties of the er ( pinton et al . 1998 ) , this latter organelle was also investigated . 2 shows the calibrated [ ca ] values in the two compartments . to obtain reliable quantitative estimates of the [ ca ] in the lumen of these two organelles , their [ ca ] needs to be decreased during both the reconstitution of aeq with coelenterazine and the subsequent initial phase of perfusion with krb / egta in the luminometer ( see materials and methods ) . under those conditions , the [ ca ] was < 10 m in both organelles . when the [ ca ] in the perfusion medium was switched to 1 mm , the [ ca ] in the lumen of the two compartments gradually increased , reaching in control cells plateau values of 452 46 m ( n = 7 ) in the er and 262 47 m ( n = 7 ) in the golgi apparatus , respectively . in bcl-2 transfected cells , lower steady state values were attained in both compartments ( 304 38 m , n = 7 for the er , and 186 34 m , n = 7 for the golgi apparatus , respectively ) . when the cells were treated with atp , rapid decreases in the [ ca ] of the two compartments the decrease of [ ca ] was larger and faster in controls compared with bcl-2expressing cells , reflecting the higher filling state and thus the more rapid flow through the ip3-gated channels . the data reported above differ substantially from those reported in wehi7.2 clones , where bcl-2 overexpression results in an augmentation of the er ca content ( lam et al . this discrepancy may be due to the use of transient vs. stable transfections , in which the results might reflect unique properties of the selected clone and/or the long - term expression of a foreign gene may result in adaptation phenomena that are only indirect consequences of the expressed protein . this interpretation might also explain the results obtained by kuo and coworkers in clones of mcf10a cells overexpressing bcl-2 , in which an increase in serca2 ( sarcoendoplasmic reticulum ca atpases 2 ) protein leads to accelerated ca uptake and enhanced ca loading ( kuo et al . 1998 ) . on the other hand , it is possible that , although the duration of these experiments is compatible with the time course of the apoptotic process , the events observed in transient experiments could include only some of the relevant effects of bcl-2 on the mechanisms controlling programed cell death . given that the steady - state [ ca ] in both organelles depends on the equilibrium between active accumulation and passive leak , the effect of bcl-2 could be on either process . on one hand , bcl-2 could reduce ca uptake by the serca , either by a direct effect on the pump or by reducing the resting cytosolic [ ca ] . on the other , it could increase the passive diffusion of ca from the er . to clarify this critical issue , we investigated these possibilities independently . at first , the possibility that bcl-2 affected primarily cytosolic [ ca ] ( and hence [ ca]er only indirectly ) was verified . given that aeq is not accurate enough to reveal small differences of [ ca ] 10 m , the resting [ ca]c was measured with the fluorescent indicator fura-2 ( grynkiewicz et al . to identify bcl-2overexpressing cells in single - cell imaging experiments , the cells were cotransfected with mitochondrially targeted gfp , mtgfp(s65 t ) ( rizzuto et al . gfp - positive cells were distinguished from controls by the typical fluorescence emitted upon illumination with blue light . no difference in the resting cytoplasmic [ ca ] between bcl-2transfected and control cells was observed ( data not shown ) . then , we investigated whether bcl-2 overexpression had any direct effect on the activity of the serca pump . for this purpose , the kinetics of ca accumulation were studied in permeabilized cells ( fig . 2 , bcl-2transfected and control cells were transferred to the luminometer chamber and perfused with a buffer ( ib , see materials and methods ) mimicking intracellular ionic conditions , supplemented with 100 m egta ( ib / egta ) . after permeabilization with 100 m digitonin added to the same medium and wash in ib / egta , the medium was switched to ib , containing 200 nm [ ca ] ( ib / ca ) . under those conditions , [ ca]er gradually increased , reaching , in control cells , a plateau value of 728 60 , n = 5 . in bcl-2transfected cells , the maximal rate of ca accumulation was the same as in control cells ( 13 2 vs. 12 3 m / s , n = 5 ) , arguing that the ca pumping activity is not modified . however , the steady - state value eventually attained was distinctly lower ( 504 52 m , n = 5 ) . therefore , these data exclude that the alteration of er ca homeostasis is due to a reduction in the activity of the sercas or to soluble cytosolic factors ( that are released into the medium after permeabilization with digitonin ) . rather , by demonstrating a difference only when er refilling approaches the high [ ca]er values of intact resting cells , i.e. , in conditions in which passive leak becomes significant , they suggest that bcl-2 acts on the latter process . we sought direct evidence for this interpretation by measuring the ca leak rate from the er of bcl-2expressing and control cells . after the depletion protocol , the er was first refilled by exposing the cells to extracellular [ ca ] ranging from 0.23 mm , thus resulting in different levels of steady - state [ ca]er . the serca blocker , 2,5-di-(tert - butyl)-1,4-benzohydroquinone ( tbubhq ) , was then added that initiates the release of stored ca . given that , by definition , the rates of ca uptake and leak in steady state are equal , the rate of [ ca]er decrease upon blockade of the serca must reflect the rate of ca cycling across the er membrane and , thus , of the leak rate at any given steady - state [ ca]er . b shows the relationship between [ ca]er and leak rate , which can be fitted by a power equation . in bcl-2overexpressing cells , the data are more scattered , but for each [ ca]er value , the rate of ca efflux is faster than in control cells . this result is consistent with the evidence that bcl-2 can induce cation channels of low selectivity in artificial lipid bilayers ( minn et al . previously , we have demonstrated that a reduction of 30% in steady - state [ ca]er results in a substantial activation ( > 50% ) of the so - called capacitative ca influx ( hofer et al . the reduction in steady - state [ ca]er caused by bcl-2 overexpression is thus expected to cause an activation of this pathway . however , indirect evidence ( namely , the observation that the resting cytosolic ca level is indistinguishable from controls ) suggests that this is not the case . to further investigate the problem , we measured the capacitative ca influx in bcl-2transfected and control hela cells expressing cytaeq ( fig . the cells were treated with the serca blocker ( 10 m tbubhq ) while perfused with ca - free medium ( krb / egta ) . this procedure first evoked a small transient increase in [ ca]c due to release of ca from the stores . when the release of stored ca was complete ( 3 min ) , ca was readded to the medium . this maneuver evoked a second , larger [ ca]c increase via the capacitative pathway , which was markedly lower in bcl-2transfected than in controls cells ( 1.19 0.11 in controls vs. 0.68 0.13 m in bcl-2expressing cells , n = 6 ) . the question then arises as to the mechanism(s ) through which bcl-2 downregulates capacitative ca influx . two possibilities were considered : one was that the reduction of this ca influx pathway is an adaptive mechanism to the long lasting reduction in steady - state [ ca]er ; the second was that the reduction is a direct consequence of bcl-2 overexpression . to distinguish between these two possibilities , we verified whether a long - term reduction of [ ca]er , obtained independently of bcl-2 , could cause a comparable downregulation of the capacitative ca influx . capacitative ca influx was thus compared in controls and in cells whose [ ca]er was reduced to values similar to those of bcl-2overexpressing cells by prolonged incubation with a low ( 100 m ) extracellular [ ca ] . in cells that had been maintained at low external [ ca ] , and thus experienced a long - term reduction in the resting [ ca]er levels , the [ ca]c peak evoked by activation of the capacitative ca influx was markedly lower than in controls ( 0.59 0.11 m , n = 5 ) . of interest , the level of steady - state [ ca]er in cells incubated in low extracellular ca was very similar ( 269 46 m , n = 4 ) to that measured in bcl-2overexpressing cells incubated in normal ( 1 mm ) extracellular ca concentration . the simplest explanation of these results is that the decrease in capacitative ca influx is an adaptive consequence to the prolonged reduction in steady - state [ ca]er , though , at the moment , it can not be excluded that other factors ( e.g. , a reduction in plasma membrane potential ) contribute to this effect . this reduction in capacitative ca influx , on one hand , appears critical to maintain a resting [ ca]c indistinguishable from that of control cells , despite a major reduction of [ ca ] in the er ; on the other hand , it could account , at least in part , for a higher resistance of cells expressing bcl-2 to apoptotic cell death , as the reduction of this ubiquitous ca influx pathway may prevent the overload of ca , considered one of the key events in apoptosis . in agreement with this view , ma and coworkers recently demonstrated that serca overexpression , by causing er ca overload , increases spontaneous apoptosis ( ma et al . finally , our data are not in contrast with previous results showing that the serca inhibitor , thapsigargin , activates programed cell death ( bian et al . 1997 ) . in this latter case , in fact , the ca depletion is acute ( 12 min ) and complete , whereas in bcl-2transfected cells , the drop in [ ca]er is modest and develops slowly . thus , thapsigargin not only causes a massive activation of the capacitative ca influx pathway , but also such a drastic reduction in the level of [ ca]er to interfere with the basic activity of er chaperonins . we have demonstrated that overexpression of bcl-2 causes a reduction of the steady - state [ ca ] levels , both in the er and in the golgi apparatus , the two main agonist - sensitive ca stores . we also showed that this alteration affects the ca signal to which the effector systems are exposed , in particular those located in local environments highly dependent on the filling state of the intracellular stores ( such as the mitochondria ) . the chronic reduction in [ ca ] of the er results in a downregulation of the capacitative ca influx . the alterations in ca handling provoked by bcl-2 overexpression could be part of the mechanisms through which bcl-2 exerts its role in the control of cell growth . targets of bcl-2 effects on ca homeostasis could be : the er itself , where sorting and trafficking of proteins , e.g. , enzymes or membrane receptors , are modulated by luminal [ ca ] ( booth and koch 1989 ) ; the mitochondria , where ca accumulation regulates the production of atp ( robbgaspers et al . 1998 ; the concentration of which is a key determinant of the apoptosis vs. necrosis choice , leist et al . 1997 ) , and , possibly , the release of the apoptotic cofactor cytochrome c ( krajewski et al . 1999 ) ; or the cytoplasm , where bulk or local [ ca ] increases activate enzymatic effectors , such as the ca - dependent protease , calpain ( carafoli and molinari 1998 ; squier et al . 1999 ) , or the phosphatase , calcineurin ( wang et al . 1999 ) . whether , and how , these routes interact , and how they are affected by the alteration in ca homeostasis caused by bcl-2 , awaits further investigation . Output:	the mechanism of action of the oncogene bcl-2 , a key regulator of the apoptotic process , is still debated . we have employed organelle - targeted chimeras of the ca2 + -sensitive photoprotein , aequorin , to investigate in detail the effect of bcl-2 overexpression on intracellular ca2 + homeostasis . in the er and the golgi apparatus , bcl-2 overexpression increases the ca2 + leak ( while leaving ca2 + accumulation unaffected ) , hence reducing the steady - state [ ca2 + ] levels . as a direct consequence , the [ ca2 + ] increases caused by inositol 1,4,5 trisphosphate ( ip3)-generating agonists were reduced in amplitude in both the cytosol and the mitochondria . bcl-2 overexpression also reduced the rate of ca2 + influx activated by ca2 + store depletion , possibly by an adaptive downregulation of this pathway . by interfering with ca2 + -dependent events at multiple intracellular sites , these effects of bcl-2 on intracellular ca2 + homeostasis may contribute to the protective role of this oncogene against programed cell death .
PubmedSumm2015	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the hospitalization rates and the social impact of diverticular disease in western countries have drastically increased . it is estimated that > 1 of 4 european citizens is affected by diverticula of the colon and therefore could potentially develop advanced clinical disorders associated with this condition . acute diverticulitis affects between 10% and 25% of people with diverticulosis , although these epidemiological data are very dated . the prevalence of diverticular disease is age - related , affecting about 60% of people over 80 years of age regardless of sex . in this sense , and in consideration of the progressive increase of the mean age of the population , diverticula - related diseases are a common cause of hospitalization . in addition to patients with acute complicated and uncomplicated diverticulitis , there are others who are affected daily by functional disorders associated with diverticular disease , with significant social costs and loss in productivity . the therapeutic request of the latter is to restore an acceptable quality of life . over the past 20 years the minimally invasive techniques allow us to obtain excellent results in the short term , in full compliance with the standard of care . the aim of this study is to analyze the effectiveness of laparoscopic sigmoid colectomy for the elective treatment of diverticular disease . this analysis was carried out through the evaluation of short- and long - term surgical outcomes and through the responses of patients by an appropriate validated questionnaire for calculating the perceived quality of life before and after the surgical procedure . our hypothesis is that the laparoscopic treatment can ensure both adequate postoperative results in full compliance with the standards of care and an increase in quality of life . a consecutive unselected series of 94 patients undergoing laparoscopic sigmoid colectomy at our division from january 1 , 2008 to december 31 , 2012 was analyzed . the indications for elective surgery were both multiple episodes of uncomplicated acute diverticulitis and single episodes of complicated acute diverticulitis with primary resolution by conservative treatment or laparoscopic drainage of abdominal abscess . all patients eligible for surgery were candidates for laparoscopic treatment . only those unsuitable from an anesthesiologic point of view or who had not given their consent to laparoscopy were treated by traditional surgery . preoperative contrast - enhanced computed tomography of the abdomen , colonoscopy , and barium enema were used in all cases . we collected patients- , surgery- and hospital stay related data , as well as the short- and long - term outcomes , as summarized in table 1 . comorbidity of each patient was assessed by charlson comorbidity index . from 2008 to 2011 , patients were subjected to mechanical bowel preparation using polyethylene glycol 70 + 70 g/2 l the day before surgery , whereas from 2011 , no patients underwent any bowel preparation . all patients were treated with both short - term broad - spectrum intravenous antibiotics ( ceftizoxime plus metronidazole ) and antithrombotic prophylaxis by low molecular weight heparin according to body weight . a nasogastric tube and a urinary catheter were placed after induction of general anesthesia in all cases . in selected cases we placed a left ureteral stent before starting the surgical procedure . all the operations were performed or supervised by the same team ; the team was fully trained in both colorectal and minimally invasive surgery . we used a 4-port medial - to - lateral standardized laparoscopic technique , as summarized in the following steps . surgical instrumentation was standardized , and dissection was performed by harmonic scalpel ( harmonic ace , ethicon endo - surgery , llc , guaynabo , puerto rico ) . prospective study data analyzed abbreviations : asa , american society of anesthesiology ; bmi , body mass index . the patient was placed in a classical lloyd - davis position with the left arm abducted and the right one along the body . the surgeon and the camera - holder stood on the right side of the patient and the assistant surgeon on the left . during the procedure , the operating table was tilted toward the right and ranged between trendelenburg and reverse trendelenburg positions depending on the operative steps . pneumoperitoneum was inducted to a pressure of 12 mm hg by the insertion of a 10-mm disposable hasson trocar about 5 cm cranially to the umbilicus at the midline . an accurate peritoneal inspection was carried out through a 30 camera to evaluate whether the case was suitable for laparoscopic surgery . then 3 trocars were placed : one 10/12-mm trocar in the right flank ( t2 ) and 2 10-mm trocars in the right lower quadrant ( t3 ) and in the left flank ( t4 ) , respectively . if we needed to mobilize the colonic splenic flexure , we started by opening the gastrocolic ligament with access to the lesser sac . the transverse mesocolon was dissected from right to left , exposing and preserving the capsule of the pancreatic tail . the left colonic angle was thus freed from splenic adhesions by the section of the splenocolic ligament . a lateral dissection completed the liberation of the splenic flexure . we continued then by an upward traction of the left colon sigma , which enabled a medial to lateral approach . we dissected the peritoneum in a caudal to cranial direction , and , after identifying both the inferior mesenteric artery ( ami ) and vein , we separated the white line of toldt from the gerota fascia under the arch of treitz , with right identification of the left ureter and gonadal vessels . ami and inferior mesenteric vein were isolated and dissected by 10-mm clips ( ligaclip , ethicon endo - surgery ) , respecting both the inferior mesenteric and the hypogastric plexus . the left sigmoid colon was laterally freed through the dissection of the left parietocolic ligament along the line of monk . the intestine was distally resected at the level of the upper rectus by an endoscopic linear stapler ( echelon flex 60 , ethicon endo - surgery ) with blue - load ( 3.5-mm 6 rows , ethicon endo - surgery ) . from that site , adequately protected ( steri - drape , 3 m healthcare , st . paul , minnesota ) , the left colon sigma was exteriorized with subsequent sigmoidectomy and insertion of the anastomotic anvil inside the proximal left colon . an end - to - end colorectal anastomosis using the knight - griffen technique was made by a transanal circular stapler ( endoscopic curved intraluminal stapler ils 29 mm , ethicon endo - surgery ) . the omentum was then relocated whenever possible to cover the anastomosis and finally both the port sites and minilaparotomy were synthesized for layers . all patients were mobilized early with removal of the urinary catheter , with the exception of those preoperatively suffering from colovesical fistula , in which the catheter was maintained for 7 days . criteria for the discharge included absence of symptoms , tolerance of a minimum of 3 meals without restrictions , and passage of stool . all adverse events that occurred within 30 days after surgery were considered complications . anastomosis was routinely checked by water - soluble contrast enema on postoperative days 6 or 7 . the term anastomotic leakage defines all conditions with clinical or radiological anastomotic dehiscence , with or without the need for surgical revision . the short- and long - term follow - up were conducted at 5 and 30 days after discharge and at 6 and 12 months after surgery , respectively . the qualitative long - term assessment was carried out by subjecting patients to the validated questionnaire gastrointestinal quality of life index ( giqli ) , via telephone or direct interview between 6 and 12 months after surgery , after obtaining proper consent . continuous and categorical variables were expressed as mean sd and as percentage value , respectively . the long - term qualitative assessment has been graphically represented with box - and - whiskers plot and compared using student t test . statistical analysis was performed using r ( version 2.15.1 ; the r foundation for statistical computing ) . institutional review committee approval was not required because the data of the present study were collected in the course of routine clinical practice and , therefore , are considered valid according to the informed consent signed by each patient for any surgery or other procedure . the study protocol conforms to the ethical guidelines contained in the world medical association declaration of helsinki ethical principles for medical research involving human subjects adopted by the 18th wma general assembly , helsinki , finland , june 1964 , then revised in tokyo 2004 . the patient was placed in a classical lloyd - davis position with the left arm abducted and the right one along the body . the surgeon and the camera - holder stood on the right side of the patient and the assistant surgeon on the left . during the procedure , the operating table was tilted toward the right and ranged between trendelenburg and reverse trendelenburg positions depending on the operative steps . pneumoperitoneum was inducted to a pressure of 12 mm hg by the insertion of a 10-mm disposable hasson trocar about 5 cm cranially to the umbilicus at the midline . an accurate peritoneal inspection was carried out through a 30 camera to evaluate whether the case was suitable for laparoscopic surgery . then 3 trocars were placed : one 10/12-mm trocar in the right flank ( t2 ) and 2 10-mm trocars in the right lower quadrant ( t3 ) and in the left flank ( t4 ) , respectively . if we needed to mobilize the colonic splenic flexure , we started by opening the gastrocolic ligament with access to the lesser sac . the transverse mesocolon was dissected from right to left , exposing and preserving the capsule of the pancreatic tail . the left colonic angle was thus freed from splenic adhesions by the section of the splenocolic ligament . a lateral dissection completed the liberation of the splenic flexure . we continued then by an upward traction of the left colon sigma , which enabled a medial to lateral approach . we dissected the peritoneum in a caudal to cranial direction , and , after identifying both the inferior mesenteric artery ( ami ) and vein , we separated the white line of toldt from the gerota fascia under the arch of treitz , with right identification of the left ureter and gonadal vessels . ami and inferior mesenteric vein were isolated and dissected by 10-mm clips ( ligaclip , ethicon endo - surgery ) , respecting both the inferior mesenteric and the hypogastric plexus . the left sigmoid colon was laterally freed through the dissection of the left parietocolic ligament along the line of monk . the intestine was distally resected at the level of the upper rectus by an endoscopic linear stapler ( echelon flex 60 , ethicon endo - surgery ) with blue - load ( 3.5-mm 6 rows , ethicon endo - surgery ) . from that site , adequately protected ( steri - drape , 3 m healthcare , st . paul , minnesota ) , the left colon sigma was exteriorized with subsequent sigmoidectomy and insertion of the anastomotic anvil inside the proximal left colon . an end - to - end colorectal anastomosis using the knight - griffen technique was made by a transanal circular stapler ( endoscopic curved intraluminal stapler ils 29 mm , ethicon endo - surgery ) . the omentum was then relocated whenever possible to cover the anastomosis and finally both the port sites and minilaparotomy were synthesized for layers . all patients were mobilized early with removal of the urinary catheter , with the exception of those preoperatively suffering from colovesical fistula , in which the catheter was maintained for 7 days . criteria for the discharge included absence of symptoms , tolerance of a minimum of 3 meals without restrictions , and passage of stool . anastomosis was routinely checked by water - soluble contrast enema on postoperative days 6 or 7 . the term anastomotic leakage defines all conditions with clinical or radiological anastomotic dehiscence , with or without the need for surgical revision . the short- and long - term follow - up were conducted at 5 and 30 days after discharge and at 6 and 12 months after surgery , respectively . the qualitative long - term assessment was carried out by subjecting patients to the validated questionnaire gastrointestinal quality of life index ( giqli ) , via telephone or direct interview between 6 and 12 months after surgery , after obtaining proper consent . continuous and categorical variables were expressed as mean sd and as percentage value , respectively . the long - term qualitative assessment has been graphically represented with box - and - whiskers plot and compared using student t test . statistical analysis was performed using r ( version 2.15.1 ; the r foundation for statistical computing ) . institutional review committee approval was not required because the data of the present study were collected in the course of routine clinical practice and , therefore , are considered valid according to the informed consent signed by each patient for any surgery or other procedure . the study protocol conforms to the ethical guidelines contained in the world medical association declaration of helsinki ethical principles for medical research involving human subjects adopted by the 18th wma general assembly , helsinki , finland , june 1964 , then revised in tokyo 2004 . patient- and disease - related data are listed in table 2 , and intraoperative and short - term results are summarized in table 3 . in all cases , the definitive histological examination of the specimen demonstrated the presence of diverticula in association with diverticulitis and peridiverticulitis , conditioning sometimes an inflammatory pseudotumor . we had 3 cases ( 3.2% ) of conversion to open laparotomy due to prolonged anatomical difficulties that did not allow us to safely continue the laparoscopic procedure . postoperative complications , classified according to the clavien - dindo system , are summarized in table 4 . we recorded 2 cases of reoperation ( 2.1% ) , one for a postoperative bleeding and the other for a fecal peritonitis from a nearly complete early anastomotic dehiscence . this latter case was fatal , hence the mortality rate of 1.06% . in the remaining discharged patients we observed no recurrence of diverticular disease during the clinical follow - up , although there was evidence of 3 cases ( 3.19% ) of incisional hernia . figure 1 shows the results carried out by the giqli validated questionnaire before the procedure and at follow - up of 9.6 2.7 months . the preoperative ( 97.1 5.8 ) and late ( 129.6 8.0 ) feedback data were statistically significant . complications according to clavien - dindo classification system box - whiskers plots show giqli scores before surgery and at follow - up . given the increased prevalence of colonic diverticular disease in the last decades , the surgical treatment of patients with diverticulitis in both elective and emergency situations has become an important topic of discussion . however , despite a considerable number of papers , the indications , choice of technique , and timing for surgery are still subject to debate , lacking level 1a evidence . laparoscopic surgery has revolutionized the treatment of many diseases of surgical interest . in 1991 , jacobs et al published the world 's first series of laparoscopic - assisted colorectal procedures wherein of 20 reported cases , 5 were laparoscopic sigmoidectomies for diverticular disease . since then , the minimally invasive treatment of diverticular disease has proved feasible and effective . laparoscopy minimizes postoperative pain and respiratory distress , reducing the length of hospital stay and improving the return to an active life . the largest published series of laparoscopic sigmoid colectomies for diverticular disease , > 500 , has shown excellent results in terms of conversion rate ( 2.8% ) , anastomotic leakage ( 1.4% ) , and overall mortality ( 0.2% ) . furthermore , the only meta - analysis available today , by siddiqui et al , showed a statistically better outcome for laparoscopic surgery compared with open surgery in terms of time of canalization , hospital stay , and morbidity rate . our results are consistent with those of other investigators in the literature , some of which are summarized in table 5 . the mean length of hospital stay is higher than that shown in some series , but not in all , as we routinely submit patients to a water - soluble contrast enema in postoperative days 6 or 7 . we use the enema to check for proper anastomotic transit and the absence of subclinical leakage . this assessment produces values well - correlated to the morbimortality , allowing us to objectively stratify the risk through the analysis of 19 tracer diseases , each of which contributes to the total score on a scale from 1 to 6 . although the mean value obtained ( 1.2 1.5 ) may seem rather low , it indicates a mild - to - moderate risk case group . in fact , a charlson score > 5 usually indicates a severe comorbidity , noting that in the original work by charlson et al a score > 3 was associated with a 10-year survival of 45% . body mass index is excluded from this assessment of comorbidity ; however , in our cohort , it was 26.2 kg / m , which is perfectly in line with the european population 's mean of 26.5 kg / m . however , according to the guidelines of the world health organization , these body mass indexes connote a condition of preobesity . in our experience , a body mass index > 25 is not a contraindication to laparoscopic surgery ; in fact , we firmly believe that obese people can benefit from minimally invasive colorectal surgery versus conventional procedures . elective laparoscopic treatment reports in the literature abbreviations : mnc , monocentric ; mtc , multicentric ; rct , randomized clinical trial . due to the inflammatory nature of the disease to be treated , the operative time is relatively higher compared with the time for the same surgery for cancer . moreover , the primary surgical goals are to completely remove the colonic segment affected by diverticula and to perform a totally tension - free anastomosis ; to do this it was necessary perform full mobilization of the colonic splenic flexure in > 50% of cases . mobilization of the left angle of the colon is a challenging laparoscopic step that results in a substantial increase in operating time . in order to identify a proper training pathway in laparoscopic colorectal surgery , jamali et al have collected the opinions of 35 experts in the field and then published a ranking of difficulty for 12 minimally invasive procedures of the lower digestive tract . their results showed a significant increase in difficulty between the simple sigmoidectomy and the sigmoidectomy with mobilization of the splenic flexure . the learning curve in laparoscopic colorectal surgery aims not only to achieve a technical proficiency but also to ensure adequate functional or oncological results . the breakpoint is estimated at around 50 to 60 procedures , as indicated by tekkis et al . although in our view , a proper attitude resulting from a progressive training in minimally invasive surgery could lower these numbers . this choice comes from the need to standardize the assessment of surgical outcomes . to date , there is no consensus on how to define and classify the severity of complications following surgery . often , the dichotomy between major and minor complications , without further explanations , is proposed . this raises the risk of assessing differently the results of surgery , with highly variable outcomes for similar procedures . to overcome these potential biases , in 2004 , dindo and clavien proposed a classification system that estimates the severity of complications depending on the resources used to solve them , and they tested it on a prospective cohort of 6336 patients undergoing elective surgery between 1988 and 1997 . after a single episode of acute uncomplicated conservatively treated diverticulitis , the probability of developing a second episode within 5 years is between 25% and 30% , with a proportional increase in the risk of diverticulitis complicated by free perforation and peritonitis . in addition , the rate of patients who remain symptomatic despite having no further hospitalizations after a first acute episode varies between 40% and 80% , resulting in an increase of the health and social costs . for some years , the guidelines from the american society of colon and rectal surgeons and the european society for endoscopic surgery have recommended an elective surgical treatment after 2 episodes of uncomplicated diverticulitis and after a single episode of complicated diverticulitis but these recommendations have been criticized by some investigators and are not currently included in the latest version of american society of colon and rectal surgeons guidelines . the current trend seems to prefer a tailored approach to each patient individually , assessing the medical history , the answer to the first acute episode , and the chronic symptoms . indeed , in 2010 , mkel et al published the review of a large20-year series of patients hospitalized for acute diverticulitis , and they concluded that 2 acute episodes would not justify an elective surgical treatment . but the epidemiological considerations by peppas et al on > 30,000 cases have estimated a hospital readmission rate of 18.6% in patients treated conservatively compared with 6.1% in patients undergoing surgery . patients in our series underwent laparoscopic surgical treatment after 1 or 2 episodes of uncomplicated diverticulitis in 78% of cases , as well as after a single episode in all cases of complicated diverticulitis . this confirms our current view , which is to undergo surgery after the first acute episode in patients under age 60 years and after 2 episodes in the remaining patients unless contraindicated otherwise . the indications for surgery are inspired by the observation that a growing number of acute episodes is directly proportional to both the conversion rate and the incidence of postoperative complications , whereas patients with < 3 acute episodes can effectively benefit from a minimally invasive surgical treatment . moreover , we believe that some factors such as lower comorbidity and the search for improvements in quality of life can motivate an earlier treatment in patients < 60 years . most of the literature that aimed to study the surgical treatment of diverticular disease almost exclusively analyzed the short - term outcomes , leaving for consideration the functional ones . the long - term outcomes are reported only by a few investigators and are contradictory . egger et al have reported that 25% of the 124 patients had bad functional outcomes symptoms such as painful constipation , abdominal distension , and diarrhea although the severity was not quantified . ambrosetti et al analyzed a series of 43 patients who underwent elective laparoscopic sigmoidectomy and concluded that minimally invasive surgery had given excellent functional outcomes , despite the 9.3% of patients who reported the occurrence of new symptoms of abdominal pain that the investigators then explained as a concomitant irritable bowel syndrome . raue et al reported that the quality of life evaluated by the european organization for research and treatment of cancer quality of life questionnaire ( eortc - qlq - c30 ) did not differ between open and laparoscopic technique , neither at 30 days nor at 12 months after surgery . the sigma trial interim results showed , after 6 weeks from the operation , a significantly better quality of life for the laparoscopic group , assessed by the sf-36 questionnaire ; this outcome decreased at the 6-month follow - up . a prospective assessment through the validated giqli questionnaire was made by forgione et al who analyzed both the pre- and postoperative results of 46 patients who underwent minimally invasive treatment for diverticular disease and concluded that laparoscopic sigmoidectomy produced statistically significant results in terms of short- and long - term quality of life . our experience confirms these latest results , with postoperative giqli mean values higher by > 20% compared with preoperative values , indicate a significant improvement of the quality of life of patients . the use of the giqli questionnaire was partially discussed and criticized because it evaluates not only the typical symptoms of the lower digestive tract but also more general symptoms . on the contrary , we believe that this feature defines the real indication for an overall assessment of performance status , psychological well - being , and quality of life after colorectal surgery . after surgical resection of the sigmoid colon and rectum , some patients report symptoms such as fecal incontinence , soiling , urgency , and difficulty to discriminate between liquids and gases . the etiology of pdd is not clear , but it could have multiple causes : an iatrogenic denervation of the left and sigmoid colon , a hypogastric plexus injury in the course of rectal mobilization , or a sphincter lesion caused by the insertion of the stapler . during laparoscopic sigmoidectomy , the incomplete mobilization of the rectum minimizes the hypogastric plexus injuries . however , some investigators suggest as a possible cause of pdd the potential injury of both the ascending fibers from the pelvic plexus and the descending fibers from the inferior mesenteric plexus . these lesions may be caused by a so , some investigators proposed ami preservation with contextual section of the arterial branches proximal to the sigmoid colon . although this topic requires further research leading to higher levels of evidence , our experience does not confirm the need for a distal arterial ligation . the results in terms of quality of life and bowel comfort stressed the effectiveness of a technique that involves ami ligation at this origin from the aorta , with careful identification of perivascular nerve plexus thanks to the magnification of the image given by laparoscopy . in addition , the high ligation allows precise identification and compliance with the anatomical fascial planes , minimizing any technical difficulties due to distorsion by previous flogistic episodes , and thus reducing drastically the conversion to laparotomy rate . the elective laparoscopic treatment of colonic diverticular disease is an effective and safe option that produces adequate postoperative results and ensures a satisfactory functional outcome for the patient . the obtained results confirm the appropriateness of both the choice of technique and the timing for surgery . Output:	background and objectives : to analyze the short- and long - term outcomes of laparoscopic sigmoid colectomy for the elective treatment of diverticular disease.methods:a consecutive unselected series of 94 patients undergoing elective laparoscopic sigmoid colectomy for diverticular disease from 2008 to 2012 was analyzed . we collected patients- , surgery- and hospital stay related data , as well as the short- and long - term outcomes . operative steps , instrumentation , and postoperative cares were standardized . comorbidity was assessed by charlson comorbidity index . complications were classified using the clavien - dindo classification system . the qualitative long - term assessment was carried out by subjecting patients to the validated gastrointestinal quality of life index questionnaire before and after surgery.results:the mean age of our cohort was 61.3 11.0 years with a charlson comorbidity index of 1.2 1.5.mean operative time was 213.5 60.8 minutes and estimated blood loss was 67.2 94.3 ml . we had 3 cases ( 3.2% ) of conversion to open laparotomy . the rates of postoperative complications were 35.1% , 6.3% , 2.1% , and 1.06% , respectively , for grades 1 , 2 , 3b , and 5 according to the clavien - dindo system . length of hospital stay was 8.1 1.9 days , and we have not recorded readmissions in patients discharged within 60 days after surgery . median follow - up was of 9.6 2.7 months . we observed no recurrence of diverticular disease , but there was evidence of 3 cases of incisional hernia ( 3.19% ) . the difference between preoperative and late gastrointestinal quality of life index score was statistically significant ( 97.1 5.8 vs 129.6 8.0).conclusions : elective laparoscopic treatment of colonic diverticular disease represents an effective option that produces adequate postoperative results and ensures a satisfactory functional outcome .
PubmedSumm2016	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: multidrug resistance ( mdr ) of cancer constitutes a significant challenge to clinical oncology and serves as one of the major reasons for tumor recurrence . cancer stem cells ( cscs ) were discovered in various types of cancer , including breast , colon , colorectal , lung , glioma , and pancreatic cancers . recently , cscs have been linked to the mdr since cscs are often enriched in tumors after conventional chemotherapies ; and cscs were found more resistant to conventional chemotherapeutics than differentiated tumor cells . mechanisms that cscs utilize to achieve drug resistance include entering a quiescent cell cycle status , possessing a high dna repair capacity , avoiding apoptosis , upregulating atp - binding cassette transporter , and overexpressing of detoxification enzymes such as aldehyde dehydrogenase 1 . the distinct susceptibility to conventional chemotherapeutics between cscs and differentiated tumors leads to enrichment of the csc populations in some tumors treated by conventional chemotherapeutics . what aggravates this alarming situation is cscs ability to recapitulate heterogeneity and hierarchy . therefore , cscs not only are one important reason for mdr but also play a critical role in progression and metastasis of cancer . consequently , a failure of eradicating cscs in tumors could have a huge adverse impact on the prognosis of cancer . due to this rationale , an effective way to elimination of cscs becomes critical to prevent mdr development and results in a favored prognosis during cancer treatment by conventional chemotherapeutics . because of the role of cscs in tumorigenesis and the mdr , there is an intensive , ongoing effort to discover new compounds that have selective toxicity to cscs . proposed mechanisms behind the selective toxicity of sali include p38 mapk activation , accumulation of reactive oxygen species , inhibition of wnt signaling pathways , and inhibition of p - glycoprotein activity by sali . consistent with its selective toxicity to cscs and the role of cscs in causing mdr , sali can sensitize a wide spectrum of tumor cell lines that would be otherwise resistant to conventional cancer chemotherapeutics such as paclitaxel ( ptx ) . while using sali to eradicate cscs may have immediate clinical benefits , such an application is currently hindered by poor solubility of sali , as well as its toxicities to healthy tissues and cells in mammals . on the other hand , nanocarriers have in general been shown to increase tumor accumulation of small molecule drugs , redistribute their accumulation in organs , and/or resolve their solubility issues . therefore , a nano delivery system is highly desired that can help sali overcome the solubility issue and increase its accumulation in tumors while reducing its distribution in off - target organs . recently , a hyaluronic acid based nanogel was shown in vitro to facilitate the uptake of sali to cscs , which are cd44 positive . the increased uptake compared to free sali was attributed to interactions between hyaluronic acid and cd44 . meanwhile , another group used peg - b - pcl polymeric micelles to deliver sali as encapsulated drugs and found that the micelles improved csc - specific toxicity of sali in vivo using a mouse subcutaneous , xenograph model of human breast cancer . however , the improvement was marginal as the overall therapeutic outcomes resulted from micelle - delivered sali and free sali treatments were indifferent statistically . we speculated that one reason for the aforementioned suboptimal therapeutic outcomes of sali may be that the micellar carrier failed to accumulate more sali in tumors than the free form of sali , and hence did not result in a stronger inhibition to tumor growth . the speculation can not be answered as the report did not include pharmacokinetics and tumor accumulation evaluations . here we seek to develop a novel and biocompatible nanoparticle ( np ) carrier to deliver sali to clinically and biologically relevant orthotopic breast tumors . we hypothesize that the np using sali as its hydrophobic , together with our new encapsulation method , can deliver satisfactory pharmacokinetics and tumor accumulation of sali , which in turn lead to a better therapeutic outcome of sali . to this end , we first obtained nps self - assembled from the conjugates between sali and itep , a biodegradable recombinant polypeptide we invented recently ( m. chen , personal communication ) ; the immune tolerance nature of itep may prove an advantage when the carrier is administered repeatedly and tested in clinical trials , in which cases the immunogenicity of therapeutics becomes a focal point of consideration . sali nps to encapsulate free sali , which is a novel strategy for sali delivery . then , we found that encapsulated sali was released in a controlled manner and maintained csc - selective toxicity of sali . we also discovered that coencapsulation of sali with two additives , -tocopherol and dmha , noticeably improved the loading efficiency and drug release profile of the sali . another benefit of the resulting nps is that they significantly retarded in vivo release of the sali and therefore led to better pharmacokinetics and tumor accumulation profiles of the sali . through tumor regression study , we demonstrated that the encapsulated sali was able to inhibit the growth of bulk tumors because of these desired properties conferred by the nps . lastly and most importantly , the np - delivered sali depleted cscs in breast tumors more efficiently than free sali . all chemicals , unless otherwise described , were purchased from thermo fisher scientific inc . organic solvents including acetonitrile ( acn ) , dichloromethane ( dcm ) , dimethylformamide ( dmf ) , isopropanol , and methanol were purchased from thermo fisher scientific inc . the cell culture media and supplements including rpmi-1640 ( with 2 mm l - glutamine ) , media 199 , and fetal bovine serum ( fbs ) were purchased from life technologies , inc . all antibodies were purchased from biolegend ( san diego , ca , usa ) including pe anti - mouse cd24 antibody ( cat . # 101808 ) , apc anti - human cd44 antibody ( cat . 4t1 , a highly metastatic murine cell line derived from a spontaneous syngeneic breast cancer of balb / c mice , was purchased from american type culture collection ( rockville , md , usa ) . 4t1 cells were maintained in monolayer cultures within an rpmi 1640 medium supplemented with 10% fbs . female balb / c mice that were 2428 days old ( 1819 g ) were purchased from charles river laboratories international , inc . all the animal experiment protocols were approved by the institutional animal care and use committee at the university of utah . iteps or itep sali conjugates were measured using the malvern zetasizer nano ( malvern , chester county , pa , usa ) at 25 m in pbs at 37 c using a low - volume disposable sizing cuvette or a clear disposable zeta cell . before size measurements , all samples were reduced overnight in a 100 mm tcep solution to reduce possible disulfide bonds . the instrument settings include the following : material ri = 1.59 , material absorption = 0.010 , water dispersant ri = 1.330 , viscosity = 0.6864 cp . the instrument was allowed to automatically optimize the count rate , duration , and attenuator . specifically , for zeta potential determination , the dispersant dielectric constant was set as 74.4 . 5.0 mg of itep sali nps and 3.0 mg of salinomycin were codissolved in 125 l of dmf and mixed well . in some cases , 13.34 mg of dmha ( sigma - aldrich , st . louis , mo , usa ) and 0.87 mg of -tocopherol ( sigma - aldrich , st . the ratio of the above components was based on our independent trial experiments and previous reports . the mixture was then supplemented with 750 l of milli - q water in a dropwise manner . the mixture was stirred for 0.5 h at room temperature before being dialyzed against 3 the complete removal of dmf was critical and was monitored using a characteristic absorption of dmf at 210 nm . the product solution was filtered through 0.45 m acrodisc syringe filters with supor membrane ( pall corporation , port washington , ny , usa ) and concentrated by amicon ultra-15 centrifugal filter units ( mw cutoff = 3000 da ) . the loading efficiency was defined as in the following equation : in vitro release profile of sali from itep sali nps was measured by a previously described dialysis method with minor modification . sali nps with or without encapsulated sali were diluted in 0.5 ml of milli - q water containing 4% bsa and kept in dialysis bags ( spectrum laboratories , inc . , rancho dominguez , ca , usa , mw cutoff = 8000 da ) . the bags were then immersed into 100 ml of pbs solution ( ph = 7.4 ) in a beaker , respectively . the entire dialysis systems were shaken at 100 rpm at 37 c . at predetermined time points , the 0.5 ml sample solutions in the bags were mixed and 10 l of sample was collected from each sample . sali inside the bags was presumed and measured as unreleased drug in the nps . the relationship of the percentage of sali release ( f ) with time ( t ) was fitted using the following equation and graphpad v5.0.k is the release rate constant : pharmacokinetics studies were carried out according a protocol modified from a previous report . sali nps or sali were injected in a 100 l solution into balb / c mice via the tail vein at 5 mg / kg free sali or encapsulated sali dose . the sali solution used for the injection contained 1% dmso to dissolve sali ( sigma - aldrich , st . about 50 l blood samples were collected by submandibular bleeding into tubes containing 150 l of pbs with heparin at 1,000 u / ml . blood plasma was collected from blood samples after samples were centrifuged to pellet blood cells . next , 100 l of isopropanol was added to each plasma sample to precipitate proteins . the supernatants were collected ; concentrations of sali in the supernatant were determined by the precolumn derivitization with dnph . the changes of plasma sali concentrations over time were fitted to a two - compartment model using graphpad v5.0 . the details are described below : using the graphpad , we obtained the initial concentration ( cp(0 ) ) , the distribution rate constant ( a ) , and elimination rate constant ( b ) directly . other pk parameters were calculated from the known parameters using following equations : for the biodistribution study , balb / c mice were inoculated subcutaneously with 10 4t1 cells in 50 l of pbs at # 4 mammary fat pad on the right side of mouse abdomen . sali np3s or sali was injected intravenously into the mice at 5 mg / kg sali or an equivalent sali dose , respectively . at 12 h postinjection , hearts , livers , spleen , lungs , kidneys , and tumors water solution on ice for 3 min using a brinkmann polytron homogenizer ( brinkmann instruments , inc . , next , 3 ml of the above abstract solution was used for liver tissues while 1 ml of the solution was used for heart , spleen , lungs , kidneys , and tumor tissues . after sali was extracted from the homogenized tissues into the solution , the samples were centrifuged at 14,000 rpm for 10 min at 4 c . supernatants containing sali were collected , and sali quantity was determined by precolumn derivitization with dnph . the final tissue distribution was normalized to percentage of initial dose averaged by weight of each organ ( % id / gram ) . 10 4t1 mammosphere cells or 10 regular 4t1 cells were seeded in wells of 96-well plates in 100 l of rpmi-1640 medium , which contained serially diluted sali , paclitaxel ( ptx ) , or the nps in tests for different studies . after the incubation with these drugs , live cells were quantified using the celltiter 96 aqueous one solution cell proliferation assay ( promega , madison , wi , usa ) . the plates were read with a spectramax m2 plate reader ( molecular devices , inc . , sunnyvale , quantities of live cells in each well were represented by the absorbance values at 490 nm of that well . consequently , cell viabilities in each treated well were expressed as the absorbance values in percentage after the values were normalized with the mean absorbance value of the wells containing untreated cells . the viability data was fitted into a sigmoidal dose response curve using the graphpad prism 5.0 ( graphpad software , inc . the ic50 and 95% confidence index ( ci ) were obtained from the fitting . balb / c mice were inoculated subcutaneously with 4t1 cells as described earlier . at the seventh day after inoculation and when tumors all reached to or above 100 mm , the mice were randomly assigned into three groups and treated with the control ( 1% dmso pbs treatment ) , free sali ( 5 mg / kg in 1% dmso pbs ) , or itep the length and width dimensions of tumors were measured by a caliper every other day . tumor volumes were estimated using the following formula : tumor volume = ( length width/2 ) . the mice were sacrificed the day after the fifth treatment . the tumors were harvest to determine their weight and csc populations . single cell suspensions of cells were prepared from 4t1 tumor tissues that we collected from the tumor growth study according to a previously published protocol . an approximately 300 mg of tumor mass was incubated with 3 ml of medium 199 ( life technologies , inc . ) with 250 u / ml ultrapure collagenase iii ( worthington biochemical , inc . , lakewood , nj , usa ) . the tissue digestion lasted for 2 h at 37 c while shaken at 100 rpm . after digestion , 3 ml of serum - containing m-199 medium was added to inhibit the activity of collagenase , and cells were filtered through a 45 m nylon mesh and washed with rpmi-1640 containing 10% fbs . cells were counted , transferred to a 5 ml tube , and washed twice with dpbs with a 0.1% heat - inactivated fbs at 4 c for 5 min at 1,000 rpm . antibodies were then incubated with the cells for 30 min on ice at a 1:80 dilution as recommended by the manufacturer of the antibodies . after the incubations , cells were washed twice with the above washing medium and then were resuspended in 0.4 ml of dpbs/0.1% fbs containing 1 g / ml dapi per million cells . flow cytometry analyses of the cells were performed on a cytek dxp ( cytek development , inc . , the gating for cscs was set up based on the cd44 and cd24 expression profile of the cells collected from 4t1 mammospheres . to study mammosphere - forming ability of tumors , the above - mentioned single tumor cells were seeded in wells of 6-well cell culture plate at a density of 500 cells / well . the culture medium was supplemented with b27 ( 1x , invitrogen , ca , usa ) , 20 ng / ml egf , 20 ng / ml bfgf ( bd biosciences , franklin lakes , nj , usa ) , 4 g / ml heparin ( sigma - aldrich , mo , usa ) , and 0.5% methylcellulose ( stem cell technologies , vancouver , bc , v5z 1b3 , canada ) . after 7 days of culture , number of mammospheres in each well was counted and analyzed . to design a nanocarrier to deliver sali to solid tumors through the enhanced permeability and retention ( epr ) effect , we took advantage of a self - assembling principle of amphiphiles that amphiphlies with strong , spatially separated amphiphilicity are able to assemble into micelle - like nps . since sali is hydrophobic with a logd value of 3.24 at ph 7.4 , we paired it with a hydrophilic itep that we recently invented ( chen , personnel communication ) . we expected that attaching sali to one terminus of the itep would introduce sufficient , segmented amphiphilicity and the amphiphilicity can drive the conjugates to form nps . to this end , 32 cysteines were appended to the c - terminal of the itep as conjugation sites ( figure s1 in the supporting information ) . the resulting new itep ( figure s2 in the supporting information ) maintains the hydrophilic nature of the parent itep ( figure s3 in the supporting information ) . to connect sali to the new itep , sali was first connected with the mpbh linker ( figures s4 and s5 in the supporting information ) ; then the formed mpbh sali conjugates were connected with the new itep ( see the supporting information ) . after itep sali conjugates were purified , we found that the conjugates displayed a hydrodynamics diameter of 109.7 45.9 nm at 37 c according to dls measurements ( figure 1a ) . the size of the conjugates was much larger than that of the unconjugated itep , 7.9 1.49 nm , which suggests a np structure of itep sali conjugates . sali nps are narrowly dispersed with a low polydispersity index ( pdi ) value of 0.217 . the nps have a critical micelle concentration ( cmc ) of 15.32 m and a neutral charge ( figures s6 and s7 in the supporting information ) . the nps were not toxic , due presumably to the nonrelease of sali from the nps ( figure 1b ) . the nonrelease result is reasonable since sali is connected with mpbh through a stable amide bond and the mpbh , in turn , is connected to the iteps through a stable thioester bond . ( a ) hydrodynamic diameters of the unconjugated itep which has a 32-cysteine appendix ( blue filled area ) and itep sali conjugate ( black filled area ) . , the unconjugated itep was reduced in 100 mm tcep solution overnight to cleave possible disulfide bonds . ( b ) viability profiles of regular 4t1 cells after they were exposed to different concentrations of sali or itep sali nps for 48 h. the ic50 of free sali is 4.8 m . the nps do not show any toxicity up to 100 m . since we have attained the itep sali nps , next we tried to use the nps to deliver sali in an encapsulated form . using a modified cosolvent method , we were able to load free sali into the nps . sali np2s have a mean diameter of 195.0 95.7 nm and a pdi of 0.288 ( figure 2a ) . next , through an in vitro release study , we found that the encapsulated sali was released from itep sali np2 at a fairly rapid release rate with a half - life of 1.0 h ( figure 2b and table 1 ) . ( b ) the release profile for sali from tep - sali np2 , itep the quantity of released sali was determined by hplc in combination with a precolumn derivatization with dnph . the sali release data were fit to a release model described in the supporting information . ( c ) the loading efficiency of sali by np2 , np2 plus dmha , or np3 . the * indicates that the differences between the np3 and np2 or np2 plus dmha are statistically significant with a p < 0.0001 analyzed by one - way anova . ( d ) viability profiles of 4t1 mammosphere cells after they were exposed to different concentrations of sali or itep sali np3 for 48 h. the ic50 of free sali is 0.49 m with a 95% ci = 0.400.60 m . the ic50 of the np3 is 0.48 m with a 95% ci = 0.350.59 m . because sali has an ionizable carboxyl group with a pka around 4.0 that could confer negative charges to sali at a neutral ph , we suspected the negative charges may have rendered instability to sali encapsulation in hydrophobic cores of itep the instability , in turn , may have led to the rapid release and low loading efficiency of sali we observed . in order to test the conjecture and to attain a carrier having a slower release rate and higher loading efficiency of sali , we coencapsulated sali with dmha , a positively charged hydrocarbon that has been used to neutralize encapsulated , negatively charged chemicals inside micelles . the results from our in vitro release study showed that coencapsulation of dmha and sali slowed down the release of sali from the np2s . the release half - life of sali from the np2 with dmha was 2.9 h , which is significantly longer than that of the np2s without dmha ( figure 2b and table 1 ) . the resulted loading efficiency of 30.2 6.8% was same as that of the np2s without dmha ( figure 2c ) . one early study pointed out that dmha was not as effective as its more hydrophobic analogue , n , n - dimethyloctadecylamine ( dmoa ) , in stabilizing the encapsulation of negatively charged chemicals . dmoa , however , was more toxic than both dmha and sali ( figure s8 in the supporting information ) , which excluded it from serving as an additive for the sali coencapsulation . we reasoned that , to exploit the coencapsulation system of dmha and sali , we might need to supplement more hydrophobicity to it . therefore , we introduced a second additive , -tocopherol ( vitamin e ) , to the system . the -tocopherol was able to double loading efficiency of rapamycin due to its ability to increase the hydrophobic core of micelles . we hypothesized that coencapsulation of -tocopherol together with sali and dmha would not only provide a sufficiently hydrophobic environment for sali but also neutralize the charge of sali . this would mean that sali can be trapped more stably inside of the hydrophobic core of the nps . consequently , we could see an improvement of both loading efficiency and release profiles of the coencapsulated sali . the hypothesis was proven true as the coencapsulation led to a very high loading efficiency , 75.0 6.17% ( figure 2c ) . in addition , the newly generated nps , including sali and two additives ( termed itep sali np3s ) , have a mean diameter of 179.9 43.0 nm and share a size similar to that of np2 ( figure 2a ) . importantly , the release rate of sali from the np3s is remarkably slower than that of np2s or np2s with dmha only ( figure 2b ) . the release half - life of np3 was 4.1 h , which is significantly longer than 2.9 h of the np2s with dmha only or 1.0 h of the np2s ( table 1 ) . together , these results showed that the paired additives , dmha and -tocopherol , improved both loading efficiency and release kinetics of encapsulated sali in itep sali nps . sali np3 solubilized sali in aqueous solution , rendering a maximum solubility of 7,320 230 g / ml , which represents a 430-fold increase , given that the intrinsic solubility of sali is only 17 g / ml . lastly , the results of the in vitro cytotoxicity study showed that itep sali np3s possess the same level of selective toxicity to cscs as free sali ( figure 2d ) . here , we used cells harvested from 4t1 tumor mammospheres as a model for cscs because these cells carry characteristic phenotypes of cscs , such as a cd24/cd44 phenotype and a mammosphere - forming capacity ( figure s9 in the supporting information ) . the ic50 of the np3 to 4t1 mammosphere cells was 0.48 m , which , statistically , was the same as the ic50 of free sali , 0.49 m ( figure 2d ) . this similarity is understandable as we expect that the encapsulated sali is released from the np3s during the incubation with the cells and exerts its toxicity the same as free sali . sali np3s was administered to mice at a dose of 5 mg / kg intravenously , plasma concentrations of free sali or encapsulated sali were sampled up to 12 h ( figure 3a ) . these concentration data fit best into a two - compartment pharmacokinetics model . according to the fitting results ( table 2 ) , the auc of the np3-delivered sali was 192.10 m h , which was approximately 6 times greater than that of the free sali , 30.70 m h. similarly , free sali s clearance was 6 times faster than that of the encapsulated sali ( clsali , 216.87 l/(h g ) versus clnp3 , 34.66 l/(h g ) ) . the elimination half - life of the np3-delivered sali was approximately 5.2 times longer than that of the free sali . meanwhile , in summary , the results support a conclusion that the np3s significantly retard the clearance of the encapsulated sali from the systematic circulation , which , together with sizes of the np3s ( 179.9 nm in diameter ) , might lead to greater tumor accumulation of sali via the epr effect . indeed , there was a 2.4-fold increase of tumor sali accumulation resulting from using the np3s . at 12 h postinjection , the mean sali concentration in tumors of those mice receiving free sali was 1.17 0.17% id / g . in contrast , the mean concentration for the np3-delivered sali was 2.85 0.12% id / g ( figure 3b ) . at the same time , the np3s reduced sali accumulation in heart and lungs ( figure s10 in the supporting information ) . ( a ) plasma sali concentrations after they were administered as free form ( blue line ) or as encapsulated form in itep sali np3 ( red line ) through intravenous injection . the concentrations were determined by hplc in combination with a precolumn derivatization with dnph . the sali plasma concentration was plotted on the log scale based on 10 as a function of time postinjection . ( b ) tumor accumulation of sali after they were administered as free form ( blue bar ) or as encapsulated form in itep sali np3 ( red bar ) through intravenous injection . the presented data are for tumor samples collected at 12 h post intravenous injection at 5 mg / kg . the quantities of sali were expressed as percentage of initial dose normalized by weight of tumor , % id / gram . the * indicates that the difference between sali and itep sali - np3 is statistically significant with a p = 0.0031 analyzed by one - way anova . since itep sali np3s extended the systematic circulation of sali and increased tumor accumulation of sali , it was intriguing to see whether such improvements could strengthen the selective inhibition of cscs by sali in vivo . to test this hypothesis , we inoculated murine 4t1 syngeneic tumor cells in mammary fat pads of balb / c mice to establish an orthotopic tumor model . once the tumors grew to desired sizes , we treated the mice with 5 doses of pbs as a control , 5 mg / kg free sali , or itep after 5 treatments , we analyzed csc frequencies in these tumors using cd24 and cd44 markers . the cscs in tumors were defined operationally as those cells that have the same cd24 and cd44 expression profiles as those cells collected from 4t1 mammospheres ( figures s9 and s11 in the supporting information ) . for the control group , free sali was able to lower the frequency significantly to 64.32 5.72% ( p < 0.05 ) . more importantly , the nps further reduced the value to 55.62 3.73% , which was significantly lower than those of both the control group and free sali - treated group ( p < 0.05 ) ( figure 4a ) . these comparison results suggest that sali is able to kill cscs in vivo just as they are able to in vitro ( figure 2b ) , and , more significantly , itep ( a ) the csc frequencies of the 4t1 orthotopic tumors treated by controls ( pbs ) , free sali , or itep sali np3 . the * symbols indicate that the mean css frequency of the np - treated tumor is significantly lower than that of control- or free sali - treated tumors ( p = 0.0004 and p = 0.0252 , respectively ) , and that the mean frequency of the sali - treated tumors is significantly lower than that of the control - treated tumors ( p = 0.0133 ) . ( b ) mammospheres formed from 4t1 orthotropic tumors after these tumors were treated by controls ( pbs ) , free sali , or itep the * symbols indicate that the mammosphere - formation ability of the np - treated tumor is significantly lower than that of control- or free sali - treated tumors ( p = 0.0035 and p = 0.0125 , respectively ) , and that the mammosphere - formation ability of the sali - treated tumors is significantly lower than that of the control - treated tumors ( p = 0.0358 ) . ( c ) the viability profile of regular 4t1 cells versus 4t1 mammosphere cells after they were exposed to different concentrations of ptx for 72 h. the ic50 of ptx to regular 4t1 cells 5.79 nm with a 95% ci = 4.906.86 nm . the ic50 of ptx to 4t1 mammosphere cells 40.48 nm with a 95% ci = 37.6447.30 nm . the capacity is relevant to the drug resistance of tumors , thus they are very significant to cancer therapy . to investigate functional significance of the np3s effect in reducing cscs in 4t1 orthotopic tumors , we examined mammosphere - forming ability of the cells collected from 4t1 tumors after these tumors were treated by the np3 , free sali , and pbs . we found that the np3-treated tumors had a significantly lower ability to generate mammospheres than the other two treatment groups ( figure 4b ) . there were on average 42.2 mammospheres formed per 500 np3-treated tumor cells , which were significantly less than those formed from free sali- or pbs - treated tumors ( 54.7 and 61.0 mammospheres per 500 tumor cells , respectively ) . putting together , the superior effects of the np3 over free sali in eliminating cscs and impeding the mammosphere - forming ability of 4t1 tumors are significant because they suggested that the np3 was effective to alter cell composition of the tumors , and , more importantly , to diminish tumorigenesis and drug resistance potential of the tumors . in a separate study , we found that the cells isolated from mammospheres were more resistant to ptx than general 4t1 cells cultured as monolayers ( figure 4c ) . the ic50 of ptx to the mammosphere cells was 7 times higher than the ic50 to regular 4t1 cells , 40.50 nm versus 5.79 nm ( p < 0.05 ) . the results are consistent with earlier reports that cscs play a critical role in promoting the tumor mdr . therefore , the aforementioned enhancing effect of itep sali nps to the csc - specific toxicity of sali may have implications for overcoming the mdr . since cscs play a role in tumorigenesis and maintain the hierarchy of tumors , depletion of cscs in tumors may lead to a tumor regression or stabilization . thus , we examined whether the superiority of the np3-delivered sali over free sali in depleting cscs can translate into an advantage of tumor growth inhibition or tumor regression . our results showed that neither the np3s nor free sali were able to abolish tumors or to stop tumor growth ( figure 5a ) . the growth - inhibition effect was especially prominent for the nps as the mean tumor weight of the np3-treated mice at day 16 was significantly smaller than that of the control group : 436.8 137.9 mg versus 762.6 193.8 mg ( p < 0.05 ) , respectively ( figures 5b and 5c ) . while it is clear the np3-delivered sali had an impact on overall tumor growth , the impact was not as potent as its impact on cscs inside the 4t1 tumors . in contrast to the result that the np3-treated 4t1 tumors had a significantly smaller csc population than free sali - treated 4t1 tumors after five treatments ( figure 4a ) , the mean weights of the np3-treated and the free sali - treated tumors were not statistically different ( figure 5c ) . one possible reason for the discrepancy is that cells in 4t1 tumors might , on average , be less sensitive to sali than the csc subpopulation in the tumors . indeed , our results showed that sali was 10 times less toxic to regular 4t1 cells than to cells isolated from 4t1 mammospheres ( figure 5d ) . thus , it is possible that , as a result of lower toxicity of sali to differentiated tumor cells and stromal cells , the increased sali accumulation did not have the same degree of impact on the overall tumor growth as on cscs . in addition , the np3s did not completely eradicate the csc subpopulation in the tumors . the surviving cscs could reignite tumor growth that counteracted any tumor growth inhibition contributed by the np3s . as a result ( a ) the size changes of 4t1 orthotopic tumors after they were treated by five doses of controls ( pbs ) , free sali , or itep sali np3 . both free sali and the np - encapsulated sali were administered at 5 mg / kg . the tumor dimensions were measured at the same day of dosing except for the last measurement , which was performed at day 16 post tumor inoculations . ( b ) photos of the tumors collected at day 16 after the inoculations . ( c ) the mean tumor weights of each group of mice at the day 16 post tumor inoculations . the * indicates a significant difference between the connected two means ( p = 0.0307 ) . ( d ) the viability profile of regular 4t1 cells versus 4t1 mammosphere cells after they were exposed to different concentrations of itep sali np3 for 48 h. the ic50 of the np3 to regular 4t1 cells 4.56 m with a 95% ci = 3.495.95 m . the ic50 of the np3 to 4t1 mammosphere cells 0.48 m with a 95% ci = 0.350.59 m . in this study , we designed and characterized an itep - based sali delivery system that improved the pharmacokinetics and tumor accumulation of sali , as well as strengthened its specific csc - inhibition effect of sali . the results , in principle , validate the use of nanocarriers to boost the efficacy of csc - specific drugs . because cscs were suggested to play a role in causing tumor mdr and recurrence , this enhancement effect of nanocarriers represented by itep iteps were chosen as building blocks for the delivery system because they bring three primary advantages to the system . first , the nonimmunogenic nature of iteps could save the carrier from the risk of any carrier - specific immunogenicity and allow multiple dosing of the carrier . in contrast , if a pharmaceutical protein or polypeptide induces immune responses during its application , the responses could severely compromise its function . second , recombinant polypeptides such as iteps are biodegradable so the carriers built upon the polypeptides are biodegradable and biocompatible as well . indeed , itep carriers have not revealed any adverse effect in our in vivo studies . finally , the sequences , length , and hydrophobicity of iteps can be definitely controlled and adjusted using genetic engineering approaches . this property of iteps becomes an advantage since it allows for the generation of a wide spectrum of new iteps or itep nps to precisely meet a specific delivery need . the versatility of itep carriers offers the possibility to further optimize delivery systems not only for sali but also for other csc - targeting drugs . examples of such optimization include appending itep with protein - based targeting ligands such as those antibodies recognizing csc - specific antigens . the sali delivery system reported here represents a novel strategy to deliver and use sali . sali contributed the hydrophobicity to the itep sali conjugates and served as payloads at the same time . previously , hydrophobic drugs were used as a hydrophobic component of micelle - forming amphiphilic conjugates and an encapsulated payload in the micelle . what we are reporting is the first application of this delivery strategy to sali . a stable encapsulation represents the primary advantage of this strategy , which happens when a hydrophobic drug is packed inside a hydrophobic core contributed by the same drug . when charged hydrophobic drugs such as sali are trapped inside a hydrophobic core of micelle - like nps , the charges may destabilize the encapsulation , which was observed in both previous studies and this current study . to cancel out the charges , it is reasonable to coencapsulate an oppositely charged , hydrophobic molecule with the charged drugs . dmoa was believed to be a good candidate to serve as coencapsulate additives for negatively charged drugs such as sali , because it is hydrophobic and positively charged . however , our data revealed strong toxicity of dmoa , especially in the comparison to sali , which excluded dmoa for the coencapsulation with sali . dmha , a positively charged analogue of dmoh with no apparent toxicity , is a reasonable alternative . however , dmha is not as hydrophobic as dmoa due to its short hydrocarbon chain . to address the insufficient hydrophobicity of the sali dmha coencapsulation system , we tried using a second hydrophobic chemical to supplement more hydrophobicity . we observed improvements in both the loading efficiency and the release profile of sali when dmha and -tocopherol were used in combination . we believe that this innovative , dual - additive approach may find its application in many encapsulation - based delivery systems , given that there could be many cases where the drugs to be encapsulated do not have an ideal charge . sali nps , has a higher accumulation in the tumor and a longer system circulation than those of free sali . consistent with these advantages , itep sali nps deplete cscs and decrease mammosphere - forming capacity of the 4t1 tumors more efficiently than free sali . however , the nps do not inhibit tumor growth more effectively , a result consistent with a previous report that was obtained by using a xenograph tumor model . in fact , neither the nps nor free sali stabilized or abolished tumors . there are at least two possible reasons for the discrepancy between the csc eradication effect and the overall tumor inhibition effect of the nps . first , itep sali nps did not eradicate cscs completely , a deficiency that may contribute to tumor re - regrowth . the regrowth , in turn , may have canceled out some tumor inhibition effect of the nps . given this possibility , it is important to develop new carriers to expand the advantage of the sali carrier in accumulating sali in tumors . the carriers that could meet this need may be either a carrier that offers more stable encapsulation and slower release or a carrier that recognizes tumor cells through its targeting moieties . the second reason stems from the sali s toxicity characteristics , its higher toxicity to cscs compared to differentiated cells in tumors . so , it is reasonable to find that cscs , rather than bulk tumors , respond well to the increased sali accumulation caused by the itep the toxicity characteristics advocate for a combination therapy that , concurrently or sequentially , uses the nps together with conventional chemotherapeutics such as ptx . the combinational therapy promises to target all cell types in tumors , and hence is able to either stabilize or abolish the tumors . sali nps is their application in preventing tumor recurrence or mdr , given that the nps have been shown to strengthen the toxicity of sali to cscs and that cscs have been shown as one factor driving the tumor mdr . Output:	salinomycin ( sali ) has selective toxicity to cancer stem cells ( cscs ) , a subpopulation of cancer cells that have been recently linked with tumor multidrug resistance ( mdr ) . to utilize its selective toxicity for cancer therapy , we sought to devise a nanoparticle ( np ) carrier to deliver sali to solid tumors through the enhanced permeability and retention effect and , hence , to increase its exposure to cscs . first , hydrophobic sali was conjugated to a hydrophilic , immune - tolerant , elastin - like polypeptide ( itep ) ; the amphiphilic itep sali conjugates self - assemble into nps . next , free sali was encapsulated into the nps alone or with two additives , n , n - dimethylhexylamine ( dmha ) and -tocopherol . the coencapsulation significantly improved the loading efficiency and release profile of sali . the resulting nps of the coencapsulation , termed as itep sali np3s , have an in vitro release half - life of 4.1 h , four times longer than itep sali np2s , the nps that have encapsulated sali only . further , the np3 formulation increases the plasma area under curve and the tumor accumulation of sali by 10 and 2.4 times , respectively . lastly , these improved pharmacokinetic and tumor accumulation profiles are consistent with a boost of csc - elimination effect of sali in vivo . in np3-treated 4t1 orthotopic tumors , the mean csc frequency is 55.62% , a significant reduction from the mean frequencies of untreated tumors , 75.00% , or free sali - treated tumors , 64.32% . the csc - elimination effect of the np3 can further translate to a delay of tumor growth . given the role of cscs in driving tumor mdr and recurrence , it could be a promising strategy to add the np3 to conventional cancer chemotherapies to prevent or reverse the mdr .
PubmedSumm2017	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: febrile seizure is the most common seizure disorder in childhood.12 its cumulative incidence is approximately 3% , but it is quite varied in nigeria with values ranging from 10 - 18%.17 the international league against epilepsy ( ilae ) defines febrile seizure as a seizure occurring in childhood after one month of age , associated with a febrile illness not caused by an infection of the central nervous system , without previous neonatal seizures or a previous unprovoked seizure , and not meeting criteria for other acute symptomatic seizures.8 it is often characterized by a generalized tonic - clonic convulsion lasting less than 15 min , fever of non - central nervous system infection with full regaining of consciousness after the convulsion and occurring commonly in children between 3 months and 5 years.12 there are two types of febrile seizures : the simple and complex types.138 the simple type is characterized by an episode of generalized tonic - clonic seizure lasting less than 15 min in 24 h while in the complex type the convulsions are multiple , lasting more than 15 min and associated with focal findings after the ictal period . while the majority of febrile seizures are simple ( 70 - 75% ) , 9 - 35% of them are complex.38 a positive family history for febrile seizures can be elicited in 25 - 40% of patients with febrile seizures.8 studies also show a high concordance rate in monozygotic rather in dizygotic twins.8 the febrile seizure gene has been mapped to chromosome 19p and 8q13 - 21.1 in addition to the positive history in a first or second degree relative , occurrence of febrile seizure has been associated with : human herpes virus-6 infection,9 influenza viral infection10 and iron deficiency anaemia.11 the overall recurrence rate is 30%.38 predictors of recurrence include : complex seizures , positive family history , onset at less 12 months and temperature < 40 c.138 management is achieved through controlling fever by tepid sponging or giving antipyretics , aborting ongoing convulsion with an anticonvulsant and treatment of the underlying cause of fever . prognosis is usually good even though a low proportion ( 2 - 4% ) go on to develop epilepsy.12 however , mortality can arise from poor or unorthodox management.7 kaduna is in northwestern nigeria a region with high under - five mortality rate in the country and , where care givers have demonstrated inappropriate management of febrile illnesses in children.1315 the aim of the study was to review the current status of febrile seizures in kaduna metropolis . we reviewed hospital records of 635 children to determine the rate and descriptions of febrile seizures among admissions in the department of paediatrics , 44 nigeria army reference hospital ( narh ) , a tertiary health facility in kaduna , between june 2008 and june 2010 . all who had seizures or seizure related disorders were reviewed for history of fever , type of seizure , level of consciousness using the glasgow coma scale ( gcs),16 outcome of cerebrospinal fluid ( csf ) analyses , urea and electrolytes , random blood sugar , blood film for malaria parasitemia , full blood count , electroencephalogram ( eeg ) and cultures of throat swab , urine , stool and blood , where indicated . other parameters reviewed were age , sex , social class , actions taken by parents and outcome of hospitalization . social class classification was that of the parents and based on oyedeji 's classification17 and utilized by akpan et al,18 ; grade i- professionals , senior public servants , large scale businessmen ; grade ii - intermediate civil servants , senior school teachers ; grade iii - junior school teachers , drivers and artisans with secondary education ; grade iv - petty traders , laborers , messengers and grade v - unemployed , students , and subsistence farmers . those reviewed fulfilled the following criteria for febrile seizures12 : had a history of fever associated with the onset of seizurehad seizures described as generalized tonic - clonic convulsion , an episode in 24 h lasting less than 15 min ( simple febrile seizure ) or greater than 15 min or multiple convulsions in 24 h ( complex febrile seizure)non - central nervous system infection as indicated by absence of bacterial isolates in the csf or the combination of pleocytosis , reduced csf glucose and elevated csf protein level19full regaining of consciousness after seizure as indicated by a normal gcs . had a history of fever associated with the onset of seizure had seizures described as generalized tonic - clonic convulsion , an episode in 24 h lasting less than 15 min ( simple febrile seizure ) or greater than 15 min or multiple convulsions in 24 h ( complex febrile seizure ) non - central nervous system infection as indicated by absence of bacterial isolates in the csf or the combination of pleocytosis , reduced csf glucose and elevated csf protein level19 full regaining of consciousness after seizure as indicated by a normal gcs . discrete variables were further assessed for significance using chi - square and fisher 's exact where applicable . discrete variables were further assessed for significance using chi - square and fisher 's exact where applicable . out of 635 cases , 79 ( 12.4% ) children had seizures or seizure related disorders [ table 1 ] . of these 17 , 21.5% of the seizure related disorders and 2.7% of the total cases , fulfilled the criteria for febrile seizures . meningitis seen in 33 children was the commonest ( 41.8% ) among the seizure related disorders . disease conditions among 79 patients with seizure related disorders there were 11 males and 6 females ( m : f , 1.8:1).there was a male preponderance . the mean age was 2.2 years1.1 with a range of 9 months to 5 years and a peak age of 3 years . twelve ( 70.6% ) were in the upper social classes ( i - iii ) while the others were in the lower classes ( iv and v ) . age and sex distribution of 17 patients with febrile seizures all the 17 eligible children presented with fever and convulsions [ table 3 ] . other symptoms were cough , catarrh , abdominal pain , headache , diarrhea and sore throat . the main physical and laboratory findings were under weight , inflamed pharynx , anaemia , malaria parasitemia and neutrophilia . comparatively , only the presence of convulsions and coma were significantly ( p < 0.05 ) commoner in the 79 children presenting with seizures or seizure related disorders as against the others without seizures . clinical features of 17 patients with febrile seizures eleven ( 64.7% ) received an antipyretic ( acetaminophen ) or an anti - inflammatory ( ibuprofen ) drug . six ( 35.3% ) were given antimalarials syrups ( chloroquine , sulphadoxine / pyrimethamine , artesunate or quinine ) while 2 were given an antibiotic ( metronidazole ) for diarrhea . seven ( 41.2% ) had traditional herbal mixture given orally or rubbed on the skin during or after a seizure . five ( 29.4% ) consulted patent medicine sellers ( pms ) and received an antimalarial ( chloroquine , artesunate , or quinine syrup ) and an antibiotic ( chloramphenicol , co - trimoxazole or ampicillin / cloxacillin ) . thrusting of an object ( mainly the finger or spoon ) in the mouth during the seizure was the major physical intervention undertaken . actions taken by caregivers of 17 of patients with febrile seizures drug administration was characterized by inadequate and irregular dosing . fourteen ( 82.4% ) had simple febrile seizures while 3 ( 17.6% ) had the complex variety . there was a positive family history in 5 ( 29.4% ) cases : 2 among those with simple seizures and all the three with complex seizures . it was the first episode in 11 ( 64.7% ) cases and all simple seizures . in 6 ( 35.3% ) of the cases experiencing a second seizure , the causes were , only malaria in 7 ( 41.2% ) cases , acute respiratory infections alone in 5 ( 29.4% ) and a combination of malaria and ari in the remaining 5 ( 29.4% ) . where indicated an antimalarial ( artesunate / lumefantrine combination ) and/or antibiotic ( amoxicillin for those without prior antibiotic administration and cefuroxime for those with previous antibiotic administration ) the prevalence of febrile seizures among hospitalized patients in this study ( 2.7% ) was low compared to that seen at other centers in nigeria.72021 this might be attributable to home treatment , presentation at lower levels of health care delivery or an actual reduction of its incidence in the community . has also been reported in other studies.21 tailor et al , opined that the comparatively earlier maturation of the female brain could provide protection against potential triggers such as fever.22 others have inferred that such observation could be as a result of cultural attitude that encourages earlier presentation of males.2123 the higher number of patients in the upper social classes could be as a result of this group 's capacity to access and afford health care.24 the observation of a positive family history in some of the patients support a genetic basis for its occurrence and this has been previously highlighted.123 furthermore , the occurrence of most seizures after the first day of onset of symptoms underscores the need to urgently identify and adequately manage underlying conditions . also , the incidence of all the complex seizures in the subsequent episodes of febrile seizures suggests a higher risk of its occurrence among those with recurrent febrile seizures . it calls for closer monitoring of fever in those who have had an episode of simple febrile seizure . inappropriate home management of fever , which has also been observed by audu and ogala in zaria,14 and ndifon et al , in calabar,25 was also noted in the study . the implication is the subsequent development of severe or complicated forms of disease among children as observed by eseigbe et al.15 however , it unveils the beneficial potential of caregivers , if properly trained on home management of common childhood illnesses , in child health care delivery in the country . patent medicine sellers are part of the informal health sector in the country and licensed to sell mainly over the counter drugs . however , they have been found to prescribe and sell proprietary drugs , inappropriately and incompetently , beyond their statutory limits . due to their accessibility and affordability , the pms were patronized for the management of febrile seizures in this study and they rendered extracurricular services inappropriately . it can be concluded , as did akuse et al , that with proper training pms have the potential to improve control of common diseases in the community . the use of traditional medicine observed in the study is in conformity with the fact that its use is a viable part of the complex health care system in nigeria.2627 furthermore , it underscores the need to address its role in health care delivery in the country . the use of traditional medication , and its attendant challenges , in the treatment of febrile convulsions and other seizure related disorders have been documented in other parts of the country.212829 ojukwu et al , reported that mortality with febrile seizures was associated with traditional medicine ingestion.7 considering the degree of utilization , traditional medicine if properly harnessed could play a more significant role in health care in the country . an example is seen with valuable use of artesunate , obtained from a chinese herbal extract , in the treatment of malaria.30 some of the pre - hospital interventional practices carried out by the care givers in this study and that observed in other similar studies,212829 such as thrusting of objects in the mouth of a convulsing child , application of onion squash to the eyes , applying scarification marks to the body and burning of the feet , are harmful . its occurrence emphasizes the need to improve public awareness about seizure related disorders and their pre - hospital management . malaria and ari , major causes of childhood illness in the country,3132 were the identifiable causes of febrile seizures in this study . this was similar to findings of osaghae and mukwuzi - odum in benin city21 where both conditions were associated with 80% of the children with febrile convulsions . it highlights the need to vigorously pursue already existing programs such as millennium development goal 633 targeted at combating endemic diseases and , implementing new policies geared towards prevention and control of these conditions . the study revealed a low prevalence of febrile seizures among hospitalized patients with majority presenting with the first episode of the simple variety and poor pre - hospitalization management . it highlighted the need to improve community awareness , establish appropriate caregiver health seeking behavior and promote malaria and ari prevention initiatives . Output:	background : febrile seizure is the most common seizure of childhood and has a good prognosis . however its presentation is fraught with poor management , with grave consequences , in our environment . thus a review of its current status is important.objective:to review the status of febrile seizures in kaduna metropolis.materials and methods : a review of cases seen in the department of paediatrics , 44 nigeria army reference hospital , kaduna between june 2008 and june 2010.results:out of the 635 cases admitted in the department 17 ( 2.7% ) fulfilled the criteria for febrile seizures . there were 11 males and 6 females ( m : f , 1.8:1 ) . age range was from 9 months to 5 years with a mean of 2.2 years 1.1 and peak age of 3 years . twelve ( 70.6% ) were in the upper social classes ( i - iii ) . fever , convulsion , catarrh and cough were major presenting symptoms . incidence of convulsion was least on the 1st day of complaint . fourteen ( 82.4% ) of the cases were simple febrile seizures while 3 were complex . there was a positive family history in 5 ( 29.4% ) of the cases . eleven ( 64.7% ) had orthodox medication at home , before presentation , 5 ( 29.4% ) consulted patient medicine sellers and 7 ( 41.7% ) received traditional medication as part of home management . malaria and acute respiratory infections were the identifiable causes . standard anti - malaria and anti - biotic therapy were instituted , where indicated . all recovered and were discharged.conclusion:there was a low prevalence of febrile seizures among the hospitalized children and a poor pre - hospitalization management of cases . it highlighted the need for improved community awareness on the prevention and management of febrile seizures .
PubmedSumm2018	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: ostriches provide dietetic meat , valuable skin , feathers and eggs [ 3 , 4 , 27 ] that make them important alternative livestock in many parts of the world [ 57 , 13 , 22 , 23 ] . however , one of the basic reasons for the hindered development of this new agricultural activity is its low reproduction rate [ 2 , 14 , 15 , 25 ] and high housing costs . it is more profitable to keep one hen which produces 6080 eggs per season than two hens with half of that egg production . so it becomes necessary to obtain higher genetic progress of the production in laying ostrich hens . due to the development of molecular methods , e.g. microsatellite sequences , new opportunities for genetic improvement of ostrich flocks have emerged in the last decades [ 8 , 12 , 1619 ] . microsatellite sequences are widely used as genetic markers , because they occur in the genome frequently , are evenly - distributed and show wide inter - individual variation and a high rate of heterozygosity [ 9 , 17 , 20 , 21 , 26 ] . facing the above , the aim of the study was to identify specific genetic markers the material consisted of 24 unrelated african black ostrich hens kept in breeding pairs or trios at the stypuw farm , poland , which maintains the birds under conditions compliant with eu recommendations by the committee of the european convention for the protection of animals kept for farming purposes ( t - ap)draft recommendation concerning ratites ( ostriches , emus and rheas ) . the study included the collection of non - invasive material only ( feathers ) which did not require the approval of an ethics committee . the stypuw farm is under official scientific supervision of the institute of genetics and animal breeding of the polish academy of sciences ( official letter of agreement signed in 2002 ) . two groups of hens in their third laying season ( 12 per group ) were randomly completed according to maximum or minimum values of laying performance : group hp ( high productivity)with a total egg production of minimum 75 eggs per hen per season ( mean of 78.92 ; sd = 5.00 ) and group lp ( low productivity)where egg production did not exceed 25 eggs per hen per season ( mean 18.75 ; sd = 3.89 ) . ostrich genomic dna was isolated from feathers ( non - invasive methods ) using dneasy tissue kit 250 ( quiagen ) . an analysis of 30 microsatellite loci characteristic of ostrich , derived from the cau ( china agricultural university ) group was performed . one of the primer pairs has been labeled with one of the four dyes6-fam , vic , ned , pet . the amplification of selected microsatellite loci was performed using a thermal cycler ptc-200 engine ( mj research ) . the pcr was carried out in a total volume of 10 ml comprising 10 ng of template dna , 0.5 mm of each nucleotide , 100 pmol of each primer , 1.5 mm mgcl2 , 50 mm kcl , 10 mm tris hcl , 0.01 % tryton x-100 and 0.5 units of dna polymerase ( polgen ) . for all tested microsatellite loci the fluorescent pcr products were separated by electrophoresis using the four - capillary genetic analyzer ( applied biosystems 3130 ) and the computer software ( genescan ) . in addition , the computer program genemapper ( applied biosystems ) was used to determine the allele size for the individual markers automatically . the computer program genpop , version 4.1 was used to determine : heterozygosity and polymorphism information content ( pic)for evaluation the genetic variability and deviations from hardy weinberg equilibrium . deviations from hardy weinberg equilibrium ( hwe ) were tested by the chi - squared test . a preliminary study on the identification of genetic markers associated with the egg production of ostriches has earlier been conducted by kawka et al . , but based mainly on the analysis of dna fingerprinting including the genetic linkage between minisatellite dna markers and quantitative trait ( egg production ) . methods based on minisatellite dna markers did not distinguish bands specific for the high or low performance groups of hens . the results allowed to conclude neither about the potential linkage between alleles represented by specific hybridization bands or loci of genes , thereof coding for the control of egg production . it should be emphasized that the present study was based on microsatellite loci characteristic for the ostrich since it provides more detailed information and therefore is widely used in linkage mapping of farm animals qtls . analysis of the polymorphism of these loci led to the identification of alleles and loci differing between two groups of ostrich hens with the high and low laying production . table 1 shows characteristics of ostrich groups with high ( hp ) and low ( lp ) laying performance , i.e. heterozygosity expected ( he ) , heterozygosity observed ( ho ) , pic index , genetic differentiation and deviations from hardy weinberg equilibrium . the ho ranged from 0.25 to 1.00 ( lp group ) and from 0.17 to 1.00 ( hp group ) . in turn , the values of ( he ) estimated for population analyzed , ranged from 0.41 to 0.94 ( lp ) and from 0.50 to 0.93 ( hp ) . both mean values ( ho and he ) occurred relatively high ( over 0.8 ) what indicates the high genetic variability of the population in question . kawka et al . , analyzing the genetic variability within and among 3 ostrich breeds reported a mean observed and expected heterozygosity ranging from 0.463 to 0.663 and from 0.481 to 0.679 , respectively . kimwele and graves showed , that the he for an ostrich populations living in wild and kept on farms in kenya , ranged from 0.40 to 0.79 . in turn , hammond et al . in emu populations kept on farms in australia reported this ratio to vary from 0.44 to 1.table 1heterozygosites ( he , ho ) , pic , genic and genotypic linkage disequilibrium , and probability of deviation from hardy weinberg equilibrium using weir and cockerham for microsatellite loci between hp and lp groups of ostrich henslocuspichet - ohet - egenic differentiation ( exact g test)genotypic differentiation ( exact g test)hardy weinberg prob . testlphpoveralllphpoveralllphpoverallp valuep valuep valuecau10.830.820.841.000.920.960.920.920.890.630.570.63cau30.780.680.741.001.001.000.880.800.810.380.260.11cau70.710.690.720.830.920.880.800.780.780.040.040.02cau110.780.790.800.830.920.880.880.890.860.380.330.07cau140.670.780.750.830.920.880.780.880.820.110.080.24cau160.770.760.780.921.000.960.860.860.840.850.800.26cau170.830.790.820.830.920.880.920.890.870.960.940.02cau220.650.640.651.001.001.000.770.750.731.001.000.01cau230.690.700.720.921.000.960.790.800.780.140.080.02cau250.680.700.711.001.001.000.800.810.780.480.360.00cau300.790.830.851.001.001.000.890.930.900.100.070.11cau320.830.760.830.420.170.290.920.860.880.030.300.00cau340.650.570.610.830.830.830.760.690.700.950.940.25cau400.760.740.761.001.001.000.860.840.820.460.350.28cau420.760.740.770.670.580.630.860.840.830.080.210.01cau430.750.750.750.921.000.960.850.850.820.970.960.80cau440.540.590.560.920.920.920.670.720.660.930.870.02cau570.610.610.630.250.670.460.710.700.690.130.270.01cau640.790.740.771.001.001.000.880.840.830.950.930.01cau650.750.770.770.830.830.830.860.870.830.840.830.48cau680.610.730.720.831.000.920.700.840.780.060.030.65cau690.760.760.781.001.001.000.860.860.840.210.130.27cau750.800.810.810.921.000.960.890.900.870.870.790.00cau760.850.810.850.921.000.960.940.910.900.360.340.03cau780.300.410.370.500.500.500.410.500.440.220.180.62cau830.810.710.790.670.580.630.910.810.850.170.430.00cau840.710.800.770.920.830.880.810.890.840.460.490.12cau850.890.840.901.001.001.000.980.930.950.040.040.02cau970.680.510.620.750.420.580.790.610.690.410.420.35cau980.760.810.801.001.001.000.860.910.860.390.290.23pooled0.730.720.740.850.860.860.830.820.800.140.13<0.005 heterozygosites ( he , ho ) , pic , genic and genotypic linkage disequilibrium , and probability of deviation from hardy weinberg equilibrium using weir and cockerham for microsatellite loci between hp and lp groups of ostrich hens as regards the pic , the highest value of which ( more than 0.7 ) was observed for 20 loci in lp and for 22 loci in hp group . the lowest values of the pic ( 0.30 and 0.41 ) were recorded for locus cau78 in lp and hp group , respectively ( table 1 ) . almost all the microsatellite markers selected for the current analysis were characterized either by a high heterozygosity or high pic values . generally it can be assumed that the studied ostrich population remained in hardy weinberg ( hwe ) equilibrium ( table 1 ) . however , several loci showed significant ( p < 0.05 ) deviations from hwe ( cau22 , cau32 , cau42 , cau75 , cau83 and cau84 in hp cau25 , cau32 , cau57 and cau83 in lp group ) . the further wider analysis would prove , whether loci showing such disequilibrium between observed and expected genotypes could be associated with laying performance in ostrich . the more precise estimation using genic and genotypic differentiation approach of genepop showed significant differences of allele and genotype frequencies of individual loci between the two groups of layers ( hp and lp ) : cau7,cau32 , cau 68 , cau85 ( table 1 ) . however , overall analysis for all 30 loci together did not show significant differences between groups : chi - square = 72.51 ( df = 60 ) , p value = 0.12 . moreover , out of a total of 30 microsatellite loci examined , 28 showed different alleles for both groups . two microsatellite loci ( cau43 and cau68 ) had no specific alleles in any of ostrich groups . in a total pool of 243 microsatellite alleles , 152 ( 62.5 % ) were common for the two production groups . the most common alleles were observed at locus cau17 ( 8 of 10 identified alleles ) and cau16 , cau43 , cau64 and cau75 - 7 common alleles . in the locus cau7 , out of the total number of 12 alleles , only 3 were common for the studied groups of hens . ninety one ( over 37 % ) microsatellite alleles from a total pool of alleles occurring in the genome of the two analyzed ostrich groups can be considered as specific for the group . of these alleles , 40 ( 16.4 % ) were typical for hp and 51 ( 20.9 % ) for lp . the most of specific alleles occurred at the locus cau7 ( 9 of the 12 identified ) and cau85 ( 9 of the 15 identified ) ( table 2 ) . alleles specific for hp hens were identified at 23 , while for lp hens at 22 microsatellite loci . the most specific alleles for hp hens were identified at loci cau7 and cau854 alleles . thirteen microsatellite loci were characterized by only one specific allele for this group of hens ( table 2 ) . however , in the case of lp hens , the most specific alleles were observed at loci cau7 , cau32 and cau855 alleles . the one characteristic allele for these hens occurred in 7 analyzed microsatellite markers.table 2common and specific alleles for two analyzed groups of ostrich henslocusalleles common for two groups of ostrichallele specific for the grouphens with high productivityhens with low productivitycau184,86,90,94,96,1048892,98,100cau3111,115,117,119113,121cau7185,187,205189,195,207,209183,191,197,203,211cau11104,106,110,112 , 114,11898,100cau14146,148,150,152138,140,144cau16188,190,192,194 , 200,204,206198186cau17160,162,164,166 , 168,170,176,178174180cau22142,144,146,150,152148cau23167,169,177,191171,185181,183,189,195cau25199,201,203,205,207197cau30115,125,127,129,131,135119,123,137117,133cau32179,183,185,189197,199187,191,193,203,205cau34198,200,202,204192,196cau40142,144,146,148150140,152cau42192,200,202,204,206196,198184,194cau43209,211,213,215,217,219,221cau44227,229,231225cau57201,203,215,217205221cau64167,169,171,173,175,181,183159161cau65177,179,181,183,185,187191cau68263,265,267,269,271cau6998,100,106,108,110112104cau75182,184,194,198,200,204,206196cau76224,226,228,230,236218,222,232242,246,248,252cau78117,119121cau84202,204,206,208,210,212200cau85244,246,248,266,268,272226,236,262,264228,230,252,270,274cau97150,154,158,162152160,164cau98162,164,166,168,170160,178172,174 common and specific alleles for two analyzed groups of ostrich hens the relationship between microsatellite marker alleles from the rhode island red and green - legged partrigenous hens and egg production and quality traits in mapping population was studied by wardcka et al . . polymorphism of 23 microsatellite markers was investigated and 30 traits of egg production and quality measured during the laying period . the results confirmed that the analyzed microsatellite loci may be linked to the genes affecting egg production and quality traits . in turn , chatterjee et al . studied the microsatellite variability and its relationship to the other egg production traits in the chicken three of the studied microsatellite loci were found significantly ( p < 0.05 ) related to egg production traits . the results of this study indicate that between the groups analyzed , the lp hens showed significantly more specific alleles ( 56.0 % of the total pool of specific alleles ) , whereas in hp hens specific alleles consisted of 43.9 % of the total pool of these alleles . the results of the current investigation show the usefulness of microsatellite loci as polymorphic genetic markers of laying performance of ostriches as well as possible association of particular allele to egg production . identification of such markers performed for the first time in the ostrich may be useful in ostrich breeding as a new tool in further genetic improvement of ostrich flocks . Output:	the aim of the current study was to search for genetic markers , microsatellite loci associated with laying performance in ostriches . the material consisted of two groups of ostrich hens characterized by high or low laying performance ( over 75 and less than 25 eggs per season , respectively ) . the investigation covered 30 microsatellite loci characteristic for the ostrich ( the cau group ) and led to identification of significant differences in allele and genotype frequencies between the two groups of hens considered . out of a total of 30 microsatellite loci examined , 28 showed different alleles in relation to analyzed performance groups . in hens of high laying performance ( hp group , n = 12 ) , specific alleles occurred in 23 microsatellite loci ( 40 alleles of 243 identified ) , while in those of low egg production ( lp group , n = 12 ) , they occurred in 22 ( 51 alleles of 243 identified ) . the results indicate the usefulness of the microsatellite loci as the potential genetic markers associated with laying performance that can be applied for genetic improvement of ostrich flocks .
PubmedSumm2019	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: prostate cancer is the most diagnosed male malignancy and the second leading cause of cancer - related death among american men . fortunately , mortality rates have increased less than incidence rates in many countries . in brazil , according to the national cancer institute , prostate carcinoma ( pca ) is the second most common cancer , in incidence , after skin tumors ( non - melanoma ) , and , in terms of mortality , it is exceeded only by lung cancer . the etiologic factors associated with prostate cancer are varied , encompassing both host genetic and environmental influences . glycosylation is one of the most common co- or posttranslational modifications . inside a cell , complex glycosylation pathways assemble these oligosaccharides and attach them to proteins and lipids as they travel to the cell surface . furthermore , by virtue of their peripheral location , particular oligosaccharide epitopes on proteins or lipids exert key functions in important intercellular communication processes such as fertilization , immune response , pathogen anchoring , or metastasis . these particular carbohydrate epitopes are recognized by membrane - anchored carbohydrate - recognition domains of different molecules such as receptors , enzymes , antibodies , or lectins . the latter class comprises proteins of nonimmune origin that display specificity for terminal or subterminal carbohydrates , forming non - covalent bonds . on account of these specific affinities , lectins have been extensively used as histochemical probes to characterize various cell types at various stages of differentiation and maturation of cancer [ 4 , 6 , 7 ] . thus , modern lectin histochemistry has become a valuable instrument to analyze patterns of glucide composition in glycoconjugates and their modifications in the cell during malignant transformation in tumors . cratylia mollis ( camaratu bean ) is a native forage from the semiarid region of the northeast of brazil , and its seeds are considered an important lectin source ( cramoll ) , supplying multiple molecular forms ( cramoll 1 to 4 ) with different carbohydrate specificities [ 9 , 10 ] . the aim of the present investigation was to evaluate the binding pattern of two preparations containing isoforms from c. mollis seed lectin ( cramoll 1,4 , a preparation containing cramoll 1 and cramoll 4 glucose / mannose - specific and cramoll 3 galactose - specific ) in human prostate tissues . con a and pna were used for binding comparison since they possess the same carbohydrate specificity as cramoll 1,4 and cramoll 3 , respectively . formalin - fixed , paraffin - embedded tissue blocks of eight cases of normal human prostate were obtained from the obit identification service at the federal university of pernambuco , brazil ; 61 cases of bph and 82 cases of pca were obtained from the tissue bank of the university hospital of the federal university of pernambuco , brazil . patient ages varied between 31 and 69 years ( mean 44 ) at the time of diagnosis for normal tissues , 46 and 92 years ( mean 69 ) for bph , and 52 and 88 years ( mean 66 ) for pca . cramoll 1,4 and cramoll 3 were extracted , isolated , and purified from seeds of c. mollis according to correia and coelho and paiva and coelho at the glycoprotein laboratory at the biochemistry department of the federal university of pernambuco , brazil . cramoll 1,4 and cramoll 3 hrp conjugation was performed according to beltro et al . . canavalia ensiformis agglutinin ( concanavalin a ( con - a ) ) and arachis hypogea ( peanut ) agglutinin ( pna ) conjugated to hrp ( con a - hrp and pna - hrp ) were purchased from sigma ( st . basic and native polyacrylamide gel electrophoresis ( 10% w / v ) was carried out with conjugated and nonconjugated c. mollis isoforms . tissue slices ( 4 m ) were cut , adhered to albumin - treated slides , deparaffinized in xylol , and hydrated in graded ethanol ( 100%70% ) . / v ) trypsin solution for 2 minutes at 37c and afterwards with 0.3% ( v / v ) methanolic hydrogen peroxide solution for 15 minutes at 25c . sections were incubated with hrp - conjugated lectins ( con - a - hrp , cramoll 1,4-hrp , pna - hrp , and cramoll 3-hrp , at various concentrations 8 , 15 , 25 , 30 and 60 g / ml ) for 2 hours at 4c . slices were washed ( twice for 5 minutes ) with 10 mm phosphate buffer saline solution ( pbs ) ph 7.2 , containing 0.15 m nacl , after each step . lectin staining was visualized with 3,3-diaminobenzidine- ( dab- ) hydrogen peroxide in pbs for 4 minutes at 25c . thereafter , tissues were rinsed in distilled water , counterstained with haematoxylin , dehydrated in graded ethanol , cleared in xylol , and mounted . inhibition lectin - carbohydrate binding controls were performed using the specific sugars at a final concentration of 0.3 m for each lectin ( methyl--d - mannopyranoside for cramoll 1,4 and con - a ; d - galactose for cramoll 3 and pna ) . tissue sections were examined using an optical microscope ( nikon eclipse 50i , usa ) . staining intensity was determined as the pattern observed in at least 20% of cells with cytoplasm or membrane staining , in luminal secretion , corpora amylacea , and stroma and scored in four categories : 0no staining , 1weak staining , 2moderate staining , and 3intense staining . for image acquisition , an image analyses system ( software nis was analyzed by nonparametric tests ( mann - whitney test and kruskal - wallis , followed by multiple comparison posttest of dunn ) with a significance level of 95% ( p < .05 ) using graphpad prism version 5.00 . conjugation efficiency of cramoll 3 and cramoll 1,4 to hrp was evaluated using gel electrophoresis for basic and native proteins . the results showed that conjugated lectins migrated more slowly than their respective nonconjugated counterparts ( figure 1 ) . corpora amylacea were observed in 50.8% of the 61 cases of bph . the gleason score for pca ( n = 82 ) was of 2 + 2 = 4 in 3 samples , 2 + 3 = 5 in 1 , 3 + 2 = 5 in 1 , 3 + 3 = 6 in 24 , 3 + 4 = 7 in 36 , 4 + 3 = 7 in 3 , 3 + 5 = 8 in 3 , 4 + 4 = 8 in 2 , 4 + 5 = 9 in 3 , 5 + 4 = 9 in 3 , and 5 + 5 = 10 in 3 . corpora amylacea were visualized in 40.2% of pca in which the gleason score was usually 3 . cytoplasm of normal prostate cells was weakly and moderately stained by con - a and cramoll 1,4 , both at 15 normal tissue stroma was moderately stained by con - a but no staining was observed with cramoll 1,4 ( figures 2(a ) and 2(b ) ) . hyperplastic epithelial cells were stained by lectins , con - a , and cramoll 1,4 , mainly in the apical cytoplasm and cell borders . cramoll 1,4 staining pattern in bph was intense in most cases ( figures 2(e ) and 2(f ) ) . in pca samples , characterized by heterogeneous gleason scores , con - a and cramoll 1,4 staining intensity decreased while the malignancy degree increased , that is , in well differentiated tissues both lectins showed moderate staining while in undifferentiated tissues no staining was observed . pca cell staining was more evident using cramoll 1,4 ( figures 2(i ) to 2(l ) ) . figure 3(a ) presents the statistical comparison of the staining patterns of cramoll 1,4 and con - a in normal and transformed prostate tissues . pna and cramoll 3 ( 25 g / ml galactose specific ) weakly stained the normal tissue and did not present a different staining pattern between stroma and glandular tissue ( figures 2(c ) and 2(d ) ) . bph and pca neoplastic cells were not differentially stained by these two lectins , rending a non - staining or a weakly heterogeneous pattern ( figures 2(g ) and 2(h ) ) . in pca pna and cramoll 3 presented a focal and intense staining pattern in luminal secretion and apical membrane of glandular tumor cells ( figures 2(m ) and 2(n ) ) . pca stroma was moderately stained by pna and cramoll 3 . statistical comparison between staining patterns of cramoll 3 and the staining pattern for cramoll 1,4 and cramoll 3 was more intense than that observed when commercial lectins were used ( figures 4(a ) to 4(d ) ) . in pca lectin staining was completely abolished in all tissues by inhibition of the lectin carbohydrate - binding site with solutions of methyl--d - mannopyranoside ( 0.3 m ) for con - a and cramoll 1,4 and of d - galactose ( 0.3 m ) for pna and cramoll 3 . cramoll 3 and cramoll 1,4 were properly conjugated to hrp . enzyme conjugation efficiency of in - house purified proteins is very important to assure the quality of histochemical assays . con - a and cramoll 1,4 ( glucose / mannose specific ) data were similar to those reported by morales et al . using ( galanthus nivalis agglutinin gna10 g / ml ) and con - a ( 20 g / ml ) for normal and hyperplastic prostate cells . they observed that normal prostatic glands showed low content or nonaccessibility to glucose / mannose residues when compared to the hyperplastic glands in which the high staining pattern indicates a high content or accessibility to these saccharides in secretory cells ( apical cytoplasm of epithelium and luminal content ) . the lectin staining of secreted glycoconjugates with mannose / glucose residues is observed in human hyperplastic prostatic glands . the increase in glucose / mannose glycocalyx content can be related to high production of glycoconjugates bearing these residues as observed by lectin histochemistry . in prostatic hyperplastic glands the presence of acid phosphatase which contains n - linked high - mannose chains in its structure was observed by jakob et al . . the present study also suggests that the increased lectin staining in bhp could be related to an increased secretor activity of prostatic cells in this pathology . con a staining of normal prostate stroma is in disagreement with the results of arenas et al . that observed a weak stroma staining of normal , bph , and pca tissues . this occurred probably because they used different enzymatic treatments that may have cleaved expressed stroma glycoconjugates bearing glucose / mannose residues in contrast to our trypsin treatment . con - a and cramoll 1,4 different staining patterns indicate that the latter has greater diagnostic value for prostate diseases than the commercially available lectin . such differences in staining can be explained by their similar but not equal quaternary structure which influences the stabilization of the lectin - carbohydrate complex . thus , the glycoconjugates expressed in pathological prostatic tissues seem to be more likely or prone to form more stable complexes with cramoll 1,4 . in the present study , pna and cramoll 3 increased staining patterns from normal to pca tissues are in agreement with the results of janssen et al . who observed that the binding of pna increases from bph to cancers . these authors also established a significant positive correlation between the number of pna acceptors and those of prostatic - specific antigen . arenas et al . , using pna , found an increasing staining pattern of epithelial cells in normal and bph to pca . we observed that using pna as well as cramoll 3 in pca , galactose residues content increased only in luminal secretion and the apical membrane of malignant tumor glands and not in the cytoplasm of epithelium cells . the absence and/or nonaccessibility of galactose residues in epithelial cell glycoconjugates can be evaluated using a neuraminidase pretreatment instead of or together with trypsin ( as used in our work ) in order to expose galactose nonaccessible residues due to the presence of sialic acid residues . pna staining in pca stroma suggests that the development of pca is associated with an increase in galactose residues in glycoconjugates at luminal border of prostate cells and stroma . the low incidence of corpora amylacea in pca can not be used to exclude malignancy as also observed by christian and collaborators . corpora amylacea contain an amyloid substance , 2-microglobulin , a double content of sugars ( glucosamine and galactose ) in relation to proteins and sulfur atoms that suggest the presence of glycosaminoglycans [ 12 , 18 ] . our labeling for bph glands and normal prostate are similar to those observed by morales et al . . the intense lectin staining in corpora amylacea of secretory cells of bph is probably due to the accumulation of keratan sulfate after luminal secretion , a component which has been identified in both prostatic secretor cells and corpora amylacea . the origin and function of corpora amylacea have been debated but its function continues to be unknown . research indicates that corpora amylacea have a secretory origin [ 12 , 20 ] . beyond structural roles in increasing the protein stability , protecting from proteolysis , and improving the protein solubility , glycosylation promotes also general functional diversity reflected on the high structural possibilities of glycans which can be acting as key events in apoptosis , immunomodulation , and antiproliferative activity in tumor cells . and the lectins remain in focus to decipher these diverse glycan chains , being intensively used as cyto- and histochemistry tool for diagnosis and prognosis of cancer . our results showed that c. mollis lectin isoforms are able to recognize cells and an extracellular structure ( corpora amylacea ) of prostatic tissues . proving that lectins bind differently the glycocode of normal and tumor cells and are able to detect subtle neoplastic changes of histologically related or similar pathologies ; these two isoforms of a brazilian northeast native forage plant can be used as auxiliary tools in prostate cancer diagnosis compared to commercial lectins such as con - a and pna . the knowledge of the saccharide identity of cell glycoconjugates enables them to be used as targets for drug delivery and immunomodulation . Output:	lectins , proteins which selectively recognize carbohydrates , have been used in histochemistry for the evaluation of changes in glycosylation in processes of cellular differentiation and/or dedifferentiation . cratylia mollis seed lectins ( cramoll 1,4 and cramoll 3 ) , conjugated to horseradish peroxidase , were used as histochemical probes in human prostate tissues : normal ( np ) , hyperplasia ( bph ) , and prostate carcinoma ( pca ) . the staining pattern of con - a and cramoll 1,4 in bph was more intense than in np . these lectins also showed staining differences between bph and pca ; the latter showing decreased staining intensity with an increased degree of malignancy . pna and cramoll 3 stained epithelial cells similarly in all diagnoses although they did present intense staining of pca glands lumen . corpora amylacea were not differentially recognized by any of the lectins . cramoll 1,4 and cramoll 3 seed lectins present themselves as candidates for histochemical probes for prostate pathologies when compared to commercial lectins such as con - a and pna .
PubmedSumm2020	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: approximately 150,000 new cases will be diagnosed in the united states in 2008 and approximately 50,000 deaths will be attributed to the disease this year . most patients are diagnosed with colorectal cancer in the sixth and seventh decade of life . approximately 30% of colorectal cancers occur in the sigmoid , 25% occur in the rectum and 25% occur in the cecum and ascending colon . histologically , colon cancers are adenocarcinomas that form moderately to well - differentiated glands that secrete varying amounts of mucin . imaging studies are frequently used to evaluate patients in the screening , staging and surveillance of colorectal cancer . this review presents the positron emission tomography ( pet)/computed tomography ( ct ) imaging findings that may be encountered in the diagnosis , staging and follow - up of patients with colorectal cancer . the adenoma carcinoma sequence theory is well established and suggests that many colon cancers develop directly from adenomatous polyps . the malignant potential of a polyp is largely determined by its size . polyps greater than 2 cm in size have a greater than 40% risk of being cancerous , while those less than 0.5 cm are essentially at no risk for harboring malignancy . other features of a polyp that predispose to malignancy are villous architecture and degree of cellular atypia and dysplasia . the cumulative risk for developing invasive carcinoma in unresected polyps has been reported to be 2.5% at 5 years , 8% at 10 years and 24% at 20 years . with the knowledge that colon cancers develop slowly over time , most often from preexisting adenomas , screening is of great importance for prevention of colon cancer . the ideal screening test should be safe , accurate and inexpensive . while there are several screening methods currently in use , such as fecal occult blood testing , optical colonoscopy and imaging studies such as barium enema or ct colonography while imaging studies generally provide an anatomic or structural snapshot of abnormalities , pet imaging differs in that it provides information about metabolic activity and function . [ f]fluorodeoxyglucose ( f - fdg ) pet has been clinically used for the evaluation of patients with a wide variety of cancers since most malignancies , including colorectal cancer typically show increased glucose metabolism . the greatest difficulty in using pet for colonic abnormalities is the presence of physiologic uptake in the gastrointestinal tract . differences in the histology of the intestinal glands in the ascending colon , descending colon , rectum , and small intestine can cause regional differences in fdg uptake and standard uptake value ( suv ) readings . the presence of lymphoid tissue in the colon may contribute to fdg uptake . increased activation of glandular structures in the ascending colon compared with other regions could induce increased fdg uptake or even excretion into the lumen . muscular activity with peristalsis may be another minor contributor to physiologic colon uptake . aside from normal colonic uptake of fdg a study by yasuda et al . looked at 110 patients and found that precancerous adenomatous polyps can be detected incidentally on whole body images performed for other indications with a sensitivity of 24% . the investigators in this study showed that benign colonic adenomas with fdg uptake could not be distinguished from fdg avid carcinomas of the colon . the positivity rate for pet rises with increasing polyp size , with 90% positivity in lesions greater than 13 mm . another study by van kouwen et al . showed similar findings with higher detection rates with increasing size ( 72% sensitivity with size > 11 mm ) and grade of dysplasia of the adenomatous polyps . since colonic uptake is frequently seen on fdg - pet imaging , it is important to determine whether the process is focal or diffuse to help distinguish physiologic from pathologic activity . while the measurement of suv does not allow the differentiation of benign from malignant processes of the colon , the presence of focal colonic fdg uptake as an incidental finding on pet / ct justifies a colon screening examination and pet / ct fusion can be particularly helpful for localization of lesions ( fig . 1 ) . fdg - pet may also identify inflammatory diseases of the colon such as inflammatory bowel disease or diverticulitis ( fig . 2 ) . figure 1coronal mip image ( a ) of an fdg - pet scan in a patient with a history of lymphoma who presented for routine surveillance shows focal uptake in the right lower quadrant ( arrow ) corresponding to a lesion in the cecum ( arrow ) on axial fused pet / ct ( b ) which proved to be a 3-cm adenomatous polyp at colonoscopy . figure 2coronal mip pet image ( a ) of an fdg - pet scan shows a focal area of uptake in the descending colon ( arrow ) . corresponding axial ct ( b ) image shows diverticulosis and surrounding stranding ( arrow ) compatible with diverticulitis . coronal mip image ( a ) of an fdg - pet scan in a patient with a history of lymphoma who presented for routine surveillance shows focal uptake in the right lower quadrant ( arrow ) corresponding to a lesion in the cecum ( arrow ) on axial fused pet / ct ( b ) which proved to be a 3-cm adenomatous polyp at colonoscopy . coronal mip pet image ( a ) of an fdg - pet scan shows a focal area of uptake in the descending colon ( arrow ) . corresponding axial ct ( b ) image shows diverticulosis and surrounding stranding ( arrow ) compatible with diverticulitis . a few studies have investigated the feasibility of combining fdg - pet with ct colonography ( ctc ) , a relatively new technique that provides an endoluminal perspective of the colon . a prospective study by gollub et al . evaluated 17 patients with a combined pet / ct examination after colonic cleansing and insufflation of the colon with carbon dioxide . these investigators found that pet / ctc was a feasible technique allowing excellent image correlation in polyps measuring greater than 10 mm and showing promise in accurate anatomic correlation of both malignant and premalignant lesions of the colon . cost , availability and relative non - specificity make this technique less than feasible at the present time for widespread colorectal screening . once a diagnosis of colorectal cancer is established , staging becomes important for prognostication and to determine appropriate therapy . complete surgical removal of tumor , along with regional lymphatics affords the best prognosis for patients with colorectal cancer . neoadjuvant or adjuvant chemotherapy and radiation therapy are therefore increasingly administered to decrease the incidence of recurrence . in patients with rectal cancer , pre - operative therapy can help to downstage more advanced tumors , which allows sphincter preservation . once a tumor is invasive , it may extend through the layers of the colonic wall and invade adjacent structures . the greatest value of pet lies in the fact that total body coverage allows detection of distant sites of disease . pet and pet / ct are clearly limited for t staging of the primary tumor due to limited spatial resolution and inability to distinguish the layers of the colonic wall . transrectal ultrasound ( us ) and magnetic resonance imaging ( mri ) provide much better anatomic resolution and are of greater value for t staging . nodal staging can be difficult with cross sectional imaging techniques such as us , ct and mri . on cross - sectional imaging , size ( greater than 1 cm ) remains the primary criterion for predicting nodal metastasis , although it is well known that size is not an ideal indicator of disease . the advantage of pet lies in the ability to use metabolic activity to help distinguish benign from malignant adenopathy at sites away from the immediate vicinity of the primary tumor . nodes in the immediate vicinity of the primary tumor are very difficult to detect with pet due to fdg activity of the primary which may obscure small lymph nodes . small nodes are also not easily detected with pet . the overall sensitivity for nodal staging is therefore reported to be quite low , only 29% . it is important not to confuse physiologic activity in the urinary system with tumor spread in the retroperitoneum or pelvis and fused pet / ct has an advantage in anatomic localization over pet alone . this is important because limited disease spread such as to the liver may be resected for cure . resection of colorectal cancer metastases with or without hepatic arterial perfusion therapy can lead to up to 60% 10-year survival in selected patients . therefore , pre - operative knowledge of tumor extent is very important to determine if curative resection is feasible . it has been suggested that fdg - pet is more sensitive than ct in the detection of hepatic and pulmonary metastases and in identifying other sites of intra - abdominal disease . fdg - pet showed greatest accuracy in the detection of liver metastases with reported accuracy up to 99% , sensitivity up to 100% and specificity up to 98% . it is important to keep in mind that lesion size is an important criterion for detection and small hepatic lesions are still not easily detected due to relatively high background liver activity . figure 3coronal mip pet image shows a primary fdg avid tumor in the rectosigmoid ( thick arrow ) with fdg avid metastases to the liver ( thin arrows ) in a patient with newly diagnosed colorectal cancer . coronal mip pet image shows a primary fdg avid tumor in the rectosigmoid ( thick arrow ) with fdg avid metastases to the liver ( thin arrows ) in a patient with newly diagnosed colorectal cancer . pet may also identify sites of disease that may preclude surgery or change the surgical approach . several studies have shown that findings on pet and pet / ct results in change in stage and thereby alters management in up to 1/3 of patients1618 . in a study of patients with low rectal cancers , fdg - pet / ct altered treatment plans in 38% of patients largely through the detection of unsuspected inguinal adenopathy . fdg - pet has also been used to predict response to pre - operative therapy and thereby predict outcome in several different malignancies including rectal cancer2024 . in a study by guillem et al . , 15 patients with locally advanced rectal cancer underwent fdg - pet imaging before and after completion of chemoradiation . all patients showed some degree of response to pre - operative therapy based on pathologic examination . the mean percentage decrease in suvmax was 69% for patients that remained free of disease at a median follow - up of 42 months ; the suvmax decreased by only 37% in patients who eventually developed recurrence . although most patients with colorectal carcinoma undergo surgery with the intent of cure , nearly 4 out of 10 patients experience relapse of disease . over the past decade , aggressive surgical approaches to metastatic disease are being practiced and nearly 30% of patients undergo resection of recurrent disease with increased long term survival . with the use of newer chemotherapeutic agents , many lesions which are deemed unresectable can be downsized thereby allowing potentially curative surgery . even in patients who have surgically unresectable disease , increasing use of newer chemotherapeutic agents , when given early show improved survival . nearly 85% of recurrences occur within the first 3 years after surgery and nearly none occur after 5 years . various approaches to surveillance are used by clinicians from the strategy of call me if you have symptoms to the use of aggressive monitoring with regular clinic visits , periodic tumor marker assays , cross sectional imaging , ultrasound and endoscopy . recognizing the benefit of early treatment in resectable metastases despite acknowledging the lack of sufficient data to determine the optimal frequency of tests , the american society of clinical oncology ( asco ) in 2005 recommended that assay by carcinoembryonic antigen ( cea ) be performed every 3 months for the first 3 years , ct scan of the chest , abdomen and pelvis be performed every year for the first 3 years and an endoscopy at 3 years in patients with stage 2 and stage 3 colorectal carcinoma . in patients with a history of colorectal cancer , pet is commonly used as a problem solving tool when there is a high index of suspicion for recurrence as evidenced by a rising cea but when the routine diagnostic work up is equivocal . reported that fdg - pet found disease in 15 of 22 patients with elevated cea but negative diagnostic work up . they showed a positive predictive value of 89% and a negative predictive value of 100% . in a similar study , flamen et al . report a sensitivity of 75% and a positive predictive value of 79% in a retrospective study of 50 patients . however , these studies were done with stand alone pet instruments between 1993 and 1996 and between 1996 and 1999 respectively . significant technical improvements providing hybrid images and images of a higher resolution and quality have taken place since that time . another common application of pet in patients with recurrent disease is in surgical planning , particularly in patients who develop resectable metastases in the liver or lungs . identification of occult metastases in such patients would avoid unnecessary surgery in many and alter management significantly . in a meta analysis , wiering and colleagues report that fdg - pet changed clinical management in 31.6% of patients . pet in pre - surgical planning decreases the number of futile surgeries and , may also lead to increased survival by allowing for better patient selection . fdg - pet also affords some benefit in rectal cancer patients who have been treated with surgery and chemoradiotherapy with subsequent development recurrence in the pelvis . early identification of pelvic recurrence is necessary for surgery to be of any benefit and distinguishing tumor from post - treatment fibrosis can be a challenge for conventional cross sectional imaging studies ; fdg - pet can be useful in this regard ( fig . fdg - pet can show other sites of disease that would avoid unnecessary surgery . figure 4fused pet / ct image ( a ) shows an fdg avid area along the left pelvic sidewall ( arrow ) with diffuse pre - sacral thickening without a distinct mass on contrast enhanced mri ( b ) in a patient with colorectal cancer treated with chemoradiation and surgery , now with rising tumor markers . biopsy of the fdg avid area proved recurrence . fused pet / ct image ( a ) shows an fdg avid area along the left pelvic sidewall ( arrow ) with diffuse pre - sacral thickening without a distinct mass on contrast enhanced mri ( b ) in a patient with colorectal cancer treated with chemoradiation and surgery , now with rising tumor markers . fdg - pet has less spatial resolution when compared to other cross sectional imaging modalities such as ct and mri . currently , the resolution of most commercially available pet scanners is in the range of 1.3 to 1.5 cm and lesions smaller than this size may not be detected because of volume averaging . overall high background hepatic activity also makes pet for assessment of small hepatic metastases difficult . despite a few reports of relative superiority of pet in detecting hepatic metastases compared to cross sectional imaging , the lack of clear anatomic landmarks and inability to detect small lesions are major limitations with pet alone . another limitation of fdg - pet that must be kept in mind when imaging patients with colorectal cancer is the relative insensitivity for detection of mucinous tumors likely due to the paucicellularity of these tumors . the use of neoadjuvant chemotherapy before surgery can also decrease the sensitivity of pet in lesion detection . serosal metastases on the surface of the large and small bowel may still be missed , due to physiological bowel activity . pulmonary metastases , particularly small ones , may be missed because of partial volume artifacts amplified by breathing . in addition to all of the above mentioned false negatives on fdg - pet , it is important to keep in mind that inflammation can result in false positive fdg uptake . in summary , fdg - pet has been used for detection , staging and surveillance of disease in colorectal cancer patients . physiologic activity in the gastrointestinal tract can be problematic and careful correlation with fused ct images should be performed to improve specificity . fdg - pet also provides information for staging particularly with regard to the presence of distant metastatic disease . there is insufficient data to justify the routine use of fdg - pet in detecting recurrence in patients with colorectal cancer , mainly due to the lack of large randomized trials . pet is still considered a modality with emerging applications . in areas such as post - operative surveillance of colorectal carcinoma where the surgical treatment options and chemotherapy strategies are being constantly redefined , pet ct may find additional future applications , particularly with the development of new , more specific radiotracers . Output:	abstractcolorectal cancer is a common malignancy that afflicts many in the western world . imaging studies are frequently used to evaluate patients in the screening , staging and surveillance of colorectal cancer . cross sectional imaging studies such as ultrasound , computed tomography and magnetic resonance imaging provide anatomic and morphologic information about tumor and patterns of spread . positron emission tomography ( pet ) differs in that it provides information about tumor metabolism.[18f]fluorodeoxyglucose pet has been clinically used for the evaluation of patients with a wide variety of cancers since most malignancies , including colorectal cancer , typically show increased glucose metabolism . this review present the positron emission tomography / computed tomography imaging findings that may be encountered in the diagnosis , staging and follow - up of patients with colorectal cancer .
PubmedSumm2021	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the exercise of metal / metal oxide nanoparticles as a frontier between the homogeneous catalysis and heterogeneous catalysis in organic synthesis has invoked tremendous interests in the recent times . the interesting features inherited with these small particle sizes are their large surface area along with more edges and corners and distinct electronic , optical , magnetic , thermal , and chemical properties [ 35 ] . the crucial role of nanoparticles in organic transformations is their excellent catalytic activity , straightforward recoverability , better selectivity , criteria of evolution , and their versatile role in green chemistry [ 610 ] . thus , the domain of metal nanoparticle catalysis [ 1113 ] should offer opportunities for mining new chemical reactions [ 1416 ] which include the synthesis of biologically important and synthetically challenging natural products . in the context of green chemistry , organic synthesis in solvent - free reaction condition [ 1821 ] has occupied a significant position in the recent years since solvent - free reaction condition involves the best reaction medium with no medium . one of the key motifs present in the biopolymer rna [ 2326 ] is uracil , a nucleobase of the pyrimidine family which participates in various functions in our life processes . uracil derivatives also have several potent medicinal properties such as bronchodilators and anticancer [ 28 , 29 ] , antiallergic [ 30 , 31 ] , antiviral [ 32 , 33 ] , antihypertensive , and adenosine receptor antagonists [ 34 , 35 ] . recently , our research group reported a greener protocol for the synthesis of bisuracil derivatives . some of the n - substituted bisuracil analogues have been screened for bioactivities against several diseases . to explore the possible applications of the metal / metal oxide nanoparticles in organic synthesis , we have been focusing on the advancement of a protocol termed nose ( nanoparticles - catalyzed organic synthesis enhancement ) [ 3941 ] chemistry in our laboratory . to the best of our knowledge , there has been no report on nano - rod - shaped al2o3 catalyzed n , n - diformylation of bisuracil derivatives . recently , we reported n - formylation of amines catalyzed by nano - al2o3 under solvent - free reaction condition . this work inspired us to focus on nano - al2o3 catalysis for the n , n - diformylation of bisuracil analogous . therefore , in this paper , we wish to account for the same ( scheme 1 ) . nano - al2o3 draws our attention due to its crystalline size and shape , abrasive and insulating properties , less toxicity , large surface area , basic surface characteristics , high resistant towards bases and acids and excellent wear resistance [ 4044 ] . rod - shaped nano - al2o3 ( the average particle diameter is 8.12 nm and average length 25.5 nm , sbet = 185.63 m g , = 3.98 g cm , and purity is 99.99% ) were purchased from sigma aldrich and used as received . the chemicals and reagents were purchased from sigma - aldrich , merck , m / s s.d . fine chemicals pvt . ltd . , and loba chemie , and used without further purification . ir spectra were recorded as kbr pallets in a nicolet ( impact 410 ) ft - ir spectrophotometer . h and c nmr spectra were recorded in a 400 mhz nmr spectrophotometer ( jeol , jnm ecs ) using tetramethylsilane ( tms ) as the internal standard , and coupling constants are expressed in hertz . elemental analyses were carried out in a perkin - elmer chn analyser ( 2400 series ii ) . mass spectra were recorded with a waters q - tof premier and an acquity uplc spectrometer . reactions were monitored by thin - layer chromatography using aluminium sheets with silica gel 60 f254 ( merck ) . in a two - neck round bottom flask ( 50 ml ) , nanorod - shaped basic al2o3 ( 7.0 mol% , 7.12 mg ) were taken , and then 1 g ( 1.0 mmol , 414 mg ) and formic acid ( 98% , 6.0 mmol , 0.23 ml ) were added . after that , it was allowed to stir on a pre heated oil bath at 40c for the required time ( the progress of the reaction was judged by tlc ) . the reaction mixture was brought to room temperature after its completion , and ethyl acetate ( 3 10 ml ) was added and then centrifuged ( 3,000 r.p.m ) to recover the nanocatalyst . having done this , the reaction mixture was washed with water and brine , dried over anhydrous na2so4 , and concentrated in a rotary evaporator , and finally the crude product was purified by column chromatography ( 30% ethyl acetate : hexane as an eluent ) . the recovered catalyst was washed with hot ethanol ( 3 10 ml ) to remove the organic impurities , decanted , dried in an oven at 80c for 6 h , and reused for evaluating the performance in the next run in the reaction as shown in scheme 2 . with the previously reported catalyst characterizations in hand , to begin with , reaction of 6,6-diamino-1,1,3,3-tetramethyl-5,5-(benzylidene)bis[pyrimidine-2,4 ( 1h , 3h)-dione ] ( 1a , 1 mmol ) with formic acid ( 6 mmol ) was chosen as the model reaction ( scheme 2 ) . initially , the reaction was carried out without using catalyst under solvent - free reaction condition at 40c and 80c which did not yield any product ( table 1 , entries 1 and 2 ) . various solvents were also tested under the mentioned condition , but they all failed ( table 1 , entries 311 ) to provide any product . these negative results suggested that we look for an effective catalyst in the present study . next , various lewis acid - base catalysts ( table 1 , entries 1214 ) along with the nanocatalysts ( table 1 , entries 1518 ) were surveyed to observe the influence on rate and yield of n , n - diformylation of 1a which were not fruitful . interestingly , nanorod - shaped basic al2o3 stood out as a choice of catalyst at 7 mol% loading ( table 1 , entry 15 ) under solvent - free reaction condition at 40c . during the course of our experiment , we observed that at higher temperature ( table 1 , entry 19 ) and at lower / higher catalyst loading the yield of the products was poor ( table 1 , entries 2022 ) . thus , the yield of n , n - diformylation product of bisuracil derivatives is highly dependent upon the temperature and catalyst loading . with this supportive optimized reaction condition in hand , a series of bisuracil derivatives ( entries 111 ) bearing different aliphatic , aromatic , and heterocyclic moieties were examined to explore the scope and limitations of this reaction and the outcomes are presented in table 2 . it is clear from table 2 that bisuracil derivatives carrying both electron donating and electron withdrawing groups in benzene ring underwent n , n - diformylation reaction smoothly producing good yields ( table 2 , entries 18 ) . however , longer reaction time was required for bisuracil derivatives substituted with furan and alkyl groups ( table 2 , entries 911 ) . it is worth mentioning that 6-amino-1,3-dimethyluracil when treated with formic acid under the current condition gave n , n - diformylation product in lower yield ( 26% , 9 h ) . the reactions were found to be clean , and no side products were formed . to test the recyclability ( vide scheme 2 ) of nano - al2o3 , it was separated from the reaction mixture by adding ethyl acetate ( 10 ml ) , centrifuged at 3,000 rpm , to pellet out the catalyst . the separated particles were washed with hot ethanol ( 3 10 ml ) to remove the organic impurities , decanted , dried in an oven at 80c for 6 h , and reused for further reactions . the efficiency of the catalyst was found to be unaffected up to 4th run , and after that , its action started to decrease as shown in table 3 . the tons were also retained from fresh up to the 5th cycle , and after that it decreased considerably . the recovered catalyst was also investigated through powder xrd and it was compared with the fresh nano - al2o3 ( figure 1 ) . in the powder xrd of the recovered catalyst after 6th run ( figure 1 ) , the intensity of the peaks ( 4 0 0 ) and ( 1 0 0 ) weakened and became broad . it might be due to the blockage of the pores of the catalyst which caused a decrease in effective active sites and also due to the dislocation of the crystal planes after each run which in turn decreased the yield . the sem micrograph of the fresh nano - al2o3 previously reported by us was also compared with the recycled one ( figure 2 ) under the present study . as indicated in figure 2 , the recycled nano - al2o3 revealed the aggregation of the particles responsible for reducing its surface area and hence deactivated the catalyst after 4th run which caused the lower yield of product . in conclusion , we have demonstrated a novel method for synthesis the n , n - diformylation of bisuracil derivatives in good yield under solvent - free reaction condition at 40c catalyzed by recyclable nano - al2o3 rods . nano - al2o3 catalyzed organic transformations we believe that this work would find wide applications for new chemical transformations , including those which enable the synthesis of complex natural products and derivatives . Output:	a feasible nose ( nanoparticles - catalyzed organic synthesis enhancement ) protocol has been developed for n , n - diformylation of bisuracil derivatives using nano - al2o3 rods as an efficient , inexpensive , and recyclable catalyst under solvent - free reaction condition at 40c . the catalyst was reused up to the 4th cycle without affecting the rate and yield of the n , n - diformylation products appreciably .
PubmedSumm2022	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: molecular studies have reported that up to 15% of sporadic colorectal cancers and approximately 90% of syndrome of hereditarynon - polyposis colorectal cancer ( hnpcc ) , which accounts for 1 to 3 percent of all colorectal cancers , were caused by inactivation of mismatch repair ( mmr ) genes , such as msh2 , mlh1 , pms1 , pms2 , hmsh6/gtbp , and hmsh3 [ 1 - 3 ] . inactivation of one of these genes results in dna microsatellite instability ( msi ) , characterized by alterations in the length of simple repetitive microsatellite sequences found throughout the genome.while germline mutations of dna mmr genes have been identified as a causative event in hereditary nonpolyposis , development of sporadic colorectal cancer with msi - high ( msi - h ) is most commonly a result of promoter methylation , leading to epigenetic silencing of the mlh1 gene . some studies have suggested that colorectal carcinomas with msi - h tend to have an improved survival rate , and may respond differently to adjuvant chemotherapy than non - msi tumors . msi tests may be used routinely as a first - line screening tool for identification of msi - h colorectal carcinomas for diagnosis of suspected hnpcc and for determination of the clinical implications of msi - h status in sporadic colorectal cancer . however , to date , msi tests are complex , time - consuming , and expensive , and are not widely accepted as a screening test . clinical data , including family history of cancer , including amsterdam criteria and/or bethesda guidelines , have been used for selection of suspected hnpcc for msi testing . in addition to clinical data , a number of studies have suggested that msi - h colorectal carcinomas may have morphologic characteristics that differ from those of non - msi - h tumors and therefore may be used as a first - line screening tool for identification of tumors for further molecular testing . however , the utility of histology as a screening tool for msi - h colorectal carcinomas has yet to be defined . immunohistochemistry ( ihc ) for mlh1 and msh2 protein may increase accuracy of prediction and aid in selection of patients who may have msi , and it can be used as a screening tool for msi testing in colorectal cancers . however , these guidelines or histopathologic parameters are not adequate for selection of colorectal cancer patients because their results were studied independently and a false negative result may be obtained . in this study , we attempted to identify the combination of parameters from clinical data , histopathology , including ihc for mlh1 and msh2 protein that may be helpful in selection of patients with msi - h . between january 2004 and june 2006 , the 120 patients who underwent surgery for treatment of colorectal cancers at daegu catholic university hospital , daegu , korea were retrospectively enrolled in the present study . using medical records or telecommunication , the questionnaire included cancer history in first- and second - degree relatives and contained questions regarding their age at diagnosis , type of cancer , hospital at which the diagnosis was made , current age , and current status . patients with hnpcc meeting the amsterdam criteria , patients with familial adenomatous polyposis , and patients with a vague family history were excluded . of the 120 enrolled cases of primary colorectal cancer , 65 were male and 55 were female , with a mean age at the time of surgery of 62.411.1 years ( range , 36 to 83 years).twenty patients had a family history of cancer , except hepatocellular carcinoma or cervical cancer , within a second degree pedigree ( table 1 ) . among them , 10 patients had a family history of colorectal cancer , and only one patient had met the full requirement for diagnosis of amsterdam , and five were included according to bethesda guidelines . the numbers according to tumor location , which were classified as rt . sided colon ( from ileocecal valve to splenic flexure ) , lt . sided colon ( from descending colon to sigmoid colon ) , and rectum ( from recto - sigmoid colon to distal rectum ) and multiple colon ( two or more synchronous colon cancers at diagnosis ) were 27 , 23 , 69 , and 1 , respectively ( table 2 ) . tumors in which less than 10% of the cells formed glands were classified as high grade ( poorly differentiated ) , while those containing more than 50% extracellular mucin were classified as mucinous type . the numbers of well / moderately differentiated adenocarcinoma ( wd / md ) , poorly differentiated adenocarcinoma ( pd ) , and mucinous adenocarcinoma ( mu ) were 112 , 3 , and 5 , and two were diagnosed as lymphoma . the depth of invasion to mucosa / submucosa , muscularis layer , pericolic ( or perirectal area ) , or invasion to adjacent other organs were 11 , 19 , 82 , and 6 , respectively . according to international union against cancer ( uicc ) classification , 19 of these tumors were tumor - node - metastasis ( tnm ) stage i , 55 were stage ii , 37 were stage iii , and nine were stage iv . dna from blood and tissue were amplified for the five mononucleotide markers of nr21 , nr22 , nr24 , bat25 , and bat26 . amplification of nr21 , 22 , and 24 was performed using the home brew pcr system and denaturing high performance liquid chromatography ( transgenomic inc . , omaha , ne ) was used for analysis of the pcr products ( fig . a hybridization probe melting point analysis was performed using light cycler and dna master hybridization probe reagents ( roche diagnostics , mannheim , germany ) for detection of bat25 and 26 instability ( fig . one block of formalin fixed paraffin embedded colorectal cancer tissue was selected for ihc . in all cases , the block was comprised of an area of normal colonic mucosa adjacent to the tumor . sections ( 4 m ) were affixed , dried , de - waxed , and rehydrated , followed by inhibition of endogenous peroxidase activity ; they were then subjected to heat antigen retrieval . the mlh1 antibody ( g168 - 15 , pharmingen , san diego , ca ) and the msh2 antibody ( g219 - 1129 , pharmingen ) were used for ihc stain using the streptavidin biotin universal detection system ( immunotech , marseille , france ) . finally , the sections were counterstained in mayer 's haematoxylin . among 120 cases , ihc could not be performed for three microsatellite stable ( mss ) cases , and , for 117 cases , one pathologist ( h.k . oh ) , without knowledge of msi status , performed scoring of ihc staining expression . for statistical analysis , the or fisher 's exact test was used as an appropriate univariate test for selection of optimal clinico - pathologic factors between groups . we then performed multiple logistic regression for identification of clinico - pathologic factors among all selected optimal factors described above as a univariate test . for selection of clinico - pathologic factors , we used the forward selection method in multiple logistic regression . using clinico - pathologic parameters identified in multiple logistic regression , we evaluated the prediction of msi - h as the sensitivity and specificity for the single and all combinations of identified clinico - pathologic parameters . in addition , we used receiver operatring characteristic curve and area under curve to obtain the most appropriate sensitivity and specificity . between january 2004 and june 2006 , the 120 patients who underwent surgery for treatment of colorectal cancers at daegu catholic university hospital , daegu , korea were retrospectively enrolled in the present study . using medical records or telecommunication , the questionnaire included cancer history in first- and second - degree relatives and contained questions regarding their age at diagnosis , type of cancer , hospital at which the diagnosis was made , current age , and current status . patients with hnpcc meeting the amsterdam criteria , patients with familial adenomatous polyposis , and patients with a vague family history were excluded . of the 120 enrolled cases of primary colorectal cancer , 65 were male and 55 were female , with a mean age at the time of surgery of 62.411.1 years ( range , 36 to 83 years).twenty patients had a family history of cancer , except hepatocellular carcinoma or cervical cancer , within a second degree pedigree ( table 1 ) . among them , 10 patients had a family history of colorectal cancer , and only one patient had met the full requirement for diagnosis of amsterdam , and five were included according to bethesda guidelines . the numbers according to tumor location , which were classified as rt . sided colon ( from ileocecal valve to splenic flexure ) , lt . sided colon ( from descending colon to sigmoid colon ) , and rectum ( from recto - sigmoid colon to distal rectum ) and multiple colon ( two or more synchronous colon cancers at diagnosis ) were 27 , 23 , 69 , and 1 , respectively ( table 2 ) . tumors in which less than 10% of the cells formed glands were classified as high grade ( poorly differentiated ) , while those containing more than 50% extracellular mucin were classified as mucinous type . the numbers of well / moderately differentiated adenocarcinoma ( wd / md ) , poorly differentiated adenocarcinoma ( pd ) , and mucinous adenocarcinoma ( mu ) were 112 , 3 , and 5 , and two were diagnosed as lymphoma . the depth of invasion to mucosa / submucosa , muscularis layer , pericolic ( or perirectal area ) , or invasion to adjacent other organs were 11 , 19 , 82 , and 6 , respectively . according to international union against cancer ( uicc ) classification , 19 of these tumors were tumor - node - metastasis ( tnm ) stage i , 55 were stage ii , 37 were stage iii , and nine were stage iv . dna from blood and tissue were amplified for the five mononucleotide markers of nr21 , nr22 , nr24 , bat25 , and bat26 . amplification of nr21 , 22 , and 24 was performed using the home brew pcr system and denaturing high performance liquid chromatography ( transgenomic inc . , omaha , ne ) was used for analysis of the pcr products ( fig . a hybridization probe melting point analysis was performed using light cycler and dna master hybridization probe reagents ( roche diagnostics , mannheim , germany ) for detection of bat25 and 26 instability ( fig . one block of formalin fixed paraffin embedded colorectal cancer tissue was selected for ihc . in all cases , the block was comprised of an area of normal colonic mucosa adjacent to the tumor . sections ( 4 m ) were affixed , dried , de - waxed , and rehydrated , followed by inhibition of endogenous peroxidase activity ; they were then subjected to heat antigen retrieval . the mlh1 antibody ( g168 - 15 , pharmingen , san diego , ca ) and the msh2 antibody ( g219 - 1129 , pharmingen ) were used for ihc stain using the streptavidin biotin universal detection system ( immunotech , marseille , france ) . ihc could not be performed for three microsatellite stable ( mss ) cases , and , for 117 cases , one pathologist ( h.k . oh ) , without knowledge of msi status , performed scoring of ihc staining expression . for statistical analysis , the or fisher 's exact test was used as an appropriate univariate test for selection of optimal clinico - pathologic factors between groups . we then performed multiple logistic regression for identification of clinico - pathologic factors among all selected optimal factors described above as a univariate test . for selection of clinico - pathologic factors , we used the forward selection method in multiple logistic regression . using clinico - pathologic parameters identified in multiple logistic regression , we evaluated the prediction of msi - h as the sensitivity and specificity for the single and all combinations of identified clinico - pathologic parameters . in addition , we used receiver operatring characteristic curve and area under curve to obtain the most appropriate sensitivity and specificity . of the 120 tumors , 11 ( 9.2% ) were msi - h , four ( 3.3% ) were msi - low ( msi - l ) , and 105 ( 87.5% ) were mss ( table 1 ) . among the 11 patients with msi - h , there were seven females , which appeared to be higher than that of males , without statistical significance . ages ( meanstandard deviation , years ) of msi - h and mss patients were 529.6 and 63.310.9 , respectively , which was significantly different ( p=0.004 ) . of patients with a family history of cancer other than hepatocellular carcinoma or cervical cancer , the ratio of msi - h was 30% ( 6/20 ) , which was significantly higher , compared to those who had no family history of cancer ( p=0.001 ) . among patients with a family history of colorectal cancer , the ratio of msi - h was 40% ( 4/10 ) , which was significantly higher than that of those who had no family history of colorectal cancer ( p=0.002 ) . the only patient who met amsterdam criteria showed msi - h , and , among five patients who were included according to bethesda guidelines , three ( 60% ) were msi - h . the ratio of increased preoperative serum cea level was 9% ( 1/11 ) in the msi - h group and 33.3% ( 33/99 ) in the msi - l or mss group when the cut off value of cea was regarded as 5 ng / dl ( p=0.038 ) . the ratios of msi - h in tumors from the right colon and synchronous multiple colon cancers were 29.6% ( 8/27 ) and 100% ( 1/1 ) , which was higher than those of the left colon or rectum ( p=0.001 ) ( table 2 ) . among five tumors of mu , three ( 60% ) showed msi - h , which was higher than those of wd / md , or pd or lymphoma ( p=0.001 ) . except for one patient , all tumors with msi - h invaded to the pericolic ( or perirectal ) area , and no difference for status of regional lymph node metastasis , lymphovascular invasion , perineural invasion , distant metastasis , or tnm stage was observed between msi and msi - l or mss groups . the numbers for expression of mlh1 were 94 ( 92.2% ) , 3 ( 75.0% ) , and 5 ( 45.5% ) , while 8 ( 7.8% ) , 1 ( 25.0% ) , and 6 ( 54.5% ) were not expressed in mss , msi - l , and msi - h , in order , and the sensitivity and specificity of mlh1 for msi - h were 54.5% and 92.2% , respectively ( table 4 ) . in msh2 , the numbers for expression were 101 ( 99.0% ) , 4 ( 100% ) , and 10 ( 90.9% ) , while 1 ( 1% ) , 0 ( 0% ) , and 1 ( 9.1% ) were not expressed in mss , msi - l , and msi - h , respectively , and the sensitivity and specificity for msi - h were 9.1% and 99.0% . among factors that were proved to be statistically significant on a univariate test , multiple logistic regression analysis was performed for identification of parameters that were significantly affected or related to msi - h . immunohistochemical staining for mlh1 , tumor location , bethesda guidelines , and cell types showed a significant association with msi - h . msi - h tumors had at least one or more parameters among negative expression of mlh1 protein , right sided colon and/or synchronous multiple colorectal cancer , one of the bethesda guidelines , or poorly differentiated or mucinous adenocarcinima . sensitivity and specificity were calculated for all combinations of immunohistochemical staining for mlh1 , tumor location , bethesda guideline , and cell types ( table 5 ) . according to the results , three clinico - pathologic parameters with mlh1 , tumor location , and bethesda guidelines were appropriate and important factors with high sensitivity of 100% and specificity of 72.5% . of the 120 tumors , 11 ( 9.2% ) were msi - h , four ( 3.3% ) were msi - low ( msi - l ) , and 105 ( 87.5% ) were mss ( table 1 ) . among the 11 patients with msi - h , there were seven females , which appeared to be higher than that of males , without statistical significance . ages ( meanstandard deviation , years ) of msi - h and mss patients were 529.6 and 63.310.9 , respectively , which was significantly different ( p=0.004 ) . of patients with a family history of cancer other than hepatocellular carcinoma or cervical cancer , the ratio of msi - h was 30% ( 6/20 ) , which was significantly higher , compared to those who had no family history of cancer ( p=0.001 ) . among patients with a family history of colorectal cancer , the ratio of msi - h was 40% ( 4/10 ) , which was significantly higher than that of those who had no family history of colorectal cancer ( p=0.002 ) . the only patient who met amsterdam criteria showed msi - h , and , among five patients who were included according to bethesda guidelines , three ( 60% ) were msi - h . the ratio of increased preoperative serum cea level was 9% ( 1/11 ) in the msi - h group and 33.3% ( 33/99 ) in the msi - l or mss group when the cut off value of cea was regarded as 5 ng / dl ( p=0.038 ) . the ratios of msi - h in tumors from the right colon and synchronous multiple colon cancers were 29.6% ( 8/27 ) and 100% ( 1/1 ) , which was higher than those of the left colon or rectum ( p=0.001 ) ( table 2 ) . among five tumors of mu , three ( 60% ) showed msi - h , which was higher than those of wd / md , or pd or lymphoma ( p=0.001 ) . except for one patient , all tumors with msi - h invaded to the pericolic ( or perirectal ) area , and no difference for status of regional lymph node metastasis , lymphovascular invasion , perineural invasion , distant metastasis , or tnm stage was observed between msi and msi - l or mss groups . the numbers for expression of mlh1 were 94 ( 92.2% ) , 3 ( 75.0% ) , and 5 ( 45.5% ) , while 8 ( 7.8% ) , 1 ( 25.0% ) , and 6 ( 54.5% ) were not expressed in mss , msi - l , and msi - h , in order , and the sensitivity and specificity of mlh1 for msi - h were 54.5% and 92.2% , respectively ( table 4 ) . in msh2 , the numbers for expression were 101 ( 99.0% ) , 4 ( 100% ) , and 10 ( 90.9% ) , while 1 ( 1% ) , 0 ( 0% ) , and 1 ( 9.1% ) were not expressed in mss , msi - l , and msi - h , respectively , and the sensitivity and specificity for msi - h were 9.1% and 99.0% . among factors that were proved to be statistically significant on a univariate test , multiple logistic regression analysis was performed for identification of parameters that were significantly affected or related to msi - h . immunohistochemical staining for mlh1 , tumor location , bethesda guidelines , and cell types showed a significant association with msi - h . msi - h tumors had at least one or more parameters among negative expression of mlh1 protein , right sided colon and/or synchronous multiple colorectal cancer , one of the bethesda guidelines , or poorly differentiated or mucinous adenocarcinima . sensitivity and specificity were calculated for all combinations of immunohistochemical staining for mlh1 , tumor location , bethesda guideline , and cell types ( table 5 ) . according to the results , three clinico - pathologic parameters with mlh1 , tumor location , and bethesda guidelines were appropriate and important factors with high sensitivity of 100% and specificity of 72.5% . msi tests can be applied for diagnosis of hnpcc from all colorectal cancers and for detection of msi - h tumors in sporadic colorectal cancers . although a definitive diagnosis of hnpcc could be established by demonstrating a germline mutation , msi tests can be used as a screening test for suspected hnpcc before mutation analysis . as clinical and prognostic implications in sporadic type colorectal cancer with msi - h would predict development of multiple synchronous or metachronous - cancers , responsiveness of adjuvant chemotherapy or to chemoprevention related to cyclo - oxygenase-2 inhibitors as well as prognosis , it is also of great importance for identification of msi - h in sporadic type colorectal cancer in a clinical field [ 13 - 15 ] . for assessment of msi status , an international consensus meeting held in 1997 this included two mononucleotide repeats ( bat25 and bat26 ) and three dinucleotide ( d5s346 , d2s123 , and d17s250 ) repeats . tumors showing instability at two or more of these markers were defined as msi - h , and those showing instability at one repeat or showing no instability were defined as msi - l and mss tumors , respectively . in this study , we used five mononucleotide repeats , including bat25 and bat26 , instead of three dinucleotide ( d5s346 , d2s123 , and d17s250 ) repeats , because each of the dinucleotide repeats in the aforementioned panel generally show instability in only 60 - 80% of msi - h tumors . in addition , when using the bethesda reference panel , misclassifications of msi - h tumors occur if two dinucleotide markers were unstable in the absence of bat26 deletions . as reported by zhou et al . , bat26 identified the msi status in 539 of 542 tumors ( 99.5% ) , and bat26 provided the advantage of being a simple and less expensive method that might be used as a screening procedure prior to performance of mutation analysis . according to our data , sensitivity for bat26 and bat25 for our data also indicated the potential of bat26 for use as a single marker for msi testing ; however , it is not desirable , as there may be the possibility of missing msi - h by bat25 or 26 alone due to polymorphisms . introduction of the msi determination as an initial screening test for hnpcc or msi - h tumors in colorectal cancers has been reported to enable molecular detection of hnpcc in a mass population . salovaara et al . , who examined 535 colorectal cancers , reported that 12% showed msi and 3.4% had germline mutations of msh2 or mlh1.this means that at least more than 85% of colorectal cancers are mss or msi - l and application of msi tests for all colorectal cancers may be ineffective due to low sensitivity and specificity for diagnosis of hnpcc or detection of msi - h in sporadic colorectal cancer . to save time and cost of msi testing , some attempts have been made to increase its sensitivity and specificity ; these include bethesda guidelines , evaluation of clinical and histopathologic parameters , msi tests by one marker or replacement of msi by ihc of mlh1/msh2 protein , or a combination of these trials [ 7 - 10 ] . original bethesda guidelines for testing colorectal tumors for msi were proposed in 1997 to guide clinical use to aid in identification of patients who may have hnpcc , and were then revised in 2004 . bethesda guidelines were established only for diagnosis of hnpcc , not msi - h sporadic colorectal cancers . reported that 29% ( 27/92 ) of bethesda - positive patients showed high msi , compared with 6% ( 4/72 ) of patients who did not meet these criteria . according to our data , five patients were included in the bethesda guidelines and three were msi - h , including one hnpcc . use of bethesda guidelines is a suboptimal method for selection of msi testsin sporadic colorectal cancer and for application to general medical practice outside academic centers , thus , additional clinical or pathologic data are required for adequate msi - h tumor selection . as reported previously , msi - h tumors are known for having certain clinical and pathological characteristics , including a higher rate of family history of cancer , early age of onset , a tendency for cancers in the proximal colon and excess of poorly differentiated carcinomas or mucinous type , medullary type , contain signet - ring cells , and increased infiltrating t lymphocytes . reported that positive predictabilities of right sided tumor , intraepithelial lymphocytes , poor differentiation , crohn's - like reaction , mucinous tumor , peritumoural lymphocytes , and amsterdam criteria for msi - h in sporadic colorectal cancer were 23% , 35% , 26% , 26% , 20% , 24% and 25% , respectively . our data also showed similar results , indicating that predictabilities of family history of cancer , bethesda guidelines , right sided tumor , and poorly differentiation or mucinous tumor for msi - h in colorectal cancer were 30% , 30% , 28.6% and 14.3% , respectively ( data not shown ) . reported that medullary carcinoma , intraepithelial lymphocytosis , and poor differentiation were the best discriminators between msi - h and microsatellite - stable , and suggested that these histopathological evaluations can be used in prioritization of sporadic colon cancers for msi studies . however , in their study , histopathology alone failed to reliably discriminate msi - h tumors because the minority of specimens showed no major difference in morphology from the usual mss cancer . in addition , approximately 40% of msi - h cancers were not detected , and 6% were never detected by histopathology . these results suggested that these clinical and pathological parameters showed differences between msi - h and mss or msi - l , but are insufficient for clinical application to selection of patients to undergo msi tests independently . also suggested other parameters associated with msi - h , including sex , distant metastasis , and stage , however , these findings were not consistent with those of other reports , and our data also did not show any correlation with msi status . engel et al . reported that ihc was highly predictive ( 99.1% ) and specific ( 99.6% ) with regard to msi tests . however , their data showed that 14 out of 230 ( 6% ) mutations escaped detection by ihc . in our data , the sensitivity and specificity of mlh1 and msh2 for msi - h were 54.5% , 92.2% and 9.1% , 99.0% , respectively and among 11 cases of msih , seven ( 63.6% ) showed loss of expression of either one of mlh1 or msh2 . , lack of mlh1 nuclear staining was observed much more often than the absence of msh2 nuclear staining . despite a close correlation between ihc and msi , these findings suggested that ihc could not be fully recommended as a substitute for msi tests . in our attempt to find parameters showing significant differences between mss or msi - l and msi tumors by multiple logistic regression tests as a multivariate analysis bethesda guidelines , tumors from the right colon and/or synchronous multiple colon cancers , and negative expression of mlh1 protein in msi - h tumors showed statistical significance . when we applied a selection of patients with one of these three parameters , sensitivity and specificity for msi - h were 100% and 72.5% , respectively . this means that , using these parameters for selection of patients for msi testing , we were able to find all msi - h tumors , with the exclusion of 80 patients who had no msi - h , and to minimize the number of patients for msi testing . using these three parameters , we retrospectively applied 25 patients who underwent surgery before 2003 and were excluded from this study ; all five msi - h patients met at least one of these parameters , and its sensitivity and specificity for msi - h were 100% and 60% , respectively . these findings allowed us to select patients with msi - h tumors who met one of three parameters . except for the bethesda guideline , ihc , tumor location , and cell type showed 100% sensitivity and 70.6% specificity . collection of data for each patient for bethesda guidelines is not difficult ; therefore , careful history taking and routine pathologic data are very important . msi testing should be recommended for positive patients with any of the bethesda guidelines , tumor location , or mlh1 stain . bethesda guidelines , rt . sided colon cancer , and negative expression of mlh1 protein are important parameters for selection of patients with colorectal cancers for msi testing . Output:	purposealthough the incidence of microsatellite instability ( msi ) accounts for 10 - 15% of cases of colorectal cancer , its clinical application for all colorectal cancers has widened . we attempted to identify clinical and pathological parameters that may be helpful in selection of patients with msi - high ( msi - h).materials and methodsa total of 120 resected colorectal cancers were enrolled retrospectively for this msi study . polymerase chain reaction ( pcr ) and denaturing high performance liquid chromatography and/or real time pcr methods with five markers and immunohistochemistry ( ihc ) for mlh1 and msh2 were performed for analysis of cancer and blood specimens . clinico - pathologic parameters , including ihc , were investigated in order to determine their usefulness as predictive factors of msi.resultsamong 120 cases of colorectal cancer , msi was observed in 15 cases ( 12.5% ) , including 11 cases of msi - h and four cases of msi - low . patients with msi were younger , less than 50 years old , had a family history of cancer , rt . sided colon cancer and/or synchronous multiple colorectal cancer , mucinous histologic type , and serum carcinoembryonic antigen group in the normal range . results of multivariate analysis showed bethesda guidelines , rt . sided and/or synchronous multiple colorectal cancer , and negative expression of ihc for mlh1 , which was consistently associated with msi - h . msi - h colorectal tumors have met at least one of these three parameters and their sensitivity and specificity were 100% and 72.5% , respectively.conclusionbethesda guidelines , tumor location , and negative expression of mlh1 protein are important parameters for selection of patients with colorectal cancers for msi testing . msi testing is recommended for patients showing any of these three parameters .
PubmedSumm2023	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: Output:	talc deposits include asbestos minerals such as chrysotile and amphiboles that may be carried over into consumer products . optical microscopy and x - ray diffraction analyses may not reveal their presence . examples are given of electron microscopy procedures that permit detection and measurement . imagesfigure 1.figure 2.figure 3 .
PubmedSumm2024	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: self - care was introduced by orem as one of the components of self - care nursing theory . orem recognizes human and environment as a single unit and believes that human and environment , and also humans themselves , influence each other reciprocally in this unit . beliefs , social and cultural background , personal characteristics , and relationship between health care providers and clients are some of the factors which influence self - care behaviors . in addition , ethnicity , socioeconomic background , educational level , employment status , environmental factors such as pollution , sociopolitical variables , and lack of knowledge are other factors affecting self - care behaviors . the importance of self - care concept is related to human 's need for maintaining and promoting health and recovery , lack of healthcare services , inadequate access to health services for the whole community , and increased expenditure of healthcare services . self - care activities alleviate symptoms and complications of diseases , shorten recovery , and reduce hospital stay and rehospitalization rate . it has been reported that lack of self - care knowledge in patients with chronic disease , such as hepatitis , is the main reason for frequent referring to healthcare centers and rehospitalization . chronic disease , especially hepatitis , can have a significant impact on the quality of life of the patients . patients with chronic hepatitis face many physical and mental problems such as fatigue , weakness , sleep disorders , and anxiety . patients with chronic hepatitis become predisposed to depression , loss of family and community support and social isolation due to fear , complications of diseases , social and economic problems , and use of certain drugs . the chronic course of disease and its economic impacts on patients and their families may cause daily life challenges for these patients . therefore , it is necessary to provide patients with further information on the status of disease and their care plans , and to encourage them to take responsibility for their own health . the association between self - care and the outcomes has been investigated in hepatitis patients . a study has indicated that self - care was a complex process in patients with hepatitis and had numerous influences including individual , familial , and healthcare system - related . self - care behaviors facilitated participants acceptance and changed the participants perspectives regarding emotional and social support . in addition , care providers behavior , negative attitudes toward self , and fear of disclosing the disease have been reported to be the barriers to self - care activities . however , hepatitis can be prevented through self - care activities and behaviors due to mental problems . mental problems and poor socioeconomic status are known as the two main factors which influence self - care . therefore , patients living with chronic hepatitis can enhance the quality of life , functional capacity , and personal and social welfare through self - care activities . one of the gaps in the care of chronic patients , especially in patients with hepatitis , is the lack of a suitable framework to investigate the patients special needs . the purpose of this study is to explore how patients living with chronic hepatitis can practice self - care and to redesign a specific version of orem 's self - care theory for patients with chronic hepatitis . a qualitative directed content analysis approach was used to conduct this study from may 2012 to may 2013 . sometimes , there is an available theory about a phenomenon that is incomplete or may benefit from further description . in this case , qualitative research practitioners might choose to use a directed approach to content analysis . the research ethics committee of the tehran university of medical sciences approved the protocol of this study ( approval no . the potential participants were recruited from a referral center , the shahrekord hepatitis services organization . a purposive sampling method was adopted , and the sampling continued until data saturation was achieved throughout the process of data collection . the inclusion criteria were diagnosis of chronic hepatitis by a specialist , referral of patients aged 18 years and over to behavioral counseling , and being consent to participate in the study . informed consent to participate in the study was provided by the participants prior to data collection . the data were collected from 22 semi - structured interviews with 18 participants and direct observation of their behaviors in the process of data collection by the researchers . the participants were interviewed individually in the local hepatitis services organization by the first author . however , the participants could freely choose to be interviewed either in their own place of choice or the local hepatitis services organization . to access reliable and rich information , the researchers created a close and direct relationship with the participants . the data were collected through semi - structured , in - depth interviews and fieldnotes that the researchers took as they observed the participants . to familiarize with the participants and gain their trust before the formal interview , the researchers met the participants individually in an informal session and then interviewed them in a formal session . prior to the interview , the researchers obtained the informed consent from the participants to record the interviews . throughout the process of interview , initially , the orem 's pattern - based general questions were asked . if the interview was diverted from the purpose of the study , the researchers redirected the interview toward the research purposes using illustrative questions . the number and duration of each interview session varied according to the content and conditions of the participants and depended on various factors including time , desire , the participants patience and strength , and physical and mental condition , the information level of participants , and environmental conditions . the patients were asked questions during interviews , such as what does hepatitis mean to you ? , did you face difficulty in relation to your illness ? , what should the attitude toward hepatitis management be ? , how has hepatitis affected your life ? , how do you do self - care activities ? , and what education have you had in relation to the disease and how does it affects social life ? before completion of each interview , the researchers offered all the participants the opportunity to add anything to the discussion by asking the following question : is there anything else that you feel is relevant and you would like to add ? immediately after each interview , the researchers transcripted the recorded interviews word by word and then checked the interview transcripts against the recorded audio tapes to ensure their accuracy . in this study , then , the whole transcript was read by the researchers several times to achieve a general sense of the text . then , the researchers read the text and fieldnotes carefully until the sentences turned into meaning units or units of analysis . afterward , using the words similar to the participants statements , the researchers derived primary codes . the primary codes were read several times and classified according to the differences and similarities . the classified codes were merged , by directed content analysis , into the larger themes derived from orem 's self - care concepts and subthemes . the researchers categorized the classified codes that were inappropriate for being classified as subthemes of orem 's concepts into independent subthemes . these classifications were eventually merged into one of the four themes of the study so that the fitness and inconsistency of the subthemes with the existing data could be examined by the original concepts . throughout gathering and analysis of the data , the researchers repeatedly moved back and forth among the questions , checked previous questions and refined them , developed hypotheses , and paid attention to the strengths and weaknesses . the data were examined in depth through extending the existing codes till all the levels of codes were completed and no new conceptual data were available to represent the new codes . to ensure the validity of the data in this study , the researchers used prolonged engagement , check member , peer review , searching for disconfirming evidence , research credibility , and audit . the process of the study and the consistency of the data were examined by the researchers . data analysis was conducted by maxqda software ( verbi gmbh , berlin , germany ) . a qualitative directed content analysis approach was used to conduct this study from may 2012 to may 2013 . sometimes , there is an available theory about a phenomenon that is incomplete or may benefit from further description . in this case , qualitative research practitioners might choose to use a directed approach to content analysis . the research ethics committee of the tehran university of medical sciences approved the protocol of this study ( approval no . the potential participants were recruited from a referral center , the shahrekord hepatitis services organization . a purposive sampling method was adopted , and the sampling continued until data saturation was achieved throughout the process of data collection . the inclusion criteria were diagnosis of chronic hepatitis by a specialist , referral of patients aged 18 years and over to behavioral counseling , and being consent to participate in the study . informed consent to participate in the study was provided by the participants prior to data collection . the data were collected from 22 semi - structured interviews with 18 participants and direct observation of their behaviors in the process of data collection by the researchers . the participants were interviewed individually in the local hepatitis services organization by the first author . however , the participants could freely choose to be interviewed either in their own place of choice or the local hepatitis services organization . to access reliable and rich information , the researchers created a close and direct relationship with the participants . the data were collected through semi - structured , in - depth interviews and fieldnotes that the researchers took as they observed the participants . to familiarize with the participants and gain their trust before the formal interview , the researchers met the participants individually in an informal session and then interviewed them in a formal session . prior to the interview , the researchers obtained the informed consent from the participants to record the interviews . throughout the process of interview , initially , the orem 's pattern - based general questions were asked . if the interview was diverted from the purpose of the study , the researchers redirected the interview toward the research purposes using illustrative questions . the number and duration of each interview session varied according to the content and conditions of the participants and depended on various factors including time , desire , the participants patience and strength , and physical and mental condition , the information level of participants , and environmental conditions . the patients were asked questions during interviews , such as what does hepatitis mean to you ? , did you face difficulty in relation to your illness ? , what should the attitude toward hepatitis management be ? , how has hepatitis affected your life ? , how do you do self - care activities ? , and what education have you had in relation to the disease and how does it affects social life ? before completion of each interview , the researchers offered all the participants the opportunity to add anything to the discussion by asking the following question : is there anything else that you feel is relevant and you would like to add ? immediately after each interview , the researchers transcripted the recorded interviews word by word and then checked the interview transcripts against the recorded audio tapes to ensure their accuracy . in this study , then , the whole transcript was read by the researchers several times to achieve a general sense of the text . then , the researchers read the text and fieldnotes carefully until the sentences turned into meaning units or units of analysis . afterward , using the words similar to the participants statements , the researchers derived primary codes . the primary codes were read several times and classified according to the differences and similarities . the classified codes were merged , by directed content analysis , into the larger themes derived from orem 's self - care concepts and subthemes . the researchers categorized the classified codes that were inappropriate for being classified as subthemes of orem 's concepts into independent subthemes . these classifications were eventually merged into one of the four themes of the study so that the fitness and inconsistency of the subthemes with the existing data could be examined by the original concepts . throughout gathering and analysis of the data , the researchers repeatedly moved back and forth among the questions , checked previous questions and refined them , developed hypotheses , and paid attention to the strengths and weaknesses . the data were examined in depth through extending the existing codes till all the levels of codes were completed and no new conceptual data were available to represent the new codes . to ensure the validity of the data in this study , the researchers used prolonged engagement , check member , peer review , searching for disconfirming evidence , research credibility , and audit . the process of the study and the consistency of the data were examined by the researchers . data analysis was conducted by maxqda software ( verbi gmbh , berlin , germany ) . using orem 's ontological concepts in this study , the researchers derived four themes of needs in matrix of time and space , self - care agency , need for change in self - care activities and consequences of hepatitis . demographic characteristics of the participants hepatitis in the matrix of time and space , based on the theory of self - care , consists of three subthemes : general needs , developmental needs , and health diversion needs . for the participants in the present study , vital needs such as water , food , and defecation were less influenced by the disease although hepatitis affected certain dimensions of their vital needs . the balance between work and recreation was one of the basic needs affected by hepatitis . a participant said : i feel dizzy , fall to the ground , and ca nt maintain my balance . how i would do sports ? social interactions were one of the basic needs influenced by hepatitis . concerning this , it was pretty bad especially when i was told that i should nt kiss anybody , communicate with anybody . the theme of self - care agency reflects the ability and limitations that participants encounter with regard to self - care activities . this theme consists of two subthemes , ability to practice self - care and deficit in self - care . in this theme , there was a variety of factors contributing to the participants ability to practice self - care . some participants trusted the healthcare personnel to avoid transmitting the infection to others and said the personnel that they had hepatitis , but they had less confidence in other people in this regard . a participant said : if i go to hospital or dentistry , i tell the physician or nurse i have hepatitis to take care of myself and other persons . we tell the doctors to use preventive tools , but as soon as common people who have no knowledge about hepatitis discover that we have hepatitis , they avoid us . one of the obstacles reported by the patients was the fact that despite the great efforts made by them , there were still no centers of self - care education for the patients . in this regard , another participant said : quitting addiction , hepatitis treatment , and education can facilitate self - care . but i did a lot of work but there was nowhere to go to be trained for self - care . change in self - care activities was one of the themes derived in this study . this theme refers to the need for change in self - care practices perceived by participants . the participants faced many challenges in practicing self - care activities , which prevented them from doing these activities satisfactorily . this theme consists of two subthemes : changes in the healthcare system and changes in the society and people . the participants reported that a major problem in the healthcare system was lack of accountability in dealing with hepatitis patients by healthcare team . a participant said : when i went to dentist and he was going to take out my tooth , i said to him hepatitis . another participant reported : i referred for surgery and told i have hepatitis b. i saw the personnel chattering with each other regularly , i asked them why you did nt operate me . after few hours , i saw that they took me to the ward and did nt operate me . i asked them why you did nt operate me and they replied that as soon as the doctor found that you have hepatitis , he refused to do surgery . the consequences of hepatitis represents the effects of disease on the participants daily life and activities that result from hepatitis acquisition . for the participants in the present study , the disease outcomes comprised a continuum , which developed compatibility on one extreme and fear on the other in the patients accordingly , the consequences of hepatitis comprised two subthemes , aura of fear and adaptation . family behavior was one of the important points , of which some of the participants complained strongly . family 's inappropriate behaviors led to exacerbation of the disease conditions in some of the participants . in this regard , one of the participants reported : my daughter , son , husband , sisters , and brothers also know that i have hepatitis and that 's why they do n't like to have relationship with me . when someone finds out , he blames the person and says he has hepatitis , and he is afraid of transmitting hepatitis to himself . another participant said : the day my wife discovered that i have hepatitis , when she brought me a cup of tea , threw the cup in the trash . people consider opium so bad , but consider hepatitis much worse than opium and look at it as a vampire . i do not care who trusts in god , just get whatever he wants . i just read the prayer and have faith in god , and god always tells me the truth . hepatitis in the matrix of time and space , based on the theory of self - care , consists of three subthemes : general needs , developmental needs , and health diversion needs . for the participants in the present study , vital needs such as water , food , and defecation were less influenced by the disease although hepatitis affected certain dimensions of their vital needs . the balance between work and recreation was one of the basic needs affected by hepatitis . a participant said : i feel dizzy , fall to the ground , and ca nt maintain my balance . social interactions were one of the basic needs influenced by hepatitis . concerning this , it was pretty bad especially when i was told that i should nt kiss anybody , communicate with anybody . the theme of self - care agency reflects the ability and limitations that participants encounter with regard to self - care activities . this theme consists of two subthemes , ability to practice self - care and deficit in self - care . in this theme , there was a variety of factors contributing to the participants ability to practice self - care . some participants trusted the healthcare personnel to avoid transmitting the infection to others and said the personnel that they had hepatitis , but they had less confidence in other people in this regard . a participant said : if i go to hospital or dentistry , i tell the physician or nurse i have hepatitis to take care of myself and other persons . we tell the doctors to use preventive tools , but as soon as common people who have no knowledge about hepatitis discover that we have hepatitis , they avoid us . one of the obstacles reported by the patients was the fact that despite the great efforts made by them , there were still no centers of self - care education for the patients . in this regard , another participant said : quitting addiction , hepatitis treatment , and education can facilitate self - care . but i did a lot of work but there was nowhere to go to be trained for self - care . change in self - care activities was one of the themes derived in this study . this theme refers to the need for change in self - care practices perceived by participants . the participants faced many challenges in practicing self - care activities , which prevented them from doing these activities satisfactorily . this theme consists of two subthemes : changes in the healthcare system and changes in the society and people . the participants reported that a major problem in the healthcare system was lack of accountability in dealing with hepatitis patients by healthcare team . a participant said : when i went to dentist and he was going to take out my tooth , i said to him hepatitis . another participant reported : i referred for surgery and told i have hepatitis b. i saw the personnel chattering with each other regularly , i asked them why you did nt operate me . after few hours , i saw that they took me to the ward and did nt operate me . i asked them why you did nt operate me and they replied that as soon as the doctor found that you have hepatitis , he refused to do surgery . the consequences of hepatitis represents the effects of disease on the participants daily life and activities that result from hepatitis acquisition . for the participants in the present study , the disease outcomes comprised a continuum , which developed compatibility on one extreme and fear on the other in the patients . accordingly , the consequences of hepatitis comprised two subthemes , aura of fear and adaptation . family behavior was one of the important points , of which some of the participants complained strongly . family 's inappropriate behaviors led to exacerbation of the disease conditions in some of the participants . in this regard , one of the participants reported : my daughter , son , husband , sisters , and brothers also know that i have hepatitis and that 's why they do n't like to have relationship with me . when someone finds out , he blames the person and says he has hepatitis , and he is afraid of transmitting hepatitis to himself . another participant said : the day my wife discovered that i have hepatitis , when she brought me a cup of tea , threw the cup in the trash . people consider opium so bad , but consider hepatitis much worse than opium and look at it as a vampire . i do not care who trusts in god , just get whatever he wants . others believed that prayer and meditation led to relaxation . in this regard , one of the participants said : i just read the prayer and have faith in god , and god always tells me the truth . from the subtheme of general self - care needs , two categories were derived , essential needs and basic needs . the participants expressed that hepatitis affected the satisfaction of food needs greatly , in terms of defecation , the balance between work and recreation , and between loneliness and social interaction as well as prevention of life - threatening risks so that they were faced with certain challenges to satisfy their basic needs . most of the participants complained of some problems such as insomnia , malnutrition , constipation , and depression . a study conducted on chronic hepatitis patients demonstrated fear , anxiety , depression , mental problems , insomnia , and weakness in the majority of participants . in a study on patients with chronic hepatitis , 31% of the patients experienced mental disorders and substance abuse that had negative impact on the patients quality of life . hepatitis and aids found that sleep disorders were the most common problems reported by the patients . the data obtained from the participants in this study showed that hepatitis negatively affected individual , familial , occupational , and psychological function in the patients and made some of them frustrated and isolated . the results of another study have indicated that chronic hepatitis heavily influences physical , psychological , and familial function , and generally the whole person so that it causes many challenges for the patients . another theme generated from the participants data was self - care consisting of two subthemes , ability to practice self - care and deficit in self - care . the participants in the present study made great efforts to take care of themselves , but they faced many obstacles . in a study on patients with chronic hepatitis , stigmatization and discrimination by the public and healthcare personnel were derived to be one of the factors resulting in the disturbance of self - care so that these patients were not motivated to do self - care and/or suffered from isolation , mental disorders , impatience , and community avoidance . study findings have indicated that patients with chronic hepatitis , in practicing self - care activities , encounter numerous problems because they have difficulty prioritizing their care plans and making various care programs similar . the data derived from transcripts of the interviews with patients living with chronic hepatitis in this study showed that several factors , including individual , family , community , and healthcare system , contributed to doing self - care activities . in addition , socioeconomic status , family support , public and private organizations , and health conditions affected self - care activities in the present study . because self - care includes diet , sleep , exercise , weight control , smoking , alcohol , drugs , personal hygiene , eating habits and healthy lifestyle promotion , prevention and periodical medical examination , the patients may face disturbances in self - care process if one of them is delayed . another theme was change in self - care such that the healthcare team not only disregarded the treatment of patients with hepatitis but also frustrated them with annoying , discriminatory , and irresponsible behaviors ; therefore , the patients did not trust the healthcare team and were not hopeful about their help and support . 's study indicated that healthcare team did not behaved hepatitis patients appropriately , evaded the patients , did not deliver appropriate healthcare services to the patients , and labeled and dealt with them discriminatorily . staff 's behavior toward hepatitis patients differed from their behavior toward patients with other diseases such as cancer so that healthcare team delivered acceptable medical services to hepatitis patients less frequently . the findings of the present study indicated that the participants complained of lack of delivering medical and dental services by physicians and dentists and/or reported that they refused to do these services when they were informed of the disease . a study indicated that physicians and dentists did not deliver suitable services to patients with hepatitis . another study of patients living with chronic hepatitis indicated that lack of suitable communication between treatment personnel and chronic hepatitis patients led to a decline in and/or no trust among the patients , and eventually isolation and failure to adhere to therapeutic and health regimen . in this study , the theme of consequences of hepatitis covers all conditions of the participants well , and gives the reader a new insight toward hepatitis . being informed of hepatitis acquisition caused numerous mental , familial , economic , and social difficulties for the participants so that an aura of fear and apprehension was developed in the participants and very hard , excruciating circumstances governed their life . a study reported that being afraid of acquiring hepatitis was the most important stressor in the lifetime of hepatitis patients . being informed of hepatitis acquisition can cause numerous negative effects on family and friends which make patients experience certain variations in family life . the adverse effects of acquiring hepatitis can destroy the patients family and their spouses may attempt to get a divorce and abandon them . according to the findings of a study , 78% of the spouses of patients with fear of family 's behavior causes the patients to hide their disease even from their spouse and children , which , may influence the self - care behaviors in patients with hepatitis . a study in china indicated that hepatitis patients were reluctant to let their family members be informed of their disease and attempted to hide their disease even from their spouses . fear of stigmatization negatively affects patients with hepatitis such that the patients in this study were severely afraid of being stigmatized and isolated by the community . as a result a study showed that people did not have a positive attitude toward hepatitis and blamed patients with hepatitis . the self - care model , developed in this study [ figure 1 ] , is a specific self - care model for chronic hepatitis . although hepatitis is a chronic health problem , the developed self - care model is not appropriate and applicable for other chronic conditions . however , the model can be tested and examined before its application in different cultural backgrounds . use of orem 's self - care theory can not meet the need for self - care in hepatitis patients because these patients have vital sexual , respect and belonging , physical , economical , and psychological - behavioral needs , and lack adequate knowledge about self - care . Output:	background : hepatitis is an inflammatory disease which has many adverse effects on patients life because of its chronic nature . since orem 's theory of self - care is a grounded theory , the concepts and applications of this theory in patients with chronic hepatitis who have special needs may lead to some challenges . the purpose of this study was to explore self - care in patients with chronic hepatitis.methods/design:a directed content analysis was used in this qualitative study . participants were recruited from a metropolitan area . data were collected through semi - structured interviews . the verbatim transcripts of the participants interviews were analyzed according to directed content analysis.results:in this study , four themes , suggested by orem , were drawn from the data according to directed content analysis . the codes generated from the data were classified into concepts and then the concepts were assigned into these four themes . these themes were needs in the matrix of time and place , self - care agency , need for change in self - care and consequences of hepatitis.conclusion:the use of orem 's self - care theory can not meet the need for self - care in hepatitis patients because these patients have vital sexual , respect and belonging , physical , economical , and psychological - behavioral needs , and lack adequate knowledge about self - care . consequently , the specific self - care model developed in this study helps health professionals identify self - care activities in patients with chronic hepatitis .
PubmedSumm2025	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: pyelonephritis is a serious infection of the upper urinary tract ( the kidneys and ureters ) that often requires hospitalization . patients with pyelonephritis often have chills that start suddenly , fever , and pain between the ribs and hips on one or both sides of the body . based on these symptoms computed tomography ( ct ) , which allows doctors to look inside the body , is useful for letting doctors determine whether something is blocking the urinary tract . this type of imaging is often used for diagnosing pyelonephritis ; some doctors believe that a condition called perirenal fat stranding ( pfs ) , in which swelling of the fat around the kidneys is visualized on ct , is a characteristic finding of pyelonephritis . however , it is not clear whether this belief is true . we found that pfs was not specific to pyelonephritis , ie , it could be associated with other conditions . we also found that ct was not the most cost - effective way to diagnose pyelonephritis , as patients symptoms and laboratory parameters were more relevant diagnostic tools . we believe that our study is important as it evaluated pfs with respect to pyelonephritis . urinary tract infections ( utis ) are very common . in the us , they account for ~8.1 million visits to health care providers annually.1 utis are termed differently depending on the part of the body affected ; an infection of the kidney / upper urinary tract is called pyelonephritis , while an infection of the urethra is called urethritis . among these infections , pyelonephritis can be complicated by sepsis , and in many cases , hospitalization is necessary for treatment.2 classical symptoms of pyelonephritis include an abrupt onset of chills , fever , and unilateral or bilateral flank pain with costovertebral angle ( cva ) tenderness.3 in addition to these physical findings , a combination of urine analysis ( via microscopy and/or reagent strip ) , urine cultures , and blood cultures may be used to diagnose pyelonephritis . however , the diagnosis of acute pyelonephritis can be difficult for clinicians , especially among elderly patients who present with unusual symptoms , such as abdominal pain , vomiting , and diarrhea . computed tomography ( ct ) is a useful diagnostic tool for providing comprehensive anatomical and physiological information that accurately characterizes both intra- and extrarenal pathological conditions.3 imaging with ct is not unusual in the diagnosis of acute pyelonephritis and is often used for patients who do not respond to antibiotics within 72 hours or patients with underlying conditions , such as pregnancy , diabetes mellitus , and immunocompromised state , among others.4,5 swelling of the kidney , hypoenhanced renal parenchyma , and pronounced perirenal fat stranding ( pfs ) are some of the ct findings that may accompany acute pyelonephritis.6,7 yu et al8 have reported that perinephric fat stranding was found in 29.1% of patients with acute pyelonephritis . some clinicians regard pfs as a characteristic finding of pyelonephritis ( figure 1).9 however , to the best of our knowledge , no study has reported on the factors that are associated with pfs as well as the sensitivity and/or specificity of pfs in diagnosing acute pyelonephritis . thus , we aimed to evaluate the correlation between age , renal function , and frequency of pfs . this retrospective observational study was conducted at the japanese red cross ishinomaki hospital , ishinomaki , miyagi , japan . this study was approved by the ethical committee of japanese red cross ishinomaki hospital ( approval no 16 - 28 ) and was conducted in accordance with the 1964 declaration of helsinki and its later amendments or comparable ethical standards . the ethics committee deemed written informed consent not necessary due to the retrospective nature of the study . the medical records of all patients who were admitted to the japanese red cross ishinomaki hospital in the miyagi prefecture of japan between december 2013 and july 2015 were reviewed . of 468,624 patients , we selected 120 patients with the 10th revision of the international statistical classification of diseases and related health problems ( icd-10 ) code n10 in their medical records . the diagnosis of acute pyelonephritis was based on physical examination ( presence of fever , chills , and cva tenderness ) and laboratory findings ( increased white blood cell [ wbc ] count , elevated c - reactive protein [ crp ] , and demonstration of pyuria and bacteriuria on urine testing ) . we excluded eleven cases of children ( aged < 16 years ) and two cases that did not demonstrate clear findings of acute pyelonephritis . of the remaining cases , 89 had undergone ct evaluation of both kidneys ; we evaluated these patients for the presence or absence of pfs as the pyelonephritis group . these patients underwent percutaneous renal biopsy at our hospital between january 2013 and may 2016 ; each of them had undergone ct examination the day after the renal biopsy to check for bleeding complications.10 we had ruled out acute pyelonephritis prior to percutaneous renal biopsy in this group , given that acute pyelonephritis is a contraindication for renal biopsy . in this control group , ct images were obtained using an aquilion cxl system ( toshiba medical systems corp . , tochigi , japan ) . normally distributed data are presented as mean standard deviation ( sd ) , with nonparametric data presented as the median and 25th75th percentile . statistical analyses were performed using the student s t - test for parametric data , the mann whitney u test for nonparametric data , fisher s exact test for nominal categorical valuables , and logistic regression analysis for the following covariates : age , sex , and estimated glomerular filtration rate ( egfr ) . we performed 1:1 matching based on the propensity score , as patient characteristics of both the pyelonephritis and control groups were different.11,12 the purpose of using the propensity score was to reduce the influence of bias by aligning the backgrounds of both groups . in this study , logistic regression was performed with the presence or absence of pyelonephritis as a dependent variable , and age , sex , and renal function as independent variables . a propensity score for whether or not pyelonephritis develops from a background factor was prepared . all statistical data were analyzed using ezr version 3.3.1 ( saitama medical center , jichi medical university , saitama , japan ) , which is a graphical user interface for r ( the r foundation for statistical computing , vienna , austria).13 this retrospective observational study was conducted at the japanese red cross ishinomaki hospital , ishinomaki , miyagi , japan . this study was approved by the ethical committee of japanese red cross ishinomaki hospital ( approval no 16 - 28 ) and was conducted in accordance with the 1964 declaration of helsinki and its later amendments or comparable ethical standards . the ethics committee deemed written informed consent not necessary due to the retrospective nature of the study . the medical records of all patients who were admitted to the japanese red cross ishinomaki hospital in the miyagi prefecture of japan between december 2013 and july 2015 were reviewed . of 468,624 patients , we selected 120 patients with the 10th revision of the international statistical classification of diseases and related health problems ( icd-10 ) code n10 in their medical records . the diagnosis of acute pyelonephritis was based on physical examination ( presence of fever , chills , and cva tenderness ) and laboratory findings ( increased white blood cell [ wbc ] count , elevated c - reactive protein [ crp ] , and demonstration of pyuria and bacteriuria on urine testing ) . we excluded eleven cases of children ( aged < 16 years ) and two cases that did not demonstrate clear findings of acute pyelonephritis . of the remaining cases , 89 had undergone ct evaluation of both kidneys ; we evaluated these patients for the presence or absence of pfs as the pyelonephritis group . these patients underwent percutaneous renal biopsy at our hospital between january 2013 and may 2016 ; each of them had undergone ct examination the day after the renal biopsy to check for bleeding complications.10 we had ruled out acute pyelonephritis prior to percutaneous renal biopsy in this group , given that acute pyelonephritis is a contraindication for renal biopsy . in this control group , normally distributed data are presented as mean standard deviation ( sd ) , with nonparametric data presented as the median and 25th75th percentile . statistical analyses were performed using the student s t - test for parametric data , the mann whitney u test for nonparametric data , fisher s exact test for nominal categorical valuables , and logistic regression analysis for the following covariates : age , sex , and estimated glomerular filtration rate ( egfr ) . we performed 1:1 matching based on the propensity score , as patient characteristics of both the pyelonephritis and control groups were different.11,12 the purpose of using the propensity score was to reduce the influence of bias by aligning the backgrounds of both groups . in this study , logistic regression was performed with the presence or absence of pyelonephritis as a dependent variable , and age , sex , and renal function as independent variables . a propensity score for whether or not pyelonephritis develops from a background factor was prepared . all statistical data were analyzed using ezr version 3.3.1 ( saitama medical center , jichi medical university , saitama , japan ) , which is a graphical user interface for r ( the r foundation for statistical computing , vienna , austria).13 the characteristics of the enrolled patients are presented in table 1 . compared with patients in the control group , patients in the pyelonephritis group were significantly older ( 7415 years vs 6316 years , respectively , p<0.001 ) and had a significantly higher frequency of pfs ( 72% vs 39% , respectively , p<0.01 ) . other findings are as follows : 25 ( 28% ) and eight ( 9.1% ) patients in the pyelonephritis group had underlying diabetes and a history of immunosuppressant use compared to 195 ( 61% ) and 62 ( 20% ) patients , respectively , in the control group . a total of 26 ( 52% ) patients in the pyelonephritis group had cva tenderness . in terms of serological findings , the median wbc count was 11,300/l ( interquartile range ( iqr ) : 8,10017,600/l ) in the pyelonephritis group vs 6,100/l ( iqr : 4,1007,700/l ) in the control group , while the median crp was 12 mg / dl ( iqr : 4.718 mg / dl ) in the pyelonephritis group vs 0.13 mg / dl ( iqr : 0.050.56 mg / dl ) in the control group . the median lactate level was 1.6 in the pyelonephritis group ( lactate was not measured in the control group ) . in terms of urinary findings , the positive rate of wbc was 83% ( 70/85 ) in the pyelonephritis group vs 20% ( 63/318 ) in the control group , that of nitrite was 37% ( 31/85 ) in the pyelonephritis group vs 4.4% ( 14/318 ) in the control group , and that of wbc or / and nitrite was 93% ( 79/85 ) in the pyelonephritis group vs 21% ( 67/318 ) in the control group . bacteria were identified in urine cultures in 92% ( 71/77 ) of patients in the pyelonephritis group . ct imaging detected urinary stones in 40% ( 36/89 ) of patients in the pyelonephritis group vs 12% ( 39/319 ) of patients in the control group , hydronephrosis in 47% ( 42/89 ) of patients in the pyelonephritis group vs 3.8% ( 12/319 ) of patients in the control group , and renal emphysema in 3.4% ( 3/89 ) of patients in the pyelonephritis group vs 0.31% ( 1/319 ) of patients in the control group . among the 89 patients in the pyelonephritis group , 51 were subjected to contrast ct ; of these , 22 ( 43% ) had areas where renal contrast was insufficient . for all enrolled patients , the sensitivity , specificity , and positive likelihood ratio of pfs in diagnosing acute pyelonephritis were 72% ( 95% confidence interval [ ci ] , 6181 ) , 71% ( 95% ci , 6676 ) , and 2.5 ( 95% ci , 2.03.1 ) , respectively ( table 2 ) . based on our empirical speculation that pfs was affected by age , sex , and renal function , we divided patients into two groups for each of the variables : age ( younger vs older ) , sex ( male vs female ) , and renal function ( high vs low ) ; the frequency of pfs was determined for each group ( figures 35 ) . in the pyelonephritis group , ( aged < 70 years ) vs 73% in older patients ( aged 70 years ) , 84% in men vs 67% in women , and 80% in those with egfr < 35 ml / min/1.73 m vs 64% in those with egfr > 35 ml / min/1.73 m. no significant differences were found between any of these groups . in the control group , the frequency of pfs was 29% in younger patients vs 54% in older patients , 47% in men vs 28% in women , and 51% in those with egfr < 35 ml / min/1.73 m vs 28% in those with egfr > 35 ml / min/1.73 m. the reason for dividing egfr into more or less than 35 ml / min/1.73 m was because the median value of both the pyelonephritis and control groups was ~35 ml / min/1.73 m. the kidney disease : improving global outcomes guidelines classify the severity of chronic kidney disease by gfr.14 gfr 3044 is classified as g3b category ( moderately to severely decreased ) , and gfr 1530 is classified as g4 category ( severely decreased ) . therefore , the 35 cutoff values used in this study were considered to be reasonable in general , with no significant divergence from the severity classification by gfr . a multiple logistic regression model yielding odds ratios ( ors ) and 95% cis was then used to identify predictors of pyelonephritis . the model included age , sex , and egfr . in the multiple logistic regression model , the or for pfs to non - pfs was 5.0 ( 95% ci , 2.79.3 ; p<0.01 ) . next , we performed a post hoc 1:1 person match by assigning the two groups ( pyelonephritis group and control group ) to each case , matched for age , sex , and egfr on admission ( table 3 ) . conditional logistic regression analyses for these propensity score - matched data sets were used to estimate the ors and respective 95% cis.7 the or for pfs to non - pfs was 3.6 ( 95% ci , 1.96.7 ; p<0.01 ) . after adjusting for age , sex , and renal function using the propensity score analysis , the sensitivity , specificity , and positive likelihood ratio of pfs for diagnosing acute pyelonephritis was 72% ( 95% ci , 6181 ) , 58% ( 95% ci , 4869 ) , 1.7 ( 95% ci , 1.32.3 ) , respectively ( table 4 ) . using the mantel haenszel test , the or for pfs to non - pfs was found to be 2.9 ( 95% ci , 1.66.5 ; p<0.01 ) . this study revealed that pfs is affected by age , sex , and egfr , with increasing age , male sex , and lower egfr increasing the frequency of pfs . however , the presence of pfs itself was not a useful indicator of acute pyelonephritis . as shown in table 1 , pfs was found in 39% of patients , which seemed high compared to 29.1% reported by yu et al . however , in their study , the mean age of enrolled patients was 6017 years and 93% were females.8 in contrast , the mean age of enrolled patients in our study was 7415 years and only 72% were females . considering that older age and male sex increase the frequency of pfs , we believe that our findings were acceptable . previous reports did not include a control group , making it impossible to calculate the sensitivity and specificity . in our study , we included patients who had undergone renal biopsy as our control group , after acute pyelonephritis had been ruled out by the treating physician ( given that active uti is a contraindication for percutaneous renal biopsy ) . this allows us to calculate the sensitivity and specificity . in the control group , 67 ( 21% ) patients had positivity of wbc and/or nitrite in urinalysis ; however , all of them had asymptomatic bacteriuria . there were ~20% of patients with asymptomatic bacteriuria when the control group was compared to the whole population ; it might be related to the fact that the average age was higher and there were more diabetic patients and immunosuppressant users . although hypoenhanced regions in the kidney may be characteristic of acute pyelonephritis , they are not specific . for example , acute renal infarction and immunoglobulin g4-related renal disease ( igg4rd ) are accompanied by hypoenhanced regions in the kidney.15,16 in addition , acute renal infarction is also associated with fever , flank pain , and increased inflammatory markers , similar to acute pyelonephritis . however , elevated serum lactate dehydrogenase is found in almost all patients with acute renal infarction , and this may be useful in distinguishing acute renal infarction from acute pyelonephritis . these findings suggest that radiological imaging is not as useful as conventional examination in the diagnosis of acute pyelonephritis . the sensitivity of urinalysis ( > 10 wbcs / high power field [ hpf ] ) was reported to be 58%82% , while the specificity was 65%86%.17 the sensitivity of positive leukocyte esterase , nitrite , and leukocyte esterase plus nitrite tests was reported to be 74%96% , 35%85% , and 75%84% , respectively . in contrast , their specificity was 94%98% , 92%100% , and 82%89% , respectively.1821 the sensitivity of positive dipstick hematuria was 44% , while its specificity was 88%.22 the sensitivity of finding of > 1 bacterium per hpf by gram stain ( centrifuged urine ) was 93% , while the specificity was 95%.23 we demonstrated that the sensitivity of pfs in diagnosing acute pyelonephritis was 72% , while the specificity was 60% . this result shows that radiological imaging is not a particularly useful tool for the diagnosis of acute pyelonephritis . the emergency physician , in wanting to avoid missing life - threatening diseases , may order radiological imaging , including ct scan ; this may thus contribute to an increased number of ct scans performed . indeed , japan is reported to have one of the highest numbers of ct machines in the world.24,25 however , studies have shown that excessive use of diagnostic imaging is not necessarily associated with improved medical care.2628 in terms of cost , an abdominal ct scan ( ~\10,000 ) is much more expensive than a urine dipstick test ( \260 ) . as shown earlier , we believe that our findings provide a fair account of the usefulness of ct scan in diagnosing acute pyelonephritis . the ratio of the pyelonephritis group was 0.72 and that of the control group was 0.40 ; the alpha error was 0.05 , and even if the sample size was 89 , the detection power was as high as 0.99 . table 5 lists the various bacteria that were isolated from the positive urine cultures in our cohort ; the causative organisms were similar to previous reports in japan.29 we agree that our method of selection also created several limitations . primary renal disease such as glomerulonephritis , tubulointerstitial nephritis , and others may increase pfs , although the cause of renal dysfunction may have little effect on the frequency of pfs ( data not shown ) . another limitation that we did not consider was previous history of pyelonephritis , which we hope will be addressed in future studies . another limitation of our study was the exclusion of children aged < 16 years and pregnant women . however , this is because ct examination is not advisable for pregnant women and children . admittedly , the occurrence of pyelonephritis is particularly high among pregnant women , with an estimated incidence of 0.5%2%.30 therefore , it was unfortunate that we could not evaluate pfs in this population of patients . older age , male sex , and lower egfr are associated with an increased occurrence of pfs . however , with respect to the diagnosis of acute pyelonephritis , the presence of pfs itself did not have particularly high sensitivity and/or specificity compared to other tests . Output:	purposepyelonephritis , an upper urinary tract infection , is a serious infection that often requires hospitalization . however , the accurate diagnosis of acute pyelonephritis can be difficult , especially among older individuals who can present with unusual symptoms . imaging with computed tomography ( ct ) is not unusual in the diagnosis of pyelonephritis , with some clinicians regarding perirenal fat stranding ( pfs ) as a characteristic finding . however , the sensitivity and specificity of pfs in diagnosing pyelonephritis are currently unknown . we therefore sought to clarify the relevance of pfs in diagnosing acute pyelonephritis.patients and methodswe conducted a case - controlled retrospective analysis of medical records . the pyelonephritis group included 89 patients who had been diagnosed with acute pyelonephritis , while the control group included 319 patients who had undergone percutaneous renal biopsy . ct findings were available for both groups . the frequency of pfs and its sensitivity and specificity for the diagnosis of acute pyelonephritis were investigated.resultsthe mean ages of the pyelonephritis and control groups were 7415 years and 6316 years , respectively . a total of 28% of men were in the pyelonephritis group vs 61% of men in the control group . the frequency of pfs was 72% in the pyelonephritis group vs 39% in the control group . age and renal dysfunction were associated with an increased frequency of pfs . after adjusting for age , sex , and renal function using a propensity score analysis , the sensitivity , specificity , and positive likelihood ratio of pfs for diagnosing acute pyelonephritis were 72% , 58% , and 1.7 , respectively.conclusionthe presence of pfs was not useful in diagnosing acute pyelonephritis .
PubmedSumm2026	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: chemical modification of these compounds have led to a large number of mono- and di - s and n - substituted analogs showing therapeutic properties , especially antivirial , antithyroid , and antitumor activities [ 17 ] . the antimetabolites of 5,6-dimethyluracil 5-morpholinomethyl-6-methyl-2-thiouracil and 5-piperidinomethyl-6-methyl-2-thiouracil have been synthesized via the mannich reaction . however , to the best of our knowledge no work has been published on the synthesis as well as on physicochemical properties of the monochlorobenzylthio - substituted derivatives of these compounds . this fact has stimulated us to investigate the reaction of chlorobenzylation of 2-thio-5-piperidinomethyl-6-methylthiouracil and 2-thio-5-morpholinomethyl-6-methylthiouracil as well as the mannich reaction of 2-o-(m- and p-)chlorobenzylthio-6-methylthiouracils , formaldehyde , and piperidine ( or morpholine ) . recently novel pharmacological action of 2,4-di - o-(m- and p-)bromo-(chloro- and nitro-)benzylthio-5-bromouracils ( and 6-methyluracils ) as well as disulfides of n , o-(n , n- or o , o-)-di- and n , n , o - tri-(o- , m- and p-)-bromobenzyl-2-thiouracils has been found on the basis of the computer - aided drug discovery approach with the compounds program prediction of activity spectra for substances ( pass ) [ 1215 ] . since only the structural formula of the chemical compound is necessary to obtain a pass prediction , this approach was used in the present work . this paper deals with the synthesis and physicochemical properties of 16 . additionally , the analysis of biological activity spectra prediction for 16 made in this paper is a good example of in silico studies of chemical compounds . a series of six new 2-o-(m- and p-)chlorobenzylthio-5-piperidinomethyl-6-methyluracils 13 and 2-o-(m- and p-)chlorobenzylthio-5-morpholinomethyl-6-methyluracils 46 were synthesized by the reaction of 2-thio-5-piperidinomethyl-6-methyluracil or 2-thio-5-morpholinomethyl-6-methyluracil in 3 n naoh in methanol with o-(m- and p-)chlorobenzyl chlorides at room temperature for 24 h ( scheme 1 ) . compounds 1 , 3 , 4 , and 6 were also produced in well - known mannich reaction from 2-o-(and p-)chlorobenzylthio-6-methyluracils when refluxed with paraformaldehyde and piperidine ( or morpholine ) in ethanol ( scheme 2 ) . compounds 2 and 5 were not produced in the mannich reaction of relatively soft 2-m - chlorobenzylthio-6-methyluracils with paraformaldehyde and piperidine ( or morpholine ) when heated in ethanol ( scheme 2 ) . the 2 o-(m- and p-)chlorobenzylthio - containing compounds 16 were confirmed by examination of their uv / vis , ft - ir , h nmr and c nmr spectra ( tables 1 and 2 ) , and hmbc ( table 3 ) as well as elemental analyses ( table 4 ) . the h nmr and c nmr data of 16 are given in tables 1 and 2 , respectively . assignments of the h nmr and c nmr resonances of these compounds were deduced on the basis of the signal multiplicities and by the corrected application of two - dimensional nmr technique h - h cosy and hmbc . the singlets of c5ch2 of 16 are situated at 3.283.58 ppm , and the singlets of c6ch3 of 16 are situated at 2.222.33 ppm , respectively . the cnmr spectra of compounds 16 in dmsod6 showed characteristic signals in the range of 31.5532.81 ppm and 53.0453.71 ppm assigned to s ch2 and n ch2 , respectively . the thiouracil ring exhibited signals in the range 110.01114.71 ppm and 158.81162.11 ppm assigned to c5=c6 , respectively . the c nmr spectra of compounds 16 showed the presence of a methyl group from the 6-methyl-2-thiouracil ring at 20.9021.30 ppm . the hmbc spectrum clearly shows the connectivities of all hydrogen and carbon atoms involved , including quaternary carbons . the hmbc results allow an unequivocal assignment of s - substitution of benzyl group at uracil ring of 16 ( table 3 ) . the hmbc experiment is conducted without c decoupling so that correlations via one or more bond can be discerned and one - bond correlation affords double cross - peaks in the h dimension . for compounds 16 the h nmr spectrum exhibits three singlets at 4.294.46 , 3.283.58 , and 2.222.33 ppm ascribed to protons of s the hmbc of 16 shows peaks corresponding to two - bond correlations for c6ch3/c6 ( 158.81162.11 ppm ) and three - bond correlations for s the ft - ir spectra of 16 show absorption bands of medium intensities in the region 809820 cm assigned to c cl vibration as well as in the region 10351091 cm assigned to c cl vibration ( table 1 ) . the ft - ir spectra of 16 show also absorption bands in the region 28522864 cm assigned to ch2s as well as in the region 14441447 cm assigned to ch2s vibration ( table 1 ) . the uv / vis spectra of 16 show max in the range 245250 nm ( table 1 ) . in the present paper the biological activity spectra were predicted for all six synthesized compounds ( 16 ) using pass [ 1215 ] . we have also selected the types of activities that were predicted for a potential compound with the highest probability ( focal activities ) . the most frequently predicted types of biological activities are antiviral ( influenza ) , antiseborrheic , and prolyl aminopeptidase inhibitor . it ought to be pointed out that in the series of compounds 1 , 3 and 6 such activity as mucomembranous protector has also been predicted , as well as in the series of compounds 2 , 3 , 5 , and 6 such activity as prolyl aminopeptidase inhibitor . the reaction of 2-thio-5-piperidinomethyl-6-methyluracil and 2-thio-5-morpholinomethyl-6-methyluracil with o-(m- and p-)chlorobenzyl chlorides in 3 n naoh in methanol at room temperature leads to 2-o-(m- and p-)chlorobenzylthio-5-piperidinomethyl-6-methyluracils 13 and 2-o-(m- and p-)-chlorobenzylthio-5-morpholinomethyl-6-methyluracils 46 . the mannich reaction of 2-o-(and p-)chlorobenzylthio-6-methyluracils with paraformaldehyde and piperidine ( or morpholine ) refluxed in ethanol leads to 1 , 3 , 4 , and 6 . the results obtained by pass method of identification of the prospective pharmacological properties of 16 exhibit the possibility of finding new pharmacological agents from this class of compounds . the purity of all described compounds was checked by melting points , tlc , and elemental analyses . rf values refer to silica gel f254 tlc plates ( merck ) developed with chcl3 : ch3oh ( 10 : 1 ) and observed under uv light ( = 254 and 366 nm ) . uv / vis spectra were recorded with a specord uv / vis spectrophotometer in methanol . ir spectra were recorded with ft - ir bruker ifs-113 spectrophotometer in kbr pellets . the h nmr ( 300 mhz ) and c nmr ( 75 mhz ) spectra were determined with varian gemini 300 spectrometer in dmsod6 solution at a concentration between 0.25 and 0.40 m in the 5 mm sample tubes at ambient temperature . 2-thio-5-piperidinomethyl-6-methyluracil , 2-thio-5-morpholinomethyl-6-methyl - uracil , and 2-o-(m- and p-)-chlorobenzylthio-6-methyluracils have been obtained according to the literature . the synthesis of 2-o-(and p-)chlorobenzylthio-5-piperidinomethyl-6-methyluracils ( 1 and 3 ) and 2-o-(and p-)chlorobenzylthio-5-morpholinomethyl-6-methyluracils ( 4 and 6 ) is as follows . general procedure a : 0.2 g ( 0.8 mmole ) of 2-thio-5-piperidino-(morpholino-)-methyl-6-methyluracils [ 8 , 9 ] was dissolved with stirring in room temperature in 2.5 ml of 3 n naoh in methanol . next , to the solution 0.116 ml ( 0.9 mmole ) of o - chlorobenzyl chloride or 0.146 g ( 0.9 mmole ) of p - chlorobenzyl chloride was added . after stirring at room temperature for 24 hours the synthesis of 2-m - chlorobenzylthio-5-piperidinomethyl-6-methyluracils ( 2 ) and 2-m - chlorobenzylthio-5-morpholinomethyl-6-methyluracils ( 5 ) is as follows . general procedure a : 0.2 g ( 0.8 mmole ) of 2-thio-5-piperidino-(morpholino-)methyl-6-methyluracils [ 8 , 9 ] was dissolved with stirring in room temperature in 2.5 ml of 3 n naoh in methanol . next , to the solution 0.116 ml ( 0.9 mmole ) of m - chlorobenzyl chloride was added . the reaction mixture was stirred in room temperature for 24 hours and after that time 5 ml distilled water was added . the obtained crude product was collected by filtration , washed with distilled water , and dried in the exicator . the obtained dry solid was dissolved in 10 ml of chcl3 and separated by silica gel column chromatography ( merck 203400 mesh ) using the following solvent mixtures : chcl3 : ch3oh 50 : 1 ( 40 ml ) , 40 : 1 ( 40 ml ) , 30 : 1 ( 30 ml ) , 20 : 1 ( 30 ml ) , and 10 : 1 ( 20 ml ) . the fractions of 20 ml were collected . on the basis of analytical tlc fractions of product desired were obtained by combining 20 ml fractions . the synthesis of 2-o-(and p-)chlorobenzylthio-5-piperidinomethyl-6-methyluracils ( 1 and 3 ) and 2-o-(and p-)chlorobenzylthio-5-morpholinomethyl-6-methyluracils ( 4 and 6 ) is as follows . general procedure b : a mixture of 0.1 g ( 0.38 mmole ) 2-o-(or p-)chlorobenzylthio-6-methyluracils , 0.024 g ( 0.4 mmole ) paraformaldehyde , and 0.0035 ml ( 0.4 mmole ) morpholine ( or 0.0046 ml , 0.4 mmole piperidine ) was suspended in 6 ml of ethanol ( 99.8% ) and refluxed for 8 hours . the precipitated solid was isolated by filtration , dried in room temperature , and recrystallized from methanol . Output:	six new 2-o-(m- and p-)chlorobenzylthio-6-methyl-5-piperidino-(or morpholino- ) methyluracils have been prepared . the structures of these compounds were confirmed by spectroscopic ( ft - ir , uv - vis , 1h nmr , 13c nmr , and hmbc ) and elemental analyses . estimation of pharmacotherapeutic potential has been made for synthesized compounds on the basis of prediction of activity spectra for substances ( pass ) .
PubmedSumm2027	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: urticarial vasculitis is characterized by recurrent episodes of urticaria with biopsy evidence of leukocytoclastic vasculitis ( 1 ) . although urticarial vasculitis is frequently idiopathic , it has been reported to be associated with infection , malignancy , certain drugs , and connective tissue diseases such as systemic lupus erythematosus ( sle ) and sjogren 's syndrome ( 2 ) . here , we report a case wherein urticarial vasculitis preceded the manifestations of sle . in august 2005 , a 50-yr - old woman visited our hospital with a 3-month history of recurrent multiple urticarial rashes and wheals on the face , trunk and back . treatment with antihistaminic and prednisolone resulted in improvement of her skin lesions , and she thereafter received prednisolone intermittently depending on her symptom recurrence . after 1 yr , she visited our hospital due to a malar rash and pitting edema of both the legs and recurrent episodes of urticaria ( fig . laboratory tests revealed the following findings : hemoglobin level , 9.6 g / dl ; white blood cell ( wbc ) count , 1,970/l ( neutrophils 62% ) ; platelets , 26910/l ; total protein , 6.3 g / dl ; albumin , 2.7 g / dl ; blood urea nitrogen ( bun ) , 14 mg / dl ; creatinine , 1.0 mg / dl ; and c - reactive protein , 1.4 mg / l . a high titer of antinuclear antibodies ( ana ) was detected ( 1:640 , speckled ) , while the test for double - stranded dna antibody was negative . anti - smith , anti - ro , and anti - la antibodies were all positive . the c3 level ( 28.7 mg / dl ; normal range , 55 - 120 mg / dl ) , c4 level ( 6.3 mg / dl ; normal range , 20 - 50 mg / dl ) , ch50 level ( 6.7 u / ml ; normal range , 23 - 46 u / ml ) , and c1q level ( 6.6 mg / dl ; normal range , 10 - 20 mg / dl ) were markedly decreased , whereas the anti - c1q antibody level was increased to 23.3 u / ml ( normal level , < 10 u / ml ) . the c1 esterase inhibitor level was normal ( 25 mg / dl ; normal range , 6 - 25 mg / dl ) and cryoglobulin was not detected . urinary analysis showed protein , + + ; red blood cell ( rbc ) count , 6 - 10/high - power field ( hpf ) ; and 24-hr urine protein , 1.1 g / day , and the creatinine clearance rate was 44.0 ml / min . in particular , distinct infiltration of neutrophils and nuclear dust was noted at the walls of small vessels ( fig . igg was positive along the capillary loop , and c1q and c3 were positive in the mesangium . the diagnosis of sle was established because of the presence of a malar rash , proteinuria , leukopenia , high ana titer , and the presence of the anti - smith antibody . an intravenous steroid pulse ( 1.0 g / day3 days ) followed by a high - dose oral steroid ( 1 mg / kg ) were administered after the disappearance of the vasculitic wheal and angioedema . hematuria and proteinuria were resolved , and mycophenolate mofetil ( 500 mg , bid ) and hydroxychloroquine were added to the therapy . however , in july 2007 , the tapering of the steroid administration was accompanied by the recurrence of urticaria , angioedema , conjunctivitis , and episodes of abdominal pain . despite mycophenolate mofetil , dapsone and steroid therapy did not completely relieve , but instead attenuated the urticarial vasculitic lesions . urticarial vasculitis resembles urticaria , is usually painful or nonpruritic , and typically persists for more than 24 hr ( 1 ) . patients with urticarial vasculitis can be divided into two groups , those with normal complement levels and those with huv ( 1 , 2 ) . the latter are more likely to exhibit systemic manifestations , including constitutional symptoms ( fever , malaise , and fatigue ) , arthralgia , arthritis , serositis , glomerulonephritis , interstitial nephritis , and raynaud 's phenomenon ( 2 ) . angioedema - like lesions are present in as many as 40% of patients , frequently involving the lips , tongue , periorbital tissue , and hands ( 2 ) . some patients have ocular inflammation like conjunctivitis and episcleritis ( 3 ) . by definition , complement measurements with low c1q and c4 levels and variably decreased c3 levels indicated activation of the classical pathway . c1q precipitins were identified and were later confirmed to be autoantibodies against c1q ( anti - c1q autoantibody ) ( 4 ) . antibodies against c1q act as a diagnostic marker for huv ( 1 , 3 ) . however , they are not specific to huv and have also been observed in sle . anti - c1q antibodies were detected in 30% of patients with sle and 80% of sle patients with glomerulonephritis ( 5 ) . this patient also showed decreased levels of c3 , c4 , ch50 , and c1q and increased levels of the anti - c1q antibody . she simultaneously exhibited huv and sle with glomerulonephritis , both of which are possible causes of decreased complement and the presence of anti - c1q antibody . in most cases , the histological findings of urticarial vasculitis include a majority of features of completely developed leukocytoclastic vasculitis ( 2 ) . direct immunofluoroscence of skin biopsy samples from huv patients usually show immunoglobulin and complement deposition in a granular pattern in and around the blood vessels of the upper dermis and the basement membrane ( 2 ) . she experienced recurrent urticaria with vasculitis , episodes of angioedema and eye inflammation , and the serum anti - c1q autoantibody was positive . she also fulfilled the criteria of sle with urticarial lesions that persisted for more than 24 hr and left a brownish pigmentation , which is different from true urticaria . some authors suggest the possible progression of huv to sle , and others have defined huv as an sle subtype ( 6 , 7 ) . at times , a hypocomplementemic patient initially diagnosed with an idiopathic huv has an underlying disease such as sle or sjogren 's syndrome ( 1 ) . huv is associated with sle in most cases ( 54% ) , but normocomplementemic urticarial vasculitis is not strongly associated with sle ( 2% ) ( 8) . this patient suffered from urticaria and angioedema for several years . serum complement levels were not checked initially , and a skin biopsy was performed 1 yr after disease onset ; the results revealed urticarial vasculitis . however , in this case , we assume that urticarial vasculitis was the presenting symptom of sle . huv has been reported to be recurrent in some patients , with an unpredictable response to therapy ( 1 , 2 ) . intravenous methylprednisolone and cyclophosphamide or high - dose oral corticosteroids , colchicines , dapsone , hydroxychloroquine , and low - dose methotrexate have all been reported to be effective treatments ( 1 , 2 , 9 ) . in this patient , the skin lesions resolved initially with steroids , but were subsequently refractory to treatment . mycophenolate mofetil , which was administered for lupus nephritis , did not affect urticarial vasculitis . in conclusion , although huv resembles sle both clinically and immunologically , an association huv and sle is not clear yet . Output:	urticarial vasculitis is characterized clinically by urticarial skin lesions and histologically by leukocytoclastic vasculitis . hypocomplementemic urticarial vasculitis is associated with connective tissue diseases such as systemic lupus erythematosus ( sle ) . we report a case of urticarial vasculitis that preceded manifestations of sle .
PubmedSumm2028	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the sessions take place 5 days per week with a duration of 1hour delivering 600 contractions . this is how we deliver the paradigm : basically , the rats are restrained in a plastic tube , in which they are free to move . you can actually see what these contractions here look like once every 6 seconds . so , here is what we did to test this paradigm . what happens if we use this paradigm after nerve transection and stimulate the muscle for two weeks ? one group had the right hind limb stimulated for two weeks starting 2 days after surgery . we made two incisions : one for nerve transection and one to implant the electrodes . the two parameters we looked for were motor unit number estimation ( mune ) and immunohistochemistry for endplate staining . for mune there are two ways to estimate it : by electromyography ( emg ) or by muscle force . what we did was to connect the stimulator proximal to the injury and then two electrodes in the muscle . then we can calculate the mu count by dividing the maximum response by the average response from the 15 - 20 templates . after two weeks the motor unit numbers in the muscle were significantly higher , a more than 2-fold increase ( figure 1 ) . when you look at the emg you can see that there is a more robust emg . clearly , there is more reinnervation taking place . by immunohistochemistry , the endplates may be stained in red , while in green may be labelled the axons . it is pretty clear that the stimulated muscles contained greater numbers of innervated endplates and intramuscular axons . they showed a greater number of axons in the distal stump of stimulated muscle . observing a nerve section from a rat that did not receive es , proximal to be nerve injury all the axon appeared intact while at the injury site , even at high resolution , the site is pretty dark , which means there were not many axons growing . in contrast , in a stimulated rat at the site of injury there were many more axons at both high and low resolution . it was pretty clear that our muscle stimulation paradigm of early stimulation works and increases motor reinnervation . a further question is : what happens in the long run if we keep stimulating throughout the reinnervation period ? the answer is a next study , in which es lasted for 1 , 2 or 3 months . we also looked at functional metrics with the walking track analysis and tapered beam tests . we divided them into 5 categories : normal reinnervation , reinnervated by a nodal sprout , reinnervated by a terminal sprout , reinnervated but unknown ( we do nt know where the axon was from ) and of course there were denervated endplates . in fact , if you combine all those reinnervated only 10% of the muscle endplates were denervated , the rest were reinnervated . so , it might be that electrical stimulation reduces terminal sprouting , we could say this is bad , but look at the percentage : it s only 5% ( figure 3 ) . the next thing we analyzed was the functional metrics over 3 months . every week , for three months we took behavioral tests to observe functional recovery . the walking track analysis was for measuring overground locomotion and the tapered beam test for measuring skilled locomotion . for the walking track test we dip the rat s feet in ink and let them walk across a piece of paper . what we had to measure were the toe spread , the print length and the intermediate toe spread and you plug these data into a formula and you get a number called the tibial function index . we found out that it is not a really good metric for tibial nerve injuries . we have full slips and half - slips , which allows us to calculate the slip percentage . after 6 weeks we noticed the rats stopped using their backwards sliding motion they used before . finally , a video at 3 months showed that the unstimulated rats are still slipping a lot . after week 4 there is a separation taking place between unstimulated and stimulated groups of rats . in summary it is clear proof that stimulation increases functional recovery and that he reinnervation is not adversely affected at the neuromuscular junction level . the early enhancement of reinnervation can be potentially explained by evidence that stimulation of the proximal nerve stump following injury can increase the rate of axonal outgrowth across the injury site . data from earlier this week on bdnf and gdnf show that both were significantly upregulated 2 weeks after injury ( figure 5 ) . if we change the muscle , intensity and so on , can we achieve similar results ? also , we want to examine the distal stump factors during the first weeks of stimulation following nerve injury and repair . what about the combination of proximal nerve stimulation during nerve repair with daily muscle stimulation ? does muscle stimulation affect synkinesis ( nerve misdirection ) ? if you cut a nerve then the axons are not going to grow back to the original muscles . we have the soleus and the lateral gastrocnemius and they are innervated by a common branch - the lateral gastrocnemius - soleus nerve . we inject some blue dye into the soleus and this goes to the spinal cord after some time . then we do a nerve transection and repair and connect the stimulator to the soleus . after 2 months we cut that soleus nerve and we basically put the nerve in a well , for which we use red color . ideally , all the axons labeled by injection will be labeled when you dip this nerve in the well . the idea behind all this is to take all these spinal cords and we take some sections and we quantify these labeled neurons . we want to see where these motoneurons are and also we want to see if they come from the same population after injury . audience question : how stable is the reinnervated phenotype if you stop the electrical stimulation ? so far , it looks like there is , but it still needs some more research . audience question : we have heavy constraints from our surgeons because they are not open to stimulation after the surgery and during repair . clinically , i can say that we had four pediatric patients that received nerve stimulation . we did the stimulation on these patients during nerve repair for 1h and it was no problem . audience question : have it been single twitches for the stimulation of the hind leg ? kern : we have to take in account , if we stimulate seniors , we should avoid any negative effect on the regeneration process . therefore , we invited mike willand . for me it is important that we avoid damage and that we do not influence the nerve regeneration in seniors . so , i am very thankful and happy . when i saw this for the first time , it was not clear if the muscle stimulation was enhancing or inhibiting the axon regeneration . here now we can be sure that the fes stimulation will not disturb the eventual nerve reinnervation process . audience question : how stable is the reinnervated phenotype if you stop the electrical stimulation ? so far , it looks like there is , but it still needs some more research . audience question : we have heavy constraints from our surgeons because they are not open to stimulation after the surgery and during repair . clinically , i can say that we had four pediatric patients that received nerve stimulation . we did the stimulation on these patients during nerve repair for 1h and it was no problem . audience question : have it been single twitches for the stimulation of the hind leg ? kern : we have to take in account , if we stimulate seniors , we should avoid any negative effect on the regeneration process . therefore , we invited mike willand . for me it is important that we avoid damage and that we do not influence the nerve regeneration in seniors . so , i am very thankful and happy . when i saw this for the first time , it was not clear if the muscle stimulation was enhancing or inhibiting the axon regeneration . here now we can be sure that the fes stimulation will not disturb the eventual nerve reinnervation process . Output:	electrical muscle stimulation following peripheral nerve injury has been a controversial method of treatment due primarily to the inconsistent literature surrounding it . in this presentation transcript i outline ongoing experiments investigating a clinically translatable daily muscle stimulation paradigm in rats following nerve injury . results show that reinnervation of muscle and functional behavioural metrics are enhanced with daily stimulation with upregulation of intramuscular neurotrophic factors as a potential mechanism . in addition , the impact of stimulation on terminal sprouting , a mentioned negative aspect of electrical muscle stimulation , was a minor contributor to long term functional reinnervation of stimulated muscles in our studies .
PubmedSumm2029	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: when skeletal muscle fibers are excited , a cascade of events is triggered , which ultimately leads to forces exerted on the skeleton . muscle forces are needed for movements , such as locomotion , and the maintenance of body balance . there are many structures involved at several levels organization : from actin , myosin , and titin of the sarcomeric cytoskeleton , desmin of the intermyofibrillar cytoskeleton , transsarcolemmal proteins such as dystrophin and integrin of the subsarcolemmal cytoskeleton , endo- , peri- , and epimysium , as well as tendon and aponeurosis at the muscle level to inter- and extramuscular connective tissues ( e.g. , the neurovascular tract , fascia ) at the compartmental level . for understanding how forces are transmitted from sarcomere to the bony skeleton in normal and diseased muscle , it is necessary to investigate the role of each of these structural elements as well as their interaction . the present review focuses on the connective tissues that are found in the direct environment of skeletal muscles and their potential effects in muscle function during movement . the most recognized pathway of force transmission from muscle fibers to bone is via the specialized myotendinous junction and tendon , named myotendinous force transmission . classical anatomy has defined each muscle as a separate entity with a unique function at the joint(s ) it spans . this is readily apparent from biomechanical models of the musculoskeletal system in which muscles are connected to the skeleton at their origin and insertion [ 2 , 3 ] . however , many scientists from the last century were aware of mechanical interactions between muscles ( for a historical overview see ) . for example , during measurements of soleus muscle forces in the cat , upon stimulation of the lateral gastrocnemius - soleus nerve branch denny - brown noticed that it is found to be extremely difficult to avoid a slight early rise of tension , and fall in the plateau , due to the vibration or pull of gastrocnemius . more recently , nichols stated mechanical artifacts due to direct mechanical action of the stretched muscle on those isometrically constrained were indicated by essentially instantaneous latencies or by effects observed after pharmacological block of heterogenic reflexes . note that in these experiments the tendons are severed from their insertion site and individually connected to force transducers . this means that the mechanical linkage was provided by structures at the muscle belly boundary ( i.e. , the epimysium ) . the purpose of this article is first to review the initial series of systematic experiments on mechanical interactions between synergistic muscles ( i.e. , neighboring muscles which produce the same movement at the joint ) via connective tissue linkages ( named epimuscular myofascial pathways ) that revealed the presence and capacity of this phenomenon ( mechanical interactions between antagonistic muscles have been reviewed elsewhere ; see ) ; second , to discuss the current debate on the importance of epimuscular myofascial force transmission during normal movements ; and third , to discuss the potential functions of inter- and extramuscular connective tissues for pathological muscle - tendon conditions . epimuscular myofascial force transmission is defined as transmission of muscle forces to the skeleton via pathways other than the muscular origin and insertion . a direct proof of epimuscular myofascial force transmission is a difference in force exerted at the origin ( proximal ) and insertion ( distal ) of a muscle . another feature of this phenomenon is that length changes in one muscle can affect forces exerted at the tendons of muscles that are kept at a constant length . two epimuscular pathways are distinguished ( figure 1 ) : ( i ) intermuscular , if force is transmitted between two neighboring muscles via the continuous connective tissue at their muscle belly interface , and ( ii ) extramuscular , if force is transmitted between the epimysium of a muscle and an adjacent nonmuscular structure . the direct intermuscular pathway is provided by an areolar connective tissue layer at the interface between muscle bellies ( for images see ) . several structures provide an anatomical substrate for the extramuscular myofascial pathway : ( i ) the matrix supporting nerves and blood vessels , that is , the neurovascular tract ( see [ 8 , 9 ] ) ( note that the neurovascular tract is continuous with the extensive intramuscular connective tissue network , which reinforces the nerves innervating muscle fibers and the blood vessels entering the muscle . ) , ( ii ) fascia layers forming the borders of synergistic muscle groups that are continuous with more superficial layers ( e.g. , subcutaneous connective tissue ) , and ( iii ) connective tissue around tendons ( for images the reader is referred to previous publications , e.g. , [ 1013 ] ) . dissection of a limb shows a vast network of collagen - based structures linking muscles together . clearly muscles are connected by fascial sheets , loose areolar tissue , vascular links , nerves , and supporting collagen . sometimes muscle fibers originate from neighboring muscle ( e.g. , in the cat there are some lg fibers that seem to terminate in mg muscle ) . tendons appear to run together . pushing or pulling on one muscle leads to movement of a neighbor . the question is how significant these links are to the normal function of muscle . an in - depth analysis of transmission of extensor digitorum longus ( edl ) muscle force in the rat , which is embedded within the anterior crural compartment together with extensor hallucis longus ( ehl ) and tibialis anterior ( ta ) muscles , has been performed . because both the proximal and distal tendons of edl can be attached to force transducers isometric forces were measured simultaneously at the proximal and distal tendons of edl muscle as well as at the tied distal tendons of ta , and ehl muscles . these tendons can all be dissected with minimal disruption of the compartment , leaving epimuscular myofascial pathways mostly intact . by manipulating the position of the force transducers , the muscle - tendon complex length of one or all muscles as well as muscle relative position length changes of the ta + ehl complex affected the forces exerted at the proximal and distal tendons of edl , which was kept at a constant length . more specifically , lengthening ta + ehl distally increased proximal edl force ( by 37% ) , but decreased distal edl force ( by 39% ) . the mechanical interactions between synergistic muscles can be ascribed to changes in the position of one muscle relative to the other and , consequently , changes in the configuration ( length and angle ) of inter- and extramuscular connective tissues ( figure 2 ) . it may appear obvious to explain these results by mechanical effects of intermuscular connective tissue . however , edl , ta and ehl are also linked to each other via extramuscular structures . a clear example is the neurovascular tract that runs in between the muscles while giving off branches of nerves and blood vessels which enter the endo - perimysial network of the muscle [ 8 , 9 ] . equal experimental conditions were imposed before and after disruption of the connective tissue layer between edl and ta + ehl , thereby eliminating force transmission via intermuscular myofascial pathways . this significantly decreased the effects of ta + ehl length on force exerted at the distal tendon of edl . however , the interaction between ta + ehl and proximal edl force did not change . therefore , we concluded that mechanical interactions between synergistic muscles originate from both inter- and extramuscular connective tissues . besides the above - described study , there is only one other study that reports data indicating that the areolar connective tissues are stiff and strong enough to transmit force . changes in the length - force characteristics were found following disruption of the intermuscular myofascial pathway . in the above - described studies , relative displacements of muscle bellies were the result of length changes in a single muscle group . to distinguish between effects of muscle length and relative position , isolated effects of muscle relative position were studied . the muscle - tendon complex length of edl and ta + ehl was kept constant . the position of edl muscle relative to its surroundings was changed by moving both the proximal and the distal tendons to an equal extent and in the same direction . displacements of edl in distal direction decreased force exerted at the distal tendons of ta + ehl . simultaneously , distal edl force increased and proximal edl force decreased . each movement affected the proximo - distal force difference and , thus , the magnitude of net epimuscular myofascial force transmission . in addition , the sign of the force difference between proximal and distal edl forces changed . length - force characteristics of edl muscle obtained by movements of the distal tendon were significantly different from its length - force characteristics if edl muscle was lengthened by moving its proximal tendon . in conclusion , the position of a muscle relative to surrounding tissues codetermines isometric muscle force . position effects can be explained by changes in the configuration of the tissues representing the epimuscular myofascial pathways ( figure 2 ) . in general , the muscle end that is positioned farthest in a particular direction ( e.g. , distal ) the above in situ studies have shown the potential of force transmission between skeletal muscles via inter- and extramuscular connective tissues . the functional relevance of this phenomenon is dependent on the magnitude of the effects found in physiological muscle conditions . however , this mode of force transmission may be small in normal muscles during physiological conditions , because ( 1 ) the above studies all used tetanic stimulation . this rarely occurs during voluntary movement , so observations may be relevant only to lab conditions ; ( 2 ) the muscle - tendon complex length of a single muscle was changed while the length of its synergists was kept constant , compared to simultaneous length changes in synergistic muscles during normal movements ; ( 3 ) when individual muscles are stimulated alone and together , the force sums linearly which is surprising if the epimuscular pathway is used ; and ( 4 ) a recent experiment studying force transmission between cat soleus ( so ) and its synergistic muscles in an intact animal showed little epimuscular force transmission . each of these points will be discussed below . in the studies described up to this point , the effects of epimuscular pathways on muscular force transmission were tested predominantly during simultaneous maximal activation of both synergistic and antagonistic muscles . coactivation of synergistic and antagonistic muscles has been observed in the awake , freely moving animal ( e.g. , ) , but in most cases at submaximal levels of activation ( e.g. , ) . recently , using the in situ setup described above , substantial proximo - distal edl force differences ( up to 30% of maximal force at each frequency ) as well as mechanical interactions with ta + ehl were found during nerve stimulation at submaximal frequencies ( 1030 hz ) . this suggests that also at firing frequencies encountered in vivo muscle forces can be transmitted via epimuscular myofascial pathways . another experimental condition of the studies described in section 2.2 that was different from the conditions under which muscles function in vivo was changing the length of only one muscle . due to differences in moment arms between synergists [ 21 , 22 ] , the change in length of one muscle can be different from that of its neighbor , but the relative movements imposed during lengthening a single muscle were beyond the physiological range . recently , this issue was addressed by investigating proximal - distal force differences in edl muscle while lengthening edl , ta , and ehl simultaneously , as is the case during ankle movements . also in these experimental conditions , a large force difference ( up to 30% of maximal force ) was found . at submaximal stimulation frequencies , however , the difference ( 5% ) and , hence , net epimuscular myofascial force transmission became small . it should be noted that different myofascial pathways can be arranged in such a way that they exert forces on a muscle in opposite direction ( see [ 9 , figure 8 ] ) and that the proximo - distal force difference is the net result of all myofascial loads . a small difference can thus be explained by limited epimuscular myofascial force transmission or opposing myofascial loads of similar magnitude . if force transmission through epimuscular pathways is substantial , then nonlinear summation of force is expected when different muscles are activated alone and together . nonlinear force summation is defined as the difference between the force exerted when two muscle parts are excited simultaneously and the sum of the forces exerted when each muscle part is excited individually . force transmission between the medial gastrocnemius ( mg ) and lateral gastrocnemius / soleus muscles ( lg / so ) was studied in the cat hindlimb [ 25 , 26 ] . the muscles were activated by stimulation of the nerve branches to each of the muscle groups . lg and so muscles were stimulated together because of the difficult surgery required to separate their nerves ( see ) . the cat hindlimb was left intact and the foot attached to a 6-degree - of - freedom ( dof ) load cell to measure force and torque . when both muscles were stimulated together , the resulting forces and torques ( all 6 dof ) were less than the sum of the individual forces . the peak error occurred during the onset of activation where force was about 9% less compared to the plateau where steady state force was about 2% less . there was no evidence that the direction of the forces changed during simultaneous activation of the muscles compared to activation of the muscles independently . these results suggest that when both muscle groups were activated together there was increased shortening of the muscle fibers , and hence less force due to a higher velocity of shortening during force onset . thus , there is some interaction , either between the muscle bellies or between their tendons . however , the interaction was small , and during steady state , it was almost immeasurable . they share a border and a tendon and thus may be expected to show nonlinear summation . the average peak error was 8.4% and the mean average error during the contraction was 1.3% ( unpublished observations ) . note that these experiments do not preclude epimuscular force transmission , but rather suggest that in normal muscle it has little functional effect on the overall force delivered to the skeleton . to tackle some of the concerns of previous studies , a new experimental approach was developed to measure directly the mechanical interactions between muscles in conditions that simulate those present during normal movements . the latter was assured by testing the muscles in a nearly intact limb of the cat . length changes were obtained by movements of the joints and , thus , only physiological relative movements could be imposed . the mechanical interactions between the one - joint so and its two - joint synergistic muscles were studied . the muscle bellies of lg and plantaris ( pl ) muscles share an interface with so . ankle moment exerted upon the isolated excitation of so was measured at various knee angles while the ankle was kept at a constant position ( ~90 ) , using a 6-degree - of - freedom load cell coupled to a 6-degree - of - freedom robotic manipulator ( figure 3 ) . note that knee movements will only alter the length and relative position of the two - joint muscles , but not of so . we hypothesized that force transmission from so muscle fibers will be affected by length changes of its synergists through configuration changes of connective tissues between these muscles . changing the knee angle from 70 to 140 lengthened lg and pl profoundly ( 4.57.2 mm ) , as calculated using the geometric model presented by goslow et al . . in contrast to our expectations , active ankle moment generated by so and the rate of muscle relaxation were not significantly affected by changes in knee angle . these results demonstrate that the presence of relative muscle movements does not necessarily mean force transmission between muscles . to further test the apparent independency of so , an additional set of experiments was performed . with minimal disruption of the connective tissues at the muscle belly level , the distal tendon of so was dissected free from the other tendons in the achilles tendon complex , cut , and connected to a force transducer . as this eliminated force transmission to its insertion on the calcaneus , any ankle joint moment following so excitation was attributed to force transmission via epimuscular myofascial pathways to the achilles tendon . if the tendon of so was placed at its original position , corresponding to the above reported ankle joint angle , the moment exerted at the ankle was close to zero while force exerted at the distal tendon of so was near its optimal value . a substantial ankle moment was found only if so was excited at positions distant from physiological . these results confirm that for in vivo muscle lengths and relative positions force generated in so muscle fibers is transmitted to its distal tendon . the above - described nearly linear force summation between mg and lg / so as well as between rectus femoris and vastus lateralis is in agreement with this finding . others have found that human so fascicle length was not affected by changes in knee angle , as measured in both passive and maximally active conditions of the ankle plantar flexors . in contrast , recent imaging studies in humans suggest that mechanical connections between gastrocnemius and so muscles are effective also within the in vivo length range . isolated excitation of mg at a fixed angle of the ankle and knee joints elicited a decrease in fascicle length , not only in the excited muscle , but also in so [ 32 , 33 ] . however , mg activation did not cause displacements in flexor hallucis longus muscle , suggesting that not all muscles are equally connected . in addition , effects of knee movements on so muscle have been reported [ 33 , 34 ] . note that the mechanical effects ( e.g. , ankle moment ) of such displacements in so were not measured . how can the different results between the rat and cat studies be explained ? we have hypothesized that the intermuscular linkages between so and adjacent muscles within the intact cat may be slack or operate on the toe region of their lumped stress - strain curve ( figure 4 , see also ) . the steep portion of this curve and , hence , epimuscular force transmission will then be attained only with supraphysiological displacements . the stiffness of the intermuscular linkages may also be a local property , being more compliant in the proximal region of the cat ankle extensors . note that maas and sandercock lengthened the two - joint muscles by knee movements ; thus , there was more movement proximally . preliminary data suggest indeed that cat so muscle is more rigidly connected to lg distally than proximally ( sandercock and maas , unpublished observations ) . also in line with this hypothesis are the results of an earlier study in the rat in which lengthening a muscle distally resulted in substantial force changes exerted at the distal tendon of a neighboring muscle , while effects of proximal lengthening were not significant . a major difference between the experiments on epimuscular myofascial force transmission in the rat ( see above ) and the cat is the number of muscles that is activated simultaneously . in the rat studies coactivation leads to several changes within the muscle compartment that may affect the mechanical interaction between adjacent muscles . the former will result in a small change of the muscle relative position , whereas the expansion will increase the lateral tension of the connective tissue network . in a recent study , we tested the hypothesis that the net effect of coactivation is an increase in the stiffness of the epimuscular pathways ( figure 4 ) , which will facilitate force transmission between muscles . effects of antagonist coactivation on mechanical interactions between synergistic muscles in the rat forelimb were assessed . in contrast to the hypothesis , changes in force of the restrained muscle with length changes of its synergist were unaffected by antagonist coactivation . testing intermuscular interaction with other combinations of active muscles ( e.g. , excluding the activity of some synergistic muscles ) may be necessary to elucidate the effects of muscle coactivation on the magnitude of epimuscular myofascial force transmission . finally , it is also conceivable that the mechanical characteristics of the connective tissue system are different between muscle groups within an animal and across species . the muscle - connective tissue architecture and composition of each synergistic group is different . therefore , generalizing the current results to the whole musculoskeletal system should be done with caution . although mechanical interactions between synergistic muscles have been shown in many animals ( e.g. , mouse , rat , cat , locust , frog ) , differences in connective tissue mechanical properties or differences in animal size may affect the importance of such force transmission . in contrast to mammals , it has been reported that amphibians have a relatively poorly developed connective tissue network and that insects contain very little connective tissue [ 38 , 39 ] . another aspect that should be taken into account is the scaling of muscle surface area ( to the 2nd power ) versus muscle volume ( to the 3rd power ) . this means that , for example , mice have a relatively larger epimysial surface to volume ratio than humans . to date , whether these variations between species lead also to different mechanical interactions remains unclear . the contradictory findings between the rat and cat studies are not fully understood and , thus , the responsible mechanisms requires further investigation . specifically , future studies should continue to test if the magnitude of intermuscular force transmission is dependent on the number of muscles that is simultaneously activated . the importance of epimuscular myofascial pathways for muscle function during normal movements remains unclear . besides a potential role for normal muscle function street observed , after the distal tendon of a frog 's semitendinosus muscle is cut , pathological changes appear first in the distal part . we found that when part of a muscle was normal the muscle as a whole generally developed the normal amount of tension and we guessed that some cell component other than the myofilament arrays served as a tension bypass through or around the damaged areas ( ramsey and street , unpublished ) . in the same paper , street suggested , in injured whole muscle it is probable that the connective tissue sheath near damaged fibers can pick up and transfer active tension generated in normal areas and , at the same time , stabilize abnormal areas against length changes . this suggests the hypothesis that the connective tissue network may act also as a safety net for traumatic events in muscle or tendon . in other words , due to the presence of myofascial pathways , the acute effects of muscle or tendon trauma are limited and muscle function is preserved . injury within a muscle or tendon is a common occurrence , especially during sport activities ( e.g. , [ 42 , 43 ] ) . thus , it is essential that basic function is maintained while the injury is healed . in the same study by maas and sandercock that showed little epimuscular force transmission in intact so , they also showed that during complete tendon transection the so can produce substantial force . the insertion of the so on the calcaneus was completely severed , yet the so produced an extensor moment at the ankle that was about 45% of normal . there was greater shortening , 17 mm compared to 1 mm , than before tendon transection . yet function was partially maintained possibly allowing use during recovery . the results of several previous studies support the idea that connective tissues within and surrounding muscles can limit injury and support repair . it has been reported that one of the four distal tendons of rat edl ( a multitendoned muscle , see ) can be cut or considerably shortened with minimal effects on force measured at the proximal tendon [ 45 , 46 ] . this can only be explained by transmission of force from the tenotomized muscle fibers to the intact distal tendons via the endomysial - perimysial network . similar phenomena have been reported following transection of the whole distal or proximal tendon . during some of the above - described rat experiments , following such a release of the proximal edl tendon , a substantial force was still found at the distal tendon . the muscle fibers of edl are thus prevented from shortening all the way , most likely by inter- and/or extramuscular connective tissues . in a different experiment , proximal ta + ehl force was not measured , but the changes in proximal and distal edl forces clearly indicate mechanical interactions between these synergistic muscles ( figure 5 ) . in addition to the acute backup from connective tissues following muscle and tendon trauma , long - term adaptations also suggest that myofascial pathways serve temporarily as a safety net . it has been reported that the integrin - vinculin mediated connections between the subsarcolemmal cytoskeleton and extracellular matrix are temporarily reinforced in ruptured muscle fibers [ 48 , 49 ] . as a consequence , force generated within the sarcomeres of these damaged fibers will be transmitted via the endomysium . this will reduce the load on the injured site allowing repair with less chance of rerupture . in the case that not the muscle fibers but the tendon is fully or partially torn , connective tissue linkages with adjacent structures may in a similar fashion prevent further trauma and facilitate the recovery process . such adhesions have been reported in chronically tenotomized muscle in the rat , cat , and rabbit [ 5052 ] , which ultimately results in reattachment of the tendon . nonsurgical treatment ( e.g. , immobilization at low length ) is also frequently applied following tendon ruptures in humans ( e.g. , ) . preserving and restoring function after injury clearly is important in wild animals and in contrast to humans , where the damage can be treated in a hospital , most animals must maintain some degree of function while the muscle - tendon injury heals . several surgical interventions include manipulation of tendon , muscle , and/or the surrounding connective tissues ( e.g. , fasciotomy in compartment syndrome , tendon transfer in cerebral palsy ) . in tendon transfer surgery limb function is improved in a patient by cutting and reattaching a tendon to a new insertion point ( e.g. , ) . preliminary results suggest that scar tissue formation following an agonist - to - antagonist tendon transfer in the rat significantly affects transmission of forces from the transferred muscle . therefore , knowledge of the acute and long - term effects of disrupting connective tissues has important implications for surgical practice . in the last decade , the potential of force transmission between skeletal muscles via inter- and extramuscular connective tissues has been demonstrated . more recent efforts have resulted in new insights regarding effects of epimuscular myofascial force transmission in more physiologically relevant muscle conditions ( e.g. , in vivo relative muscle movements ) . while not conclusive , these insights suggest that the role of this pathway may be small in normal undamaged muscles . future studies should investigate force transmission during muscle activation patterns that resemble those of normal movements . in particular , effects of decreasing the number of muscles that are active simultaneously on the mechanical connectivity between muscles need to be investigated . a full understanding requires knowing how the deformation of the border of a muscle affects the strain throughout the muscle . while the significance of epimuscular myofascial force transmission for muscle function in vivo remains unclear , potential functions for pathological muscle - tendon conditions ( e.g. , tendon rupture ) have emerged . Output:	the classic view of skeletal muscle is that force is generated within its muscle fibers and then directly transmitted in - series , usually via tendon , onto the skeleton . in contrast , recent results suggest that muscles are mechanically connected to surrounding structures and can not be considered as independent actuators . this article will review experiments on mechanical interactions between muscles mediated by such epimuscular myofascial force transmission in physiological and pathological muscle conditions . in a reduced preparation , involving supraphysiological muscle conditions , it is shown that connective tissues surrounding muscles are capable of transmitting substantial force . in more physiologically relevant conditions of intact muscles , however , it appears that the role of this myofascial pathway is small . in addition , it is hypothesized that connective tissues can serve as a safety net for traumatic events in muscle or tendon . future studies are needed to investigate the importance of intermuscular force transmission during movement in health and disease .
PubmedSumm2030	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: type 1 diabetes patients often suffer from total loss of their functional cells , as indicated by the complete absence of insulin c - peptide in their serum . replacing cell mass by pancreatic islet transplantation has been shown to be curative in principle but is limited by the availability of donor islets , immunological complications , and transplant survival ( shapiro et al . , 2006 ) . therefore , attempts to regenerate patient - specific insulin - producing cells have been undertaken using different cell sources , including embryonic stem cells ( escs ) , induced pluripotent stem cells ( ipscs ) , hepatic cells , exocrine cells , and other endocrine cells ( al - hasani et al . , 2009 , kroon et al . , 2008 , pagliuca et al . , 2014 , sangan et al . , 2015 , , 2012 , zhou et al . , 2008 ) . in most cases , these approaches have relied on the overexpression of master regulatory transcription factors involved in normal pancreas development , and in only a few cases , small molecules or biologicals have been used ( fomina - yadlin et al . , 2010 , kubicek et al . , 2012 , pennarossa et al . , 2013 , xie et al . , 2013 , yi et al . , 2013 ) . cells are particularly attractive starting points for transdifferentiation protocols , as they are developmentally closely related to cells . these cells have been shown to replenish insulin - producing cells following extreme cell loss ( thorel et al . , 2010 , ye et al . , 2015 ) . in a genetic model , overexpression of the transcription factor pax4 converts mouse cells to cells during development ( collombat et al . , 2009 ) and when triggered in adulthood ( al - hasani et al . , 2013 ) . molecularly , the cell factor pax4 acts by directly repressing the cell master regulatory transcription factor arx ( collombat et al . , 2003 ) , and the loss of arx alone is sufficient to convert cells into cells ( courtney et al . , 2013 ) . so far , pax4- and arx - mediated and cell transdifferentiation has been observed in vivo in the endogenous environment of a pancreatic islet of langerhans ( collombat et al . , 2009 , courtney et al . , 2013 ) . it remains unclear whether these cell - type conversions occur by a cell - autonomous mechanism or are dependent on other islet cell types and/or signaling from remote organs , such as the liver . to discriminate the two scenarios , we generated cell - line models for inducible overexpression of pancreatic transcription factors and showed their applicability to identify small - molecule modulators of the transdifferentiation process . from a representative library of approved drugs , we identified the anti - malarial drug class of artemisinins as compounds that functionally counteract the cell - specific transcription factor arx , thereby overcoming a major barrier for cell transdifferentiation . we show that these compounds bind to gephyrin , a multifunctional protein that has primarily been studied in neurons ( tyagarajan and fritschy , 2014 ) . among other functions , gephyrin is essential for active gabaa receptor signaling , a pathway that has been proposed to reverse diabetes by induction of cell proliferation ( purwana et al . , 2014 , soltani et al . , 2011 ) . consequently , cells acquire cell characteristics , resulting in improved glucose homeostasis in zebrafish and rodent diabetes models . to discriminate cell - autonomous effects of transcription factor - mediated transdifferentiation from phenotypes that require paracrine and endocrine signaling in an islet microenvironment , we engineered the mouse cell line min6 to allow the inducible overexpression of arx . based on the known heterogeneity of the parental cell line ( nakashima et al . , 2009 ) , we isolated clonal derivatives with integrations of constructs for the doxycycline - controlled transcriptional activation of myc - tagged arx ( figures s1a and s1b ) . while basal expression levels of and cell genes varied in the different subclones , we observed that induction of arx overexpression consistently activated the transcription of cell genes and repressed cell - specific transcripts ( figure s1c ) in a time - dependent manner ( figure s1d ) . these changes indicate that our cell lines faithfully model the -to- fate switch upon arx overexpression and suggest that transdifferentiation can be initiated in a cell - autonomous fashion . furthermore , the cellular system we generated is directly applicable to a high - content screen for functional repressors of arx . in dmso - treated control samples , we observed a > 50% reduction in insulin protein levels after arx induction compared to uninduced cells ( figures 1a and 1b ) . to identify small molecules that counter arx function , we induced arx expression and , at the same time , added compounds from a library of 280 clinically approved small molecules selected for their structural and target diversity . we then measured insulin levels after 72 hr and selected hit compounds for their ability to maintain high insulin levels in the presence of arx while not affecting cell viability ( figure 1c ) . all compounds that appeared to counteract arx in min6 cells were then tested for their ability to induce insulin expression in the cell line tc1 ( figure 1d ) . two artemisinins , artemether and its active metabolite dihydroartemisinin , fully inhibited the arx overexpression phenotype in min6 cells while also inducing insulin expression in tc1 cells , as predicted for functional inhibitors of arx ( figure 1e ) . importantly , artemether treatment increased insulin expression in min6-arx - induced cells and in tc1 cells in a dose - dependent manner ( figures s2a and s2b ) . to further understand the effects of artemether on cells , we analyzed the relative quantitative changes in the proteome of tc1 cells treated with artemether for 72 hr . proteins with roles in oxidative phosphorylation and mitochondrial localization were significantly altered , with some of these proteins showing increased abundance and others showing decreased abundance . in line with artemisinins acting as functional arx antagonists , glucagon protein levels were reduced by more than 50% after artemether treatment ( figure 1f ) . importantly , artemether reduced the abundance of both proglucagon and processed glucagon peptides in tc1 cells ( figure 1 g ) . to understand the effect of artemether on arx , we used the min6 overexpression model to study the cellular localization of the transcription factor . we observed that artemether induced the translocation of arx from the nucleus to the cytoplasm ( figures 1h1j ) , thereby depleting arx from chromatin . based on the activity of artemether , we investigated the effects of additional artemisinin analogs in cells . the closest structural analogs , including arteether and artesunate , showed effects comparable to those of artemether . in contrast , deoxyarteether , an analog lacking the endoperoxide moiety , did not increase insulin expression in cells ( figure s2c ) . to rule out the possibility that artemether , an endoperoxide , induced insulin expression due to the generation of reactive oxygen species ( ros ) , we applied the ros inducer elesclomol . no significant change of insulin expression in cells was apparent ( figure s2d ) . finally , combined treatment with the anti - oxidant n - acetylcysteine , a known suppressor of the anti - malarial properties of artemisinins ( arreesrisom et al . , 2007 ) , abolished the effect of artemether on insulin expression in cells ( figure s2e ) . artemisinins are among the most effective therapies for malaria and , based on their rapid toxicity to plasmodia , are the preferred first - line treatment for the disease ( nosten and white , 2007 ) . despite being widely used different molecular targets have been proposed , including heme binding followed by iron oxidation in human erythrocytes ( shukla et al . , 1995 , wang et al . , 2015 ) and the inhibition of two different plasmodium - specific proteins : the sarcoplasmic / endoplasmic reticulum calcium atpase ( serca ) pfatp6 ( eckstein - ludwig et al . , 2003 ) and phosphatidylinositol 3-kinase pfpi3k ( mbengue et al . , 2015 ) potent effects of artemisinins are also observed on human cells , where these compounds have been assigned anti - inflammatory and anti - cancer properties ( ho et al . , 2014 ) . there is limited evidence for possible positive effects of artemisia extracts on animal models of type 1 diabetes ( ahmad et al . , 2014 ) and in human patients ( al - waili , 1988 ) , but the use of isolated artemisinins for the treatment of diabetes has never been proposed , and their effect in the pancreas has not been evaluated . to identify the molecular mechanism of artemisinins in pancreatic cells , we first assessed whether the homologs of proposed targets of the compounds in plasmodia were involved . we used the serca inhibitor thapsigargin on tc1 cells and did not observe increased insulin levels up to dose - limiting toxic concentrations ( figure s3a ) . next , we tested whether artemether treatment of tc1 cells resulted in reduced pi3p and phosphorylated ( p-)akt levels , indicative of pi3k inhibition ( figures s3b and s3c ) . rather than inhibition , we observed an increased signal in both readouts , thereby excluding a conserved mechanism of action based on the proposed plasmodium targets . in order to discover potential mechanisms of action of artemisinins in mammalian endocrine cells , we undertook a chemical proteomics approach ( rix and superti - furga , 2009 ) to determine the proteins that directly interact with artemisinins in tc1 cells . we coupled artesunate , an artemisinin active in tc1 cells , to solid support and performed pull - down experiments in the presence and absence of competing free artemether ( figure 2a ) . mass spectrometry identified gephyrin as the most significantly enriched specific interactor ( figures 2b and s3d ) . artemether increased gephyrin protein levels in tc1 cells ( figures 2c and s3e ) , while mrna expression levels remained unchanged ( figure s3f ) , suggesting stabilization of the protein . to examine the interaction between gephyrin and artemisinins in a cellular context , we used a fluorescently labeled artelinic acid probe . the cellular localization of the compound was nearly identical with gephyrin in costaining experiments ( figure s3 g ) . next , we asked whether gephyrin was functionally required for the insulin increase in artemether - treated tc1 cells . using a specific small hairpin rna ( shrna ) targeting gephyrin , we achieved a > 70% reduction of gephyrin protein in tc1 cells ( figure s3h ) . in contrast to control - treated cells , cells treated with an shrna targeting gephyrin ( shgephyrin ) failed to increase intracellular insulin levels after artemether treatment ( figure 2d ) . single - cell analysis of the imaging data revealed a high correlation between the increases of gephyrin and insulin in artemether - treated cells , which was largely abolished when gephyrin was knocked down ( figure 2e ) . gephyrin exerts different functions , including enzymatic activity in the synthesis of the molybdenum cofactor moco , regulation of mtor signaling via direct interaction with mtor ( mechanistic target of rapamycin ) ( sabatini et al . , 1999 ) , and structural roles in the transport of glycine and gabaa receptors to the membrane ( fritschy et al . , 2008 ) . we observed increased moco synthesis in artemether - treated cell extracts , indicative of enhanced gephyrin enzymatic activity ( figure s4a ) . moreover , a small fraction of endogenous gephyrin interacted with mtor , and artemether inhibited this interaction ( figure s4b ) . finally , several subunits of the gabaa - receptor complex , which physically interacts with gephyrin , were increased on both the mrna and protein levels following artemether treatment ( figures 3a and 3b ; figure s3f ) . rna sequencing ( rna - seq ) experiments further underlined the effects of artemether on gabaa - receptor signaling in tc1 cells . gene ontology ( go ) term enrichment analysis identified synaptic transmission among the pathways significantly altered by artemether ( figure 3c ) , and we observed the upregulation of the genes p2rx3 , vamp1 , and nrxn3 involved in the gaba - signaling pathway ( figure s4c ) . therefore , we used 3-day pre - incubation with 10 m artemether to test whether adaptive cellular responses , such as gabaa - receptor - mediated effects , could contribute to changes in tc1 cell excitability to modulate hormone release . artemether - treated cells exhibited significantly lower basal ca levels ( figures 3d and 3e ) , including coincidently reduced nuclear ( figure s4d ) and cytoplasmic ( figure s4e ) ca relative to control cultures . likewise , partial substitution of extracellular na by k ( to trigger excitation ) induced differential responses in drug - exposed tc1 cells : physiologically relevant 15 mm kcl increased the fraction of cells with ca transients , suggesting increased excitability , as compared to untreated controls ( figure 3f ) . in 50 mm kcl , which induces signal saturation due to maximal ca influx through voltage - gated ( vg ) ca channels , artemether - treated cells showed attenuated slow ca responses in the second timescale ( figure 3 g ) , which was mimicked by pre - treatment of control cells with the gabaa - receptor antagonist picrotoxin . next , we tested whether acute application of muscimol , a gabaa - receptor agonist , could potentiate kcl ( 15 mm)-induced responsiveness and whether it exhibited an additive effect with artemether . indeed , muscimol - exposed cells showed further increases in their intracellular ca when simultaneously exposed to kcl . pre - incubation with the gabaa - receptor antagonist picrotoxin abolished these effects ( figure 3h ) , confirming gabaa - receptor involvement . based on these results , we suggest that artemether pre - treatment resets intracellular ca through gabaa - receptor - mediated mechanisms and acutely modulates tc1 cell excitability by potentiating gabaa - receptor signaling . moreover , the data show that gaba can act as an excitatory stimulus in tc1 cells . since gabaa receptors gate cl ions , we sought to directly monitor intracellular cl concentrations by using mqae dye imaging . 3-day pre - treatment with artemether induced an increase in intracellular cl ( figure 3i ) , which was not further changed when treated with muscimol acutely ( figure s4f ) . similarly , we detected muscimol - induced transmembrane currents at the low - picoampere range , if any , in both control and in artemether - treated tc1 cells acutely using perforated patch recordings ( data not shown ) . even though this might be seen as contradictory to a mechanism reliant on gabaa - receptor - gated currents , we argue that the high input resistance ( > 1 g ) of all recorded cells , together with the presence of vg ca currents operating at the picoampere range ( figure s4 g , pink shading ) , and with high - amplitude delayed - rectifier k channels ( figure s4 g ) , might be sufficient to induce membrane potential oscillations because of the initial activation of vg ca channels ( depolarization ) followed by the activation of vg k channels ( repolarization ) ( figure s4h ) . accordingly , the injection of minute currents ( up to 5 pa ) in current - clamp mode was sufficient to induce quasi - action potentials , which we expect to be responsible for the generation of ca transients regulating exocytosis . thus , artemether treatment can evoke global differences in tc1 cell excitability and hormone release through the potentiation of functional gabaa receptors . to prove that gabaa - receptor signaling plays a functional role in the mechanism of action of artemisinins in tc1 cells , we combined artemether with structurally and mechanistically distinct gabaa - receptor antagonists ( figures 3j and s4i ) . while these compounds had different effects on baseline insulin expression by themselves , all compounds partially inhibited the effects of artemether on insulin expression . we next tested whether treatment with different gabaa - receptor agonists alone was sufficient to induce insulin expression in tc1 cells ( figure 3k ) . most compounds particularly , the positive allosteric modulators etifoxine and ns11394induced insulin expression , but all gaba - receptor agonists were less potent relative to artemether . gaba is normally co - secreted with insulin by cells , and suppressing glucagon secretion is one of the physiological functions of gaba signaling in cells ( li et al . , 2013 , , 1989 , wendt et al . , 2004 , xu et al . , 2006 ) . in tc1 cells , artemether treatment inhibited glucagon secretion in low - glucose conditions ( figure 4a ) . interestingly , supplementing recombinant glucagon abolished the effect of artemether on insulin induction ( figure 4b ) . these data imply that maintaining the correct concentration of glucagon in close vicinity to cells is critical for the stability of this cell type . in order to lower the extracellular glucagon concentration , we applied decanoyl - rvkr - cmk ( pci ) , a highly potent , irreversible , cell - permeable inhibitor of proprotein convertases that inactivates prohormone convertase 2 at subnanomolar concentrations ( denault et al . , 1995 ) . treatment of tc1 cells with this compound resulted in a dramatic increase of intracellular glucagon staining , consistent with the accumulation of unprocessed proglucagon ( figure 4c ) . at the same time , these cells stained positive for insulin protein ( figure 4c ) . on the transcriptional level , the decrease of glucagon secretion repressed glucagon expression and significantly upregulated the expression of cell - specific transcription factors pax4 and mnx1 ( figure 4d ; table s2 ) . these results suggest that artemether - mediated amplification of gaba - receptor signaling prevents glucagon secretion , thereby reducing extracellular glucagon and triggering the loss of cell identity . to characterize the physiological relevance of our findings , we applied artemether in vivo in three model organisms . for developmental effects of the compound , we studied zebrafish carrying a gfp transgene under the glucagon promoter and an mcherry construct controlled by the insulin promoter . 5-day treatment of zebrafish larvae with artemether altered islet morphology and caused a dose - dependent reduction in cell numbers , with cell numbers affected at higher concentrations ( figures s5a and s5b ) . next , we applied the compound in a zebrafish cell ablation model at a later developmental stage . in animals expressing an insulin - promoter - driven caspase-8 transgene that can be activated pharmacologically , cells were completely ablated by the addition of a small - molecule dimerizer at 3 days post - fertilization ( dpf ) for 48 hr . the expression of a far - red fluorophore ( e2crimson ) under the control of the insulin promoter allowed the analysis of cell numbers at 12 dpf ( figures 5a and 5b ) . based on the known regenerative capacity of zebrafish , control ablated animals had an average of eight reestablished cells at this time point . in animals ablated and treated with artemether for 4 days ( 812 dpf ) , we observed a 75% increase in cell number to 14 cells per animal on average . next , we analyzed the effects of cell ablation and artemether on glucose levels in zebrafish . while cell ablation resulted in a 34% increase in glucose levels 7 days post - ablation , artemether treatment after ablation inversed that phenotype , and animals had 42% lower glucose levels compared to unablated controls ( figure 5c ) . in addition to these developmental models , we also sought to study artemisinin effects on mature islets in wild - type mice . in line with in vitro data , we observed an increase in pancreatic islet size following 3-month treatment with artesunate in drinking water ( figures 5d and 5e ) . these changes occurred in the absence of a dramatic shift in islet subpopulations or cell proliferation ( figure s5c ) , thereby phenocopying arx loss in vivo ( courtney et al . , 2013 ) . to confirm cells as the source of new insulin - positive cells , we turned to a lineage - tracing model . we isolated islets from mice harboring a construct for cre recombinase under the control of the glucagon promoter and a floxed red fluorescent protein ( rfp ) allele integrated in the rosa26 locus . thereby , all cells that have ever activated glucagon expression are permanently labeled rfp positive . in control dmso - treated islets , we observed that only a background of approximately 1% cells were double positive for insulin and rfp . treatment of these islets for 24 hr with artemether resulted in 4-fold upregulation of this cell population ( figures 5f and 5 g ) . these data indicate that artemether induces a rapid induction of insulin expression in cells within mouse islets , resulting in long - term expansion of the insulin - producing cell population and islet mass . finally , to analyze the effects of artemether treatment in a rodent diabetes model , we ablated cells in rats with a single high - dose streptozotocin injection . animals were randomized to treatment and control groups based on matching blood glucose measurements on day 9 post - ablation and then treated orally with artemether or control vehicle for 23 days . after overnight fasting , baseline blood glucose levels in artemether - treated animals were drastically reduced compared to those of control rats ( figure 5h ) . these lower baseline glucose levels were also reflected in an oral glucose tolerance test performed in the two groups ( figures 5i and 5j ) . we next assessed the effects of artemether on cells in primary human pancreatic islets ( figure 6a ; table s3 ) . the drug caused a rapid translocation of arx to the cytoplasm and a reduction in the staining intensity for this master regulatory transcription factor ( figure 6a ) . to better visualize the artemether - induced translocation of arx to the cytoplasm , we treated intact islets and then dissociated them to single cells immediately prior to immunofluorescence staining ( figure 6b ) . we observed that active gabaa - receptor signaling was a requirement for the translocation of arx to the cytoplasm following artemether treatment , since co - treatment with artemether and the gabaa - receptor antagonist picrotoxin maintained nuclear arx localization ( figure 6c ) . we then studied the effect of artemether on hormone levels and gene expression in primary human islets . compared to control dmso - treated islets , artemether increased the number of double - positive cells expressing both insulin and glucagon ( figures s6a s6c ) . the effects of artemether or gaba on global transcriptional profiles were tested in bulk samples containing 500 cells . gene set enrichment analysis ( gsea ) revealed significantly decreased expression of exocrine - specific genes ( li et al . , 2016 ) , both with artemether and gaba ( figure s7a ; table s1 ) . in contrast , genes upregulated by both artemether and gaba significantly overlapped with an endocrine , but not an exocrine , gene set ( figure s7b ) . to investigate the cell - type - specific gene expression changes with artemether treatment , we performed single - cell transcriptomics on human primary islets . we treated intact islets with artemether or dmso , dissociated the islets immediately prior to sorting single cells to individual wells of a 96-well plate containing lysis buffer , and performed single - cell rna - seq as previously described ( li et al . , 2016 ) . we then assessed global transcriptomic alikeness between individual cells through multidimensional scaling ( figure 6d ) . we observed that compound - treated cells clustered with dmso - treated controls , indicating that up to 72 hr of in vitro treatment with artemether did not induce a whole new cell type . looking for changes in specific genes related to the compounds mechanism , we found that gabrb3 and gabrg2 were strongly upregulated in artemether - treated cells ( figure 6e ) , consistent with the findings in the mouse cell line . an increase in gephyrin and gaba - receptor subunit protein levels on some islet cells was also evident by immunofluorescence staining ( figure 6f ) . to analyze transcriptome - wide changes , we used gsea to identify differentially regulated pathways in dmso- versus artemether - treated single - cell transcriptomes . in cells , artemether downregulated some cell - type - specific transcripts ( figures s7c and s7d ) . in cells , we observed a strong and significant downregulation of cell - specific genes , including eif4a1 , cryba2 , pdk4 , and muc13 following artemether treatment ( figures 6 g and 6h ) . key cell - specific genes were upregulated in cells following artemether treatment , including gnas and abcc8 , the gene coding for the sulfonylurea receptor ( figures s7e and s7f ) . we next assessed whether the reduction in cell character and gain of cell characteristics following artemether treatment had functional consequences in intact human islets . first , we tested the expression of cell factor arx ( figure 7a ) . while we observed donor - to - donor variability of the effect size , arx expression was consistently reduced . we then investigated whether the loss in cell character corresponded to increased cell properties . normalized to total protein , artemether - treated islets contained more intracellular insulin compared to dmso - treated islets from the same donor ( figure 7b ) . finally , we tested whether this insulin increase was functional and performed glucose stimulated insulin secretion assays . while artemether treatment did not affect secretion in low - glucose conditions , artemether - treated islets from all donors tested secreted significantly more insulin compared to dmso - control - treated islets when challenged with high - glucose concentrations ( figure 7c ) . we identified artemisinins as approved drugs that can confer cell characteristics to cells . this phenotype depends on the stabilization of the protein gephyrin , which , in turn , activates gaba signaling in cells . active gaba signaling is required for the effects of artemisinins on the key cell - fate determining transcription factor arx , which translocates to the cytoplasm following drug treatment ( figure 7d ) . it is tempting to speculate that the changes in nuclear calcium ( and/or chloride ) signaling that we observed provide a direct link between extracellular glucagon , gaba - receptor signaling , and nuclear arx localization . previous studies have already connected cells key function of glucagon secretion to gaba - receptor signaling . gaba is primarily produced by cells , whereas several gabaa - receptor subunits are specifically expressed on cells ( franklin and wollheim , 2004 , wan et al . , 2015 ) . the architecture of the vasculature in islets ensures that arterioles first contact insulin - expressing cells ( bonner - weir and orci , 1982 , stagner and samols , 1992 ) , resulting in the exposure of the downstream cells to high local concentrations of factors secreted by cells , including insulin and gaba . insulin is known as a major repressor of glucagon secretion ( maruyama et al . , 1984 ) , and the action of the hormone is dependent on active akt and gaba signaling ( xu et al . , our findings suggest that gaba signaling not only affects cell function , as measured by glucagon secretion , but also feeds back to cell identity . thus , we propose an additional beneficial effect of activating gaba - receptor signaling in diabetes , beyond the modulation of the immune system and cell proliferation previously suggested ( purwana et al . , 2014 , soltani et al . , 2011 ) . a key remaining question concerns the lack of to cell transdifferentiation in diabetes . why would evolution equip cells with a high amount of epigenetic plasticity , so that loss of a single transcription factor is sufficient for their transdifferentiation , and why would a mechanism to exclude this very transcription factor from the nucleus be maintained ? however , the only conditions known , when cells convert into cells , are following the genetic manipulation of these master regulatory transcription factors ( collombat et al . , 2009 , courtney et al . , 2013 ) and a toxin - induced model of near - complete cell loss ( thorel et al . , 2011 ) . particularly in type-2-diabetic conditions , several of the factors observed to contribute to arx repression are altered in similar ways to the changes we observe with artemisinins . for example , gaba receptors accumulate on the surface of cells in high - glucose / high - insulin conditions ( xu et al . , 2006 ) . moreover , long - term treatment with gaba alone is sufficient to induce cell - mediated , -like cell neogenesis in a mouse model ( ben - othman et al . , 2016 ; however , we still only have a limited understanding of the intra - islet changes associated with diabetic conditions , including local concentrations of gaba , insulin , and glucagon , as well as the composition of gaba receptors and other factors affecting the autocrine and paracrine effects of these molecules . insufficient cell - derived gaba levels and inappropriate glucagon secretion in diabetic high - glucose conditions could , therefore , prevent endogenous cell transdifferentiation in diabetic conditions . in our cell - line model , likely , the strong transcriptional induction of several gaba - receptor subunits following drug treatment induces a specific gaba - receptor subunit composition not normally present in cells . these receptors then increase gaba responsiveness and alter baseline calcium signaling of artemether - treated cells , and the effects could potentially synergize with other aspects of gephyrin biology and pi3k signaling . artemisinin combination therapy is the treatment of choice for malaria and more than 300 million doses are dispensed annually ( dondorp et al . , 2010 ) . despite this large patient cohort , no clinical data on the effects of artemisinins on human pancreatic endocrine function are available . first , the acute life - threatening condition of plasmodium - infected patients , together with the known propensity of plasmodium infection to cause hypoglycemia , makes blood glucose levels highly variable in the short term . second , in healthy individuals , a small increase in cell number is not expected to cause a phenotype , as these cells secrete insulin in a highly glucose - regulated manner . unfortunately , currently no imaging methods are available to directly assess human cell mass . we observe rapid effects of these compounds on cell lines , isolated islets , and zebrafish larvae in the timescale of hours to days . in contrast , effects in rodent models take weeks to months , potentially because of the lower bioavailability of artemisinins in vivo . plasma half - lives of these compounds are in the range of minutes to a few hours , and the poor solubility in aqueous solutions precludes the continuous dosing using pumps ; hence , we rely on once - daily oral dosing in rats . in contrast , for malaria patients , a typical treatment cycle with artemisinins lasts only 4 days , but dosing occurs at relatively high concentrations every 8 hr . currently , clinical studies are ongoing for the long - term effects of artemisinins in cancer patients ( ericsson et al . , 2014 ) ( nct02263950 ) . therefore , investigation of the effects of long - term artemether treatment for the induction of endogenous insulin expression in c - peptide - negative type 1 diabetes patients appears clinically feasible . regardless of the direct clinical use of artemether in diabetes , our findings open up completely new avenues for transdifferentiating cells by structurally different gephyrin stabilizers , gaba - receptor transcriptional activators , and additional targets in the gaba - receptor signaling pathway , toward a treatment for type 1 diabetes . reagent or resourcesourceidentifierantibodiesinsulinsigmacat#18510dakocat#a056401 ; rrid : ab_2617169linkocat#4011 - 01glucagonabcamcat#ab92517 ; rrid : ab_10561971arxr&dcat#af7068 ; rrid : ab_10973178myc - tagcell signaling technologycat#2276histone h2bcell signaling technologycat#2934gephyrinabcamcat#ab25784 ; rrid : ab_1209349gephyrin ( figure s4b)synaptic systemscat#147111 ; rrid : ab_887719gaba - receptor subunit beta 3/4milliporecat#mab341 ; rrid : ab_2109419gaba - receptor subunit alpha 2abcamcat#ab176170gaba - receptor subunit gamma 2abcamcat#ab101036 ; rrid : ab_10672116mtorcell signaling technologycat#2972p - mtorcell signaling technologycat#5536aktcell signaling technologycat#4691p - aktcell signaling technologycat#4060p ; rrid : ab_972551actinsigmacat#a2066 ; rrid : ab_476693alpha - tubulinabcamcat#ab7291 ; rrid : ab_2241126dsredclontechcat#632496 ; rrid : ab_10013483goat anti - rabbit 633thermo fischercat#a-21070 ; rrid : ab_2535731goat anti - guinea pig 546thermo fischercat#a-11074 ; rrid : ab_2534118goat anti - guinea pig 488thermo fischercat#a-11073 ; rrid : ab_2534117donkey anti - rabbit 488thermo fischercat#a-21206 ; rrid : ab_2535792donkey anti - mouse 488thermo fischercat#a-21202 ; rrid : ab_2535788donkey anti - sheep 546thermo fischercat#a-21098 ; rrid : ab_2535752donkey anti - rabbit 546thermo fischercat#a-10040/a-10036/a-10074 ; rrid : ab_2534016rabbit anti - sheep igg - hrpsanta cruzcat#sc-2770 ; rrid : ab_656968donkey anti - rabbit hrpjackson immunoresearchcat#711 - 035 - 152 ; rrid : ab_10015282donkey anti - mouse hrpjackson immunoresearchcat#715 - 035 - 151 ; rrid : ab_2340771donkey anti - sheep hrpjackson immunoresearchcat#713 - 035 - 147 ; rrid : ab_2340771donkey anti - goat hrpjackson immunoresearchcat#705 - 035 - 147 ; rrid : ab_2313587chemicals , peptides , and recombinant proteinsartemethersigmacat#a9361artemether ( figures 5h5j)cayman europecat#11815bicucullinetocriscat#0130gabazinesigmacat#s106picrotoxinsigmacat#p1675salicylidene salicylhydrazidesigmacat#s8194etifoxinesigmacat#sml0272muscimolsigmacat#m1523vgabatrinsigmacat#v8261halothanemedchemexpresscat#hy - b1010ns11394medchemexpresscat#hy-11048zolpidemsigmacat#e006thapsigarginsigmacat#t9033gaboxadolsanta cruzcat#sc204342furosemidetocriscat#3109fg-7142sigmacat#e006elesclomolselleckcat#s1052nacetylcysteinesigmacat#a9165lynxtag_artbiolynx technologiescat#bl007ruopcienzo life sciencescat#alx-260 - 022-m001artesunatesigmacat#a3731artesunate ( figures 5d and e)tcicat#a2191deoxyarteethersanta cruzcat#sc-211235arteethersanta cruzcat#sc-217662n - hydroxy-11-azaartemisinintrccat#h804500artemisitenetrccat#a777550artemisonealsachimcat#c1740artemisitenalsachimcat#s1873dihydro artemisinintrccat#a637875recombinant glucagonsigmacat#g2044dimerizer ap20187clontechcat#635059pe2-crimson - n1 vectorclontechcat#632554methylcellulosesigmacat#m6385hoechst 33342thermo fischercat#h1399critical commercial assaysxanthine oxidase assay kitabcamcat#ab102522pi(3)p mass elisa kitecheloncat#k-3300glucagon elisaalpcocat#48-gluhu - e01-alinsulin elisaalpcocat#80-inshu - e01.1glucose assay kitbiovisioncat#k606 - 100deposited datasingle cell rna - seq of the control cellsncbi gene expression omnibusgeo : gse73727single cell rna - seq of the artemether treated cellsncbi gene expression omnibusgeo : gse84714rna - seq of bulk samplesncbi gene expression omnibusgeo : gse84592experimental models : cell linestc1novo nordiskatcc cat#crl-2934tc1-ltthis papern / amin6novo nordiskmiyazaki et al . , 1990min6-arxthis papern / aexperimental models : organisms / strains(gcga : gfp)- ( ins : ntr - mcherry ) zebrafishthis papern / ains : caspase8;ins : e2crimson zebrafishthis papern.s . and d.m . , unpublished dataglucre::rfp mousethis papern / arecombinant dnatetr - cmv - arx - myc - flagthis papern / asv40 large t - antigen lentivirusgentargetcat#lvp016-neosequence - based reagentsprimers for ins2 , actb , pax4 , actbfomina - yadlin et al . , 2010n / aprimers for mnx1 , gcg , arxthis papertable s2shgephyrinsigmacat#trcn0000098585shcontrolsigmacat#shc001software and algorithmstophat v2.0.4kim et al . , 2013https://ccb.jhu.edu / software / tophat / index.shtmlrseqc packagewang et al . , 2012http://rseqc.sourceforge.net / gene set enrichment analysissubramanian et al . , 2005http://software.broadinstitute.org / gsea / downloads.jspmootha et al . 2012https://fiji.sc / cell counter pluginkurt de vos , university of sheffield , academic neurologyhttps://imagej.nih.gov / ij / plugins / cell - counter.html further information and requests for reagents may be directed to , and will be fulfilled by the lead contact stefan kubicek ( [email protected] ) . tc1 cells were purchased from atcc ( crl-2934 ) and grown in low - glucose dmem supplemented with 10% fbs , 50 u / ml penicillin and 50 g / ml streptomycin . min6 cells with doxycycline - inducible constructs were grown in high - glucose dmem supplemented with 15% tet system approved fbs ( clonetech 631106 ) , 71 m 2-mercaptoethanol , 50 u / ml penicillin and 50 g / ml streptomycin . the mouse islets from the cell lineage - tracing animals were kept in rpmi medium supplemented with 10% fbs , 50 u / ml penicillin and 50 g / ml streptomycin . the cell culture for human islets followed established protocols ( walpita et al . , 2012 ) . all mouse work was conducted in patrick collombat s lab as previously described ( courtney et al . , 2013 ) according to french ethical regulations under the animal license nce/2012 - 85 and nce/2014 - 190 . rat experiments were conducted at physiogenex according to the standard operating procedure in place at the test facility and an agreement from the french veterinary services . all procedures were performed in accordance with the guide for the care and use of laboratory animals ( revised 1996 and 2011 , 2010/63/eu ) and french laws . transgenic ( gcga : gfp)-(ins : ntr - mcherry ) zebrafish were maintained according to the guidelines of the local authorities under licenses gz:565304/2014/6 and gz:534619/2014/4 . transgenic ins : caspase8;ins : e2crimson ( abbreviated ins : casp8 ) zebrafish embryos were generated under license bmwf-66.008/0017-ii/3b/2013 . human islets were obtained from the european consortium for islet transplantation ( geneva ) and the integrated islet distribution program iidp . this study is approved by the ethics committee of the medical university of vienna ( ek - nr . detailed donor information is provided in table s3 . to generate inducible min6 cells , we first transduced cells with a lentivirus carrying tetr dna and a neomycin resistance cassette ( gentarget , cat . # lvp017-neo ) and then isolated a single clone expressing high amounts of tetr protein , validated with the tet01 antibody ( mobitec ) . this tetr expressing cell line was then transduced with virus particles coding for flag - myc tagged ( c - terminal ) human arx cdna also expressing the puromycin resistance gene . the vector used for this work was pstk007 , a modified version of the plenti4.2 ( life technologies ) . then , a second round of clonal selection was performed where numerous clones were tested for maximum repression in the uninduced state and a high degree of induction upon treatment with doxycyline ( 1g / ml ) . all lentiviruses were produced in hek293 t cells using the virapower plasmid mix ( life technologies ) , according to the manufacturer s protocol . all virus work was done in a class ii laboratory , and before transferring the cells to a regular lab the cells were proven to be virus free using a p24 elisa kit ( cell biolabs , cat.#vpk-108-h ) . to establish the tc1 subclone used for all experiments described in this study , tc1 cells were infected with a lentiviral vector causing the expression of large t antigen under the cmv promoter to make them independent of the glucagon - promoter driven large t antigen expression . 50 nl compound per well was transferred to black optical suitable 384-well plate ( corning 3712 ) from dmso stock plates using acoustic transfer ( labcyte echo 520 ) . 3000 min6 cells per well were then plated in 50 l media containing 1 g / ml doxycycline on top of the compounds . three days after seeding , cells were fixed in 3.7% formaldehyde for ten minutes at room temperature . following washing with pbs , cells were fixed with cold pure methanol at 20c for 10 min , permeabilized with 1% triton x-100 in pbs for 30 min and blocked with 3% bsa in pbs for 30 min . 20 l primary anti - insulin antibody ( 1:2000 ) in 1.5% bsa / pbs , was added per well and incubated at 4c overnight . after washing with pbs twice , 20 l cy-3labeled donkey--guinea pig antibody ( 1:1000 ) and 10 g / ml hoechst 3342 in pbs was added per well and incubated for 1 hr . after two washes with pbs , plates were stored in 4c in dark until analysis . images were taken by an automated microscope ( perkin elmer operetta ) using a 20x objective . images were exposed for 10 ms in the hoechst channel and for 500 ms in the alexa fluor 546 channel . nuclei were identified ( harmony method c ) and cytoplasm was defined based on the nuclei ( harmony method c ) . in total 1152 wells were screened containing 280 compounds from the cemm library of unique drugs ( cloud ) ( m. licciardello , p. markt , f. klepsch , and s.k . , unpublished data ) in triplicates with control wells . hits were selected based on the intensity of insulin in the alexa fluor 546 channel and cell numbers in the hoechst channel . immunofluorescence staining for all other antibodies was performed as described above , images were acquired on an automated microscope ( perkin elmer operetta ) or a leica dmi6000 b inverted confocal system with a 63x 1.30 acs apo lens . image analysis and quantitation was performed as described above . for quantification , at least 1,000 cells were analyzed for each condition . cells were lysed and rna isolated using the rneasy mini kit ( qiagen ) according to the manufacturer s protocol . the amount of total rna was quantified using a qubit 2.0 fluorometric quantitation system ( life technologies ) and the rna integrity number ( rin ) was determined using an experion automated electrophoresis system ( bio - rad ) . the libraries for rna - seq after 24h induction in min6 cells were prepared with ribo - zero and scriptseq v2 kits ( epicenter ) following the provider s manual . other rna - seq libraries were prepared with truseq stranded mrna lt sample preparation kit ( illumina ) using sciclone and zephyr liquid handling robotics ( perkinelmer ) . library amount was quantified using the qubit 2.0 fluorometric quantitation system ( life technologies ) and the size distribution was assessed using experion automated electrophoresis system ( bio - rad ) . libraries were pooled and sequenced on the illumina hiseq platform using the 50 bp single - read configuration . reads were aligned with tophat ( v2.0.4 ) with the no - novel - juncs no - novel - indels options ( kim et al . , 2013 ) . gene expression was calculated as reads per kb per millions of reads ( rpkms ) using rpkm_count.py from the rseqc package ( wang et al . , 2012 ) and the ncbi rna reference sequences collection ( refseq ) downloaded from ucsc . sample preparation and data analysis for single cell rna - seq was conducted as previously described ( li et al . , 2016 ) . after the rna was isolated with the rneasy mini kit ( qiagen ) , it was reverse transcribed with random primers using the high capacity cdna reverse transcription kit ( applied biosystems ) . quantitative pcr was performed with power sybr green pcr master mix ( applied biosystems ) on a lightcycler 480 ( roche ) . ratiometric imaging was performed using a visichrome monochromator ( visitron systems ) on a coolsnap hq2 camera ( photometrics ) and recorded in visiview software ( visitron systems ) . krebs - ringer solution was used as basic external solution and contained : 119 mm nacl , 2.5 mm kcl , 1 mm nah2po4 , 1.5 mm cacl2 , 1.5 mm mgcl2 , 20 mm hepes , 11 mm glucose ( ph 7.4 ) . a p value of < 0.05 was considered statistically significant , and calculated by the mann - whitney rank sum test ( unpaired test ) or wilcoxon s signed rank test ( paired test ) , as appropriate . tc1 cells were assayed by patch - clamp electrophysiology using the gramicidin or amphotericin perforated - patch configuration . ionic currents and membrane potential were recorded , filtered , and analyzed using an axopatch 200 b amplifier , digidata1322 interface , and the pclamp8 software ( molecular devices ) . the external solution contained : 119 mm nacl , 2.5 mm kcl , 1 mm nah2po4 , 1.5 mm cacl2 , 1.5 mm mgso4 , 11 mm glucose and 20 mm hepes - naoh ( ph 7.4 ) . for the measurement of muscimol - induced currents the following intracellular solution was used : 140 mm cscl , 4 mm nacl , 2 mm mgcl2 , 10 mm hepes , 1 mm egta , 4 mm atp - na2 , 400 mg / l amphotericin , ( ph adjusted to 7.3 with naoh ) . for studying excitable properties of tc1 cells the intracellular solution contained : 114 mm k - gluconate , 6 mm kcl , 10 mm hepes , 5 mm egta , 4 mm atp - mg , 30 - 50 mg / l gramicidin ( ph adjusted to 7.3 with koh ) . muscimol induced cl changes , tc1 cells were preincubated with mqae for 0.5 hr ( thermo fisher scientific ) . the fluorescence was recorded with coolsnap hq2 camera ( photometrics ) and visiview software ( visitron systems ) using 350 nm excitation with visichrome monochromator ( visitron systems ) and the emission filter ( 445 50 nm ) . basal levels were measured after 1 hr incubation with mqae and washing with pbs by live cell imaging on an automated microscope ( perkin elmer operetta ) using a 20x objective and analyzed using the harmony software ( perkin elmer ) . the measurement of moco synthase was performed with xanthine oxidase assay kit ( abcam ab102522 ) . whole cell extracts were generated by lysing cells in np-40 buffer containing 150 mm sodium chloride , 1.0% np-40 and 50 mm tris , ph 8.0 supplemented by protease inhibitor cocktail ( roche ) . 30 g whole cell lysates were loaded onto a sds polyacrylamide gel for electrophoresis at 30 ma per gel , and then transferred by electrophoresis to a nitrocellulose membrane ( ge healthcare life science ) . all the blots were incubated with corresponding primary antibodies diluted 1:1000 in 5% milk at 4c overnight followed by hrp - labeled secondary antibodies ( 1:20000 ) for 1 hr at room temperature . primary antibody was incubated with dynabeads protein g ( life technologies 10765583 ) for 15 min at room temperature and washed with pbs containing 0.02% tween-20 ( pbst ) . antibody - coupled beads were then incubated with tc1-lt cell lysates overnight at 4c , washed with pbst for three times and eluted with sds - loading buffer . pi3p measurements were performed using the pi(3)p mass elisa kit ( k-3300 , echelon ) . in brief , tc1-lt cells were pretreated with artemether for three days , followed by lipid extraction and an elisa assay according to the manufacturer s guideline . cells were first incubated in high glucose medium ( 5 g / l glucose in krbs medium ) for one hour followed by incubation in low glucose medium ( 0.5 g / l glucose in krbs medium ) for another hour . supernatant after low glucose challenge was collected to measure glucagon content using a glucagon elisa ( alpco 48-gluhu - e01-al ) . transgenic zebrafish strains tg(gcga : gfp ) and tg(ins : ntr - mcherry ) were used to demarcate cells by gfp expression and cells by mcherry expression . double transgenic zebrafish were treated with indicated concentrations of artemether added to e3 medium from 26 hpf until 100 hpf . zebrafish were embedded for confocal microscopy as described previously ( distel and koster , 2007 ) . z stacks of the pancreatic islets were taken on a confocal microscope ( leica tcs sp8 x wll ) using a 25x water objective . using the measurement counting tool of the leica software on 3d models of the islets gfp - positive cells and mcherry - positive cells transgenic lines were generated using tol2 mediated transgenesis ( kawakami et al . , 2004 ) . 1 kb of the insulin promoter ( huang et al . , 2001 ) was cloned into the p5e - mcs plasmid of the tol2 gateway system ( kwan et al . , 2007 ) using the bamhi and hindiii restriction sites . 2005 ) ( a kind gift of philipp scherer ) was cloned downstream of the ins promoter in the p5e - ins plasmid . e2crimson ( pe2-crimson - n1 vector , clontech ) was cloned into the pme - mcs plasmid of the tol2 system and expressed via a separate insulin promoter inserted via the bamhi and hindiii sites . final constructs were generated by an lr recombination reaction as described ( kwan et al . , 2007 ) in pdesttol2pa , and injected together with transposase mrna into fertilized eggs to generate tg(ins : caspase8;ins : e2crimson ) , abbreviated ins : casp8 , fish . zebrafish ( danio rerio ) were maintained according to standard protocols . transgenic lines were generated and maintained in the mitfab692/b692/ednrb1b140/b140 background ( a kind gift of wolfgang driever at the university of freiburg ) . 3 days post fertilization ( dpf ) old embryos were incubated in 2 m dimerizer ap20187 ( dim ) for 48 hr at 28c in the dark . dim was added to eggwater ( 0.3 g / l coral pro salt ( red sea ) to reach the final concentration . control embryos were exposed to corresponding amounts of ethanol . at 5 dpf ( 0 days post ablation , dpa ) , larvae were rinsed in egg water and transferred to fresh egg water at a density of 20 fish/200 ml . starting at 3dpa , ablated larvae were treated with either 5 m artemether or an equivalent volume of dmso for 4 d. compound was renewed every second day . larvae were harvested at 7 dpa and fixed for 1 to 2 hr at room temperature in 4% pfa in pbs , then washed 3x 5 min with 1 x pbs/0.2% triton x-100 . to improve access of antibodies to internal structures , head and tail were cut off and the gut was cut open . larvae were incubated in blocking buffer containing 1% dmso , 1% sheep serum , 1% bsa and 1% triton x-100 in 1 x pbs for at least 60 min at room temperature . the embryos were then incubated overnight at 4c with primary antibody recognizing e2crimson ( rabbit anti - dsred , 1:100 dilution ) , washed and then reblocked and incubated in secondary antibody ( anti - rabbit alexa fluor 633 conjugated , 1:1000 dilution ) overnight at 4c . confocal images were acquired using a zeiss lsm exciter5 microscope with a 40x water objective . for glucose assays , after removal of all liquid , larvae were resuspended in 200 l ice - cold , autoclaved 1x pbs , then sonicated on ice with 6 pulses of 0.5 s , and centrifuged at 13,000 rpm for 15 min at 4c . the glucose assay was performed as described ( kimmel et al . , 2015 ) on biological replicates , using 15 l larvae sample buffer was added up to 25 l , and combined with 25 l reaction mix . after incubation at 37c for 30 min , ex / em 535/590 was read on a microplate reader . protein concentration ( a280 ) was measured on a nanodrop and used to normalize glucose concentration to protein content . insulin - expressing ( e2crimson+/alexa 633 + ) cells were counted in z stacks using the cell counter plugin of imagej . confocal images were processed by a median filter in imagej to remove speckle noise and assembled using adobe illustrator . / l glucose in krbs medium ) for one hour and in high glucose medium ( 5 g / l glucose in krbs medium ) for another hour . supernatant was collected after low and high glucose incubations to measure insulin content using an elisa kit ( alpco 80-inshu - e01.1 ) . the islet pellets after high glucose incubation were lysed in np-40 buffer and intra - islet insulin content was measured using the same kit . two months old 129/sv mice were treated with artesunate ( 1mg / ml ) in drinking water provided ad libitum for 3 months . animals were sacrificed by cervical dislocation and the tissues were fixed and sectioned as previously described ( collombat et al . , 2003 ) . for the ex vivo lineage tracing experiments , islets from 2 months old glucre::rfp animals ( containing the following transgenes : the rat glucagon promoter upstream of the phage p1 cre recombinase ( herrera , 2000 ) and the rosa26 promoter upstream of a loxp - stop - loxp - tdrfp cassette ( luche et al . , 2007 ) ) were isolated by collagenase type xi ( 1 mg / ml ) digestion followed by a density gradient to purify the islets from the acinar tissue . approximately 250 islets per group were handpicked under the microscope and treated with either 10 m artemether or dmso as control . following 24 h of treatment , the islets were dissociated , fixed , and plated into a 384-well plate . following washing with pbst and blocking with bsa , the cells were incubated with guinea pig polyclonal anti - insulin ( 1/500-linco ) for 2 h at room temperature . the cells were then washed and incubated with 488-alexa anti - guinea pig ( 1/1000-molecular probes ) together with the nuclear counter - stain dapi for 1 h. the cells were then imaged on an automated microscope ( perkin elmer operetta ) using a 20x objective and analyzed using harmony software ( perkin elmer ) . rat experiments were conducted on 2 months old male sd rats which were fasted overnight and treated with 60 mg / kg streptozotocin i.p . on day 9 post ablation animals were randomized to treatment and control groups based on matching fasting glucose levels , and treated with 20 mg / kg artemether p.o . for 7 16 d. 1% methylcellulose was used as vehicle , prepared once per week by dissolving in distilled water , heating to 40c , then stored at 4c . blood glucose were measured via tail nick using a handheld glucometer ( accu - check performa roche diagnostics ) . nmr spectra were recorded on a bruker avance iii 400 ( bruker , billerica , ma , u.s ) . mass spectra were recorded using a xevo - uplctq - ms system ( waters , milford , ma , u.s . ) . purification by flash column chromatography ( fcc ) was performed using silica gel 60 ( merck , darmstadt , germany ) , mplc was performed on a biotage isolera system ( biotage , uppsala , sweden ) . drug - affinity matrices were prepared essentially as described previously ( fernbach et al . , 2009 , 2.7 l , 40 mol ) , ethanolamine ( 9.7 l , 160 mol ) , and trimethylamine ( 15 l , 108 mol ) were added to 500 l nhs - activated sepharose 4 fast flow beads ( ge healthcare bio - sciences ab , uppsala , sweden ) and the reaction was incubated on a rotary shaker for 24 hr . beads were washed and resuspended in dmso and nhs - activated artesunate ( 100 l , 1.00 mol ) was added to the suspension , and the mixture was incubated on a rotary shaker for 24 hr . unreacted beads were blocked by addition of nhs - acetate ( 10 mol ) and triethylamine ( 25 l , 180 mol ) , followed by agitation on a rotary shaker for 24 hr . after washing with dmso and lysis buffer , beads were incubated with cell lysates . affinity chromatography and elution were performed in duplicate as reported previously ( fernbach et al . , 2009 , after elution , enriched proteins were reduced with dithiothreitol , cysteine residues alkylated by incubation with iodoacetamide and the samples digested with modified porcine trypsin ( promega , madison , wi ) . three percent ( and multiples thereof ) of the digested eluates were purified and concentrated by c18 reversed - phase material . samples were analyzed at ph 2.4 on an agilent 1200 nano - hplc system ( agilent biotechnologies , palo alto , ca ) coupled to a hybrid linear trap quadrupole ( ltq ) orbitrap velos mass spectrometer ( thermofisher scientific , waltham , ma ) utilizing the xcalibur software version 2.1 . the 15 most intense ions were isolated and fragmented by collision - induced dissociation ( cid ) . dynamic exclusion for selected ions was 60 s and a single lock mass at m / z 445.120024 ( si(ch3)2o)6 ) was used for internal mass calibration ( olsen et al . , 2005 ) with the target loss mass abundance of 0% . maximal ion accumulation time allowed was 500 ms and overfilling of the c - trap was prevented by automatic gain control set to 10 ions for a full ftms scan and 5 10 ions for msn . intact peptides were detected in the orbitrap mass analyzer at a resolution of 60,000 with a signal threshold of 2,000 counts for triggering an ms event . further details of the lcms methodology are as previously described ( maurer et al . , 2013 ) . peak extraction and conversion of raw ms files into the mgf format was performed with msconvert ( proteowizard library v2.1.2708 ) and the peak lists searched against the mouse swissprot database v2013.01_20130110 ( 24,615 sequences including isoforms obtained from varsplic.pl ( kersey et al . , 2000 ) and appended known contaminants ) with the search engines mascot ( v2.2.03 , matrixscience , london , uk ) and phenyx ( v2.5.14 , genebio , geneva , switzerland ) ( colinge et al . , 2003 ) . submission to the search engines was via a perl script that performs an initial search with relatively broad mass tolerances ( mascot only ) on both the precursor and fragment ions ( 10 ppm and 0.6 da , respectively ) . high - confidence peptide identifications are used to recalibrate all precursors and fragment ion masses prior to a second search with narrower mass tolerances ( 4 ppm and 0.3 da for cid ) . carbamidomethyl cysteine and oxidised methionine were set as fixed and variable modifications , respectively . to validate the proteins , mascot and phenyx output files proteins with 2 unique peptides above a score t1 , or with a single peptide above a score t2 were selected as unambiguous identifications . the thresholds t1 , t2 , and t3 for each search engine were set by the target - decoy method to control the false discovery rate ( fdr ) of < 1% for protein identifications and for mascot , the following thresholds were used : t1 = 14 , t2 = 40 and t3 = 10 ; phenyx thresholds were set to 4.2 , 4.75 and 3.5 , respectively ( p value < 10 ) . the validated proteins retrieved by the two algorithms were merged , any spectral conflicts discarded and grouped according to shared peptides . non - specific binders were filtered from the drug pull - downs using the saint software ( version 2.3.4 ) ( choi et al . , 2011 ) . using protein spectral counts as a measure of protein abundance and comparing the data of from the drug pull - downs with the negative control drug competition experiments , we also compared saint probabilities with the fold - reduction of spectral count upon free compound competition representing a magnitude of effect . fold - reduction was computed as the ratio of median spectral counts observed in pull - downs with / without competition . in each condition , 4 spectral counts were available for the median ( 2 biological replicates and 2 technical for each ) . quantitative proteomic analysis was performed on tc1 cells harvested after three day compound treatment and lysed with 2% sodium dodecyl sulfate ( sds ) . samples were centrifuged at 5,000 r.c.f . for 2 min and the supernatant removed . lysis buffer ( 1ml ) was added and the samples incubated for 20 min at rt . the samples were then heated for 5 min at 99c and sonicated with a covaris sonicator to reduce viscosity . the supernatant was collected . the protein amount in each sample was determined by using the bicinchoninic protein assay ( bca , pierce biotechnology , thermo scientific , il ) followed by an adaptation of the filter - aided sample preparation method ( fasp ) ( manza et al . , 2005 , winiewski et al . , 2009 ) . samples were reduced with 100 mm dithiothreitol ( dtt ) at 99c for 5 min and transferred into vivacon 500 filter units ( vivaproducts ) . sds containing buffer was removed from the sample by centrifugation and exchanged with 8 m urea in 100 mm tris - hcl buffer . proteins were alkylated with 50 mm iodoacetamide and washed with 50 mm triethyl ammonium bicarbonate ( teab ) . porcine trypsin ( promega , madison , wi ) was used to digest the proteins in an enzyme to protein ratio of 1:100 ( w / w ) . for relative protein quantitation , the set of six samples ( three untreated and three drug - treated ) were separately derivatised with tmt 6-plex reagents ( thermofisher scientific , waltham , ma ) according to the instructions provided by the manufacturer . the samples were labeled with tmt 126 , 127 , 128 , 129 , 130 , 131 , respectively . the labeled tryptic digests were pooled and concentrated by solid phase extraction ( spe ) ( macrospin columns 30 - 300 g capacity , the nest group ) . samples were basified with 20 mm ammonium formate prior to injection onto a phenomenex column ( 150 2.0 mm gemininx - c18 3 m 110 ; phenomenex , torrance , ca , usa ) on an agilent 1200 series hplc ( agilent biotechnologies , palo alto , ca ) with uv detection at 214 nm . hplc solvent a consisted of 20 mm ammonium formate , ph 10 in 5% acetonitrile and solvent b consisted of 20 mm ammonium formate , ph 10 in 90% acetonitrile . peptides were separated at a flow rate of 100 l / min and eluted from the column with a two - step linear gradient from 0 to 70% solvent b. seventy two time - based fractions were collected , acidified , and pooled into 50 hplc vials based on the uv trace . after removal of organic solvent in a vacuum centrifuge , samples were reconstituted to 10 l with 5% formic acid ( bennett et al . , 2011 ) . individual fractions were analyzed at ph 2.4 on an agilent 1200 nano - hplc system ( agilent biotechnologies , palo alto , ca ) coupled to a hybrid linear trap quadrupole ( ltq ) orbitrap velos mass spectrometer ( thermofisher scientific , waltham , ma ) utilizing the xcalibur software version 2.1 . for data acquisition . single fractions were loaded onto a trap column ( zorbax 300sb - c18 5 m , 5 0.3 mm , agilent biotechnologies , palo alto , ca ) with a binary pump at a flow rate of 45 l / min . loading and washing solvents were composed of 0.1% trifluoracetic acid ( tfa ) in water ( solvent a ) and 0.1% tfa in 70% methanol and 20% isopropanol ( solvent b ) . the peptides were eluted by back - flushing from the trap column onto a 16 cm fused silica analytical column with an inner diameter of 50 m packed with c18 reversed - phase material ( reprosil - pur 120 c18-aq , 3 m , dr . lcms solvents were composed of 0.4% formic acid ( fa ) in water ( solvent a ) and 0.4% fa in 70% methanol and 20% isopropanol ( solvent b ) . elution was achieved with a 27 min gradient ranging from 3 to 30% solvent b , followed by a 25 min gradient from 30 to 70% solvent b and , finally , a 7 min gradient from 70 to 100% solvent b at a constant flow rate of 100 nl / min . the 10 most intense ions were isolated and fragmented by higher - energy collision - induced dissociation ( hcd ) for peptide identification and relative quantitation of tmt reporter ions . dynamic exclusion for selected ions was 60 s and a single lock mass at m / z 445.120024 ( si(ch3)2o)6 ) was used for internal mass calibration ( olsen et al . , 2005 ) with the target loss mass abundance of 0% . maximal ion accumulation time allowed was 500 ms and overfilling of the c - trap was prevented by automatic gain control set to 10 ions for a full ftms scan and 5 10 ions for msn hcd . intact peptides were detected in the orbitrap mass analyzer at a resolution of 30,000 with a signal threshold of 2,000 counts for triggering an msms event . the maximum ion scan time was set to 200 ms for acquiring 1 microscan at a resolution of 7,500 . peak extraction and conversion of .raw ms files into the .mgf format was performed with msconvert ( proteowizard library v2.1.2708 ) and the peak lists searched against the mouse swissprot database v2014.03_201403131 ( 24,830 sequences including isoforms obtained from varsplic.pl ( kersey et al . , 2000 ) and appended known contaminants ) with the search engines mascot ( v2.2.03 , matrixscience , london , uk ) and phenyx ( v2.5.14 , genebio , geneva , switzerland ) ( colinge et al . , 2003 ) . submission to the search engines was via a perl script that performs an initial search with relatively broad mass tolerances ( mascot only ) on both the precursor and fragment ions ( 10 ppm and 0.6 da , respectively ) . high - confidence peptide identifications are used to recalibrate all precursor and fragment ion masses prior to a second search with narrower mass tolerances ( 4 ppm and 0.025 da for hcd ) . carbamidomethyl cysteine and the tmt 6-plex tag on peptide n - termini and lysine residues were set as fixed modifications . oxidised methionine was set as a variable modification . to validate the proteins , mascot and phenyx output files proteins with 2 unique peptides above a score t1 , or with a single peptide above a score t2 were selected as unambiguous identifications . the thresholds t1 , t2 , and t3 for each search engine were set by the target - decoy method to control the false discovery rate ( fdr ) of < 1% for protein identifications and for mascot , the following thresholds were used : t1 = 12 , t2 = 45 and t3 = 10 ; phenyx thresholds were set to 5.5 , 9.5 and 3.5 , respectively ( p value < 10 ) . the validated proteins retrieved by the two search engines were merged , any spectral conflicts discarded and grouped according to shared peptides . using the isobar software ( breitwieser et al . , 2011 ) , quantitation of the relative fold change of each protein after drug treatment ( figure 1f ) was calculated by determining the ratios of tmt reporter ion intensities . the tmt reporter ion channels were normalized based on the protein median intensities and only peptides unique to each protein group were used for the quantitation . the isobar software employs statistical models to calculate the fold change of each protein and the associated p value based on the ratios and statistical confidence of each individual reporter channel . unless otherwise indicated , p values were calculated by student s t test ( graphpad prism ) and the exact values were indicated in figure legends . gene set enrichment analysis was conducted as previously described ( mootha et al . , 2003 , subramanian et al . , 2005 ) . raw data files for the rna sequencing analysis have been deposited in the ncbi gene expression omnibus under accession numbers geo : gse84592 , geo : gse73727 , and geo : gse84714 . p.c . conceived the project and designed the study ; t.f . and j.h .- s . developed cell lines ; j.l . , conducted and analyzed cell biological experiments in rodent cell lines ; j.l . and c .- h.l . , k.l.b . , and g.s .- f . performed and analyzed chemical proteomics experiments and global proteome studies ; r.a.r . , i.b . , j.k . , m.f . , f.m.p . , t.p . , and c. bock conducted and analyzed rna - seq experiments ; a. spittler performed fluorescence - activated cell sorting ; n.s . , r.a.k . m.c . , a.v . , f.a . , and p.c . conducted and analyzed mouse experiments ; f.b . and t.s . Output:	summarytype 1 diabetes is characterized by the destruction of pancreatic cells , and generating new insulin - producing cells from other cell types is a major aim of regenerative medicine . one promising approach is transdifferentiation of developmentally related pancreatic cell types , including glucagon - producing cells . in a genetic model , loss of the master regulatory transcription factor arx is sufficient to induce the conversion of cells to functional -like cells . here , we identify artemisinins as small molecules that functionally repress arx by causing its translocation to the cytoplasm . we show that the protein gephyrin is the mammalian target of these antimalarial drugs and that the mechanism of action of these molecules depends on the enhancement of gabaa receptor signaling . our results in zebrafish , rodents , and primary human pancreatic islets identify gephyrin as a druggable target for the regeneration of pancreatic cell mass from cells .
PubmedSumm2031	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: 99mtc pertechnetate scintigraphy is routinely used to detect testicular torsion and differentiate this condition from acute epididymo - orchitis . however , differentiation of torsion from hydrocele and testicular or scrotal abscess may be difficult . in this report , we present a 5-year old child in whom epididymitis with hydrocele was misdiagnosed as testicular torsion on scrotal scintigraphy . a 5-year - old boy presented to the pediatric emergency department with a complaint of left scrotal swelling and local tenderness for 7 h. doppler ultrasound ( us ) imaging showed mild hydrocele of the left scrotum with no e / o torsion of the left testis . static images at 5 and 10 min showed a halo - like pooling of tracer in the periphery of the left hemi - scrotum with a photopenic center and normal tracer uptake in the right hemi - scrotum [ figure 1 ] . a scintigraphic diagnosis of mid - phase testicular torsion of the left testis was made and the patient was immediately operated . however , it was discovered intraoperatively that the patient had epididymitis with a hydrocele and not torsion of the left testis . static images at 5 and 10 min show a halo - like pooling of tracer in the periphery of the left hemi - scrotum ( arrow ) with a photopenic centre . normal tracer uptake is seen in the right hemi - scrotum differentiation by scintigraphy between acute torsion , hydrocele , testicular or scrotal abscess , or even inguinal hernia can be difficult , and images must be interpreted in conjunction with clinical findings from scrotal transillumination . in this situation , sonography can provide useful complementary information.[14 ] in a previous study comparing scintigraphy and us in children with scrotal pain , doppler us was able to provide the diagnosis of epididymo - orchitis in cases of false - positive scintigraphy . a peri - testicular hyperemic rim on radionuclide scrotal scintigraphy epididymitis and hydrocele have also been reported to mimic testicular torsion on scintigraphy in adults.[68 ] the present study describes a halo - like appearance on scrotal scintigraphy caused by epididymitis with co - existing hydrocele in a clinical setting , suggestive of testicular torsion in a child . Output:	scintigraphic differentiation between acute torsion , hydrocele and testicular or scrotal abscess can be difficult . doppler sonography may provide useful complimentary information toward diagnosis . the authors describe a 5-year - old child where epididymitis with hydrocele was misdiagnosed as testicular torsion on scrotal scintigraphy .
PubmedSumm2032	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: polycystic ovary syndrome ( pcos ) is characterized by oligo - anovulation , clinical or biochemical hyperandrogenism and/or polycystic ovaries ( franks , 1995 ; knochenhauer et al . , 1998 ; fauser , 2004 ) . insulin resistance ( ir ) accompanied by compensatory hyperinsulinaemia constitutes another major biochemical feature of pcos ( nestler and jakubowicz , 1996 ; dunaif , 1997 ; nestler , 1997 ; jakubowicz et al . , 2001 ; fleming et al . , 2002 ) . the syndrome affects 510% of women during reproductive age ( asuncion et al . , 2000 ) . because of the link between ir and pcos , metformin has been put forward as a drug to induce ovulation in women with pcos . in fact , metformin either alone or in combination with clomifene citrate is now the most widely used insulin - sensitizer for ovulation induction in women with pcos ( nestler , 2002 ; glueck et al . , 2003 ; the rationale of this treatment was based on only a few small studies with conflicting results ( nestler et al . , 1998 ; recently , we demonstrated in an rct among newly diagnosed , therapy naive women with pcos that there were no significant differences in either ovulation rate , ongoing pregnancy rate or spontaneous abortion rate in women using clomifene citrate plus metformin compared with women using clomifene citrate ( moll et al . , 2006 ) . since then , our results considering ongoing pregnancy rates have been confirmed in another large rct ( legro et al . , 2007 ) . however , pcos is a heterogeneous condition and thus both studies included a heterogeneous group of women . surprisingly , there is only one small study involving 32 patients with pcos that has investigated the effect of metformin in specific subgroups . this study showed in a multivariable analysis that higher insulin , lower androstenedione and less severe cycle abnormalities appeared to be independent significant parameters for better response to metformin ( moghetti et al . , 2000 ) . increasing age and high waist hip ratio ( whr ) are known risk factors in developing ir and , as such , these factors may also affect clinical response to metformin ( despres , 1993 ; legro et al . , 1999 ; pasquali et al . , 2000 ; ncep expert panel , 2001 ; laine and wilson , 2007 ) . apart from these parameters , some authors suggest that metformin would most likely be beneficial in women with high bmi ( lord , et al . , 2003 ) . thus , although many investigators claim a beneficial effect of metformin in specific subgroups of women , the evidence is limited . we therefore wanted to determine whether co - treatment with metformin improves pregnancy rates compared with the standard treatment of clomifene citrate alone in subgroups of women based on clinical and biochemical variables . the data used in this study were collected in an rct that has been reported elsewhere ( moll et al . , 2006 ) . this double blinded trial took place from june 2001 to may 2004 in 20 dutch hospitals . all patients with chronic anovulation ( a menstrual cycle 35 days ) , world health organization type ii criteria ( normogonadotrophic normoestrogenic oligo- or anovulation ) , polycystic ovaries diagnosed by transvaginal ultrasonography and a desire to conceive were randomly allocated to clomifene citrate ( 50150 mg a day for five consecutive days in the beginning of a menstrual cycle , per os ) plus metformin ( 500 mg 4/day ) or to clomifene citrate plus placebo . primary exclusion criteria were other causes of anovulation , age over 40 years and liver , kidney or heart disease / failure ( i.e. abnormal results on liver function tests , serum creatinine concentration > 95 mol / l or a history of heart disease / failure ) and sperm quality indicating male subfertility ( total motile count < 10 10 ) . randomization was carried out in the coordinating centre ( amc , amsterdam ) , using computer - generated blocks of four . the randomization was stratified per centre , and the centres received blinded , numbered containers with medication . patients continued to take the study medication until a positive pregnancy test ( 4 weeks after the first day of menstruation ) , six ovulatory cycles or clomifene citrate resistance occurred , whichever came first . women had to discontinue their medication as soon as they had a positive pregnancy test as the safety and benefit of using metformin during pregnancy have not yet been proven . the study was approved by the institutional review boards of all hospitals . written informed consent was obtained from all participants . at the start of the study , we measured height , weight , whr and plasma testosterone . total plasma testosterone concentrations were determined using an in - house radioimmunoassay , without extraction and chromatography and with tritiated testosterone as label ( pratt et al . , 1975 ) . intra - assay and inter - assay coefficients of variation were 4 - 7% and 59% , respectively . subgroup analysis was performed on age , bmi , whr , testosterone , 2 h glucose and homa ( homeostatic model assessment for assessing insulin sensitivity ) as a measure of ir ( ehrmann et al . ncep expert panel , 2001 ; fauser , 2004 ; samaras et al . , 2006 ) . since not all data were available for all women , in some analyses only a proportion of the women the parameters to be analysed were chosen , after completion of the trial , according to the most recent literature ( despres , 1993 ; legro et al . , 2000 ; ncep expert panel , 2001 ; lord , et al . , 2003 ; laine and wilson , 2007 ) . for this subgroup analysis , we explored differences in treatment effect on ongoing pregnancy rate in subgroups defined on age , bmi , whr , plasma testosterone , 2 h glucose level and ir evaluated by homa - ir . for this purpose , we performed logistic regression analysis including treatment , the subgroup indicator and the interaction between treatment and subgroup . we assessed the linearity of the association between the continuous variables age , bmi , whr , testosterone , 2 h glucose level and homa and the ongoing pregnancy rate using spline functions ( harrell et al . , 1988 ) . non - linear associations were accommodated by redefining the corresponding variables . by testing the interaction in logistic regression , if the interaction is statistically significant , this is a clear indication that the effect of treatment differs between subgroups ( matthews and altman , 1996 ) . next , to explore potential interesting subgroup features , we combined subgroups for which the interaction with treatment was p < 0.20 . in all analyses , for this subgroup analysis , we explored differences in treatment effect on ongoing pregnancy rate in subgroups defined on age , bmi , whr , plasma testosterone , 2 h glucose level and ir evaluated by homa - ir . for this purpose , we performed logistic regression analysis including treatment , the subgroup indicator and the interaction between treatment and subgroup . we assessed the linearity of the association between the continuous variables age , bmi , whr , testosterone , 2 h glucose level and homa and the ongoing pregnancy rate using spline functions ( harrell et al . , 1988 ) . non - linear associations were accommodated by redefining the corresponding variables . by testing the interaction in logistic regression , if the interaction is statistically significant , this is a clear indication that the effect of treatment differs between subgroups ( matthews and altman , 1996 ) . next , to explore potential interesting subgroup features , we combined subgroups for which the interaction with treatment was p < 0.20 . in all analyses , baseline characteristics are presented in table i. out of the 111 women in the clomifene citrate plus metformin group , 44 ( 40% ) reached an ongoing pregnancy versus 52 ( 46% ) of the 114 women in the clomifene citrate group , a difference which was not significant [ relative risk ( rr ) 0.87 ; 95% confidence interval ( ci ) 0.641.2 ] . a significantly larger proportion of patients in the metformin group discontinued treatment because of side effects ( 18/111 versus 6/114 ; rr 2.9 ; 95% ci 1.27.1 ) . the total dropout rate in both treatment arms was not statistically different ( 28/111 versus 21/114 ; rr 1.3 ; 95% ci 0.822.2 ) . baseline characteristics of patients with pcos who were treated with clomifene citrate plus metformin or clomifene citrate alone . homa , homeostatic model assessment for assessing insulin sensitivity ; whr , waist hip ratio . the data for 2 h glucose level were available in 80% of patients , and for homa , in 50% of patients . in women 28 years of age , the effect of metformin versus placebo showed a small trend towards a different effect compared with the effect of metformin versus placebo in women < 28 years ( p - value of interaction 0.11 ) ( table ii ) . in women with a whr 0.85 , the effect of metformin versus placebo was significantly different from the effect of metformin versus placebo in women with a whr < 0.85 ( p - value of interaction 0.012 ) ( table ii ) . pregnancy rates for subgroups of women with pcos treated with clomifene citrate plus metformin or clomifene citrate alone . we did not find significant differences in effect of treatment for groups based on bmi , plasma testosterone , 2 h glucose values or homa . on the basis of these findings , we combined age and whr and created four subgroups . we found a significantly different chance of reaching an ongoing pregnancy of metformin versus placebo between these subgroups ( p - value of interaction 0.014 ) ( table iii ) . a positive effect of metformin versus placebo on pregnancy rate was found in older women ( 28 years ) with a whr 0.85 , a negative effect of metformin versus placebo in young women regardless of their whr and no effect in the older , not viscerally obese women ( fig . 1 ) . relative risks and 95% confidence intervals for combined subgroups of women treated with clomifene citrate ( clomid ) plus metformin or clomifene citrate alone with respect to pregnancy rates . pregnancy rates for combined subgroups in women with pcos treated with clomifene citrate plus metformin or clomifene citrate alone . in the present analysis , there was a significantly different chance of an ongoing pregnancy of metformin versus placebo between subgroups , based on age and whr . a positive effect of metformin versus placebo on pregnancy rate was found in older women with a high whr , a negative effect of metformin versus placebo in young women regardless of their whr and no effect in older , not viscerally obese women . we have included women with pcos , who had never used clomifene citrate and were seeking treatment for their fertility problems for the first time . our study group reflects the largest group of women with pcos a fertility clinic will see and treat . our results contradict those from the only other subgroup analysis , where no differential effect of metformin was shown in subgroups of women based on age and whr ( moghetti et al . , 2000 ) : this study did show that in multivariable analysis higher insulin , lower androstenedione and less severe cycle abnormalities appeared to be significant parameters for better response to metformin , variables we did not assess in our study . however , our data may be more valid , because we analysed over 200 patients , 10 times more than in the moghetti et al . ( 2000 ) subanalysis and because our study was based on a double - blinded rct as opposed to the open protocol used in the previous subanalysis . the most likely reason why metformin has a beneficial effect on pregnancy rates in older and viscerally obese women is that ir is common in these women ( cowie et al . , however , we did not find a significant interaction between pregnancy rates and the outcome of other surrogate markers of ir , including bmi , plasma testosterone , 2 h glucose value and homa . this is probably due to the fact that whr is the most powerful predictor of ir of all surrogate markers ( rexrode et al . , 1998 ; lord et al . , 2003 ; alberti et al . , 2006 ; lord et al . , 2006 ) . we did not measure ir directly by means of the gold standard , the glucose clamp technique , since this is considered to be too invasive and burdensome to be part of a large clinical trial in outpatients ( legro et al . the lack of effect of metformin in patients with high bmi , increased plasma testosterone , higher 2 h glucose levels and homa may be real , if confirmed by further studies . however , another possibility is that the present study did not have enough power to show a difference . this might especially hold true for homa , since we only had data on half of the patients . a limitation of the study is that we did not collect all parameters in all patients . in particular , fasting insulin is missing in 49% of the patients . when comparing baseline characteristics of the patients for whom data on insulin were available with those for whom data were not available , we found no significant differences ( data not shown ) . the lack of data appears , therefore , to be due to a random effect of not executing the complete study protocol rather than a deliberate omission of patients with certain clinical or biochemical characteristics . after our study protocol was written ( 1999 ) and our study performed ( 20012004 ) , it became clear that testosterone assays show a wide inter - laboratory variability ( kane et al . , 2007 ; rosner et al . , 2007 ) . when we had finished the trial , it was impossible to adjust for this . the calculations performed in this trial were therefore performed without correcting for this inter - laboratory imprecision , as is the case in all previous literature . it is possible that the variation is sufficiently high to render the result not significant . this relatively new understanding of the variability of testosterone assays may have great impact , not only on existing but also on future literature on pcos , especially in multicentre studies . our rct was not powered for these subgroup analyses ; therefore , the data need to be interpreted with great caution . nonetheless , we feel that our data argue for performing a trial comparing metformin and clomifene citrate with clomifene citrate alone in older visceral obese women with pcos . although our initial randomized trial showed no benefit for metformin , our current findings suggest that metformin may be an effective addition to clomifene citrate in infertile , older and visceral obese women with pcos . randomized studies should be performed specifically for this subgroup , or data from other existing trials should be re - analysed before we can consider implementing these findings in clinical practice . Output:	backgroundan rct among newly diagnosed , therapy naive women with polycystic ovary syndrome ( pcos ) showed no significant differences in ovulation rate , ongoing pregnancy rate or spontaneous abortion rate in favour of clomifene citrate plus metformin compared with clomifene citrate . we wanted to assess whether there are specific subgroups of women with pcos in whom clomifene citrate plus metformin leads to higher pregnancy rates.methodssubgroup analysis based on clinical and biochemical parameters of 111 women randomized to clomifene citrate plus metformin compared with 114 women randomized to clomifene citrate plus placebo . the data for age , bmi , waist hip ratio ( whr ) and plasma testosterone were available in all women , 2 h glucose in 80% of women and homeostatic model assessment for assessing insulin sensitivity ( homa ) in 50% of women.resultsof the women who were allocated to the metformin group , 44 women ( 40% ) reached an ongoing pregnancy . in the placebo group , 52 women ( 46% ) reached an ongoing pregnancy . there was a significantly different chance of an ongoing pregnancy for metformin versus placebo between subgroups based on age and whr ( p = 0.014 ) . there was a positive effect of metformin versus placebo on pregnancy rate in older women ( 28 years ) with a high whr , a negative effect of metformin versus placebo in young women ( < 28 years ) regardless of their whr and no effect in older , not viscerally obese women . no significant differences in effect of treatment were found for groups based on bmi , 2 h glucose , homa or plasma testosterone.conclusionsmetformin may be an effective addition to clomifene citrate in infertile women with pcos , especially in older and viscerally obese patients .
PubmedSumm2033	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the lipid benzoquinone sorgoleone ( 2-hydroxy-5-methoxy-3-[(8z , 11z)-8 , 11 , 14-pentadecatriene]-p - benzoquinone ) is a potent allelochemical that is produced as an oily exudate from the root hairs of sorghum ( sorghum bicolor ) ( czarnota et al.2001 ; dayan et al.2007 ) , along with a variety of structural analogues ( kagan et al.2003 ) responsible for inhibiting weeds and harmful algae ( einhellig and souza 1992 ; nimbal et al.1996 ; rimando et al.2003 ; uddin et al.2009 , 2012 ; weston and czarnota 2001 ) . sorgoleone is released from the roots of sorghum continually during the growing season , which may prolong its presence in soil ( dayan 2006 ; dayan et al.2009 ; weidenhamer 2005 ) . sorgoleone , its resorcinol analogue , and other related hydroquinones are produced solely by living root hairs and are exuded as golden - coloured droplets from the tips of root hairs ( czarnota et al.2003a ; dayan et al.2009 ) . however , sorgoleone biosynthesis is linked intrinsically to the presence of living root hairs ( czarnota et al.2001 ; yang et al.2004 ) . plants use a variety of sequestration and transport mechanisms to move and export bioactive products safely into the rhizosphere ( weston et al.2012 ) . specific root exudates and rhizodeposits are important messengers for chemical communication systems that help regulate the interactions between roots and soil organisms , and mediate processes in response to environmental stressors ( uren 2000 ) . roots , when they are under stress or encountering challenges in the rhizosphere , often react by releasing low - molecular - weight compounds , including amino acids , organic acids , and phenolics , as well as proteins . these secretions can protect the plant , thus initiating a negative form of communication in the rhizosphere ( bertin et al.2003 ) . alternatively , these exudates can elicit symbiotic responses that initiate legume rhizobium nitrogen fixation or attract common soil microbes , which are positive forms of communication ( mathesius and watt 2011 ; peters et al.1986 ; shi et al.2011 ) sorgoleone biosynthesis appears to occur exclusively in root hair cells , which in sorghum appear as cytoplasmically dense cells filled with numerous osmiophilic deposits ( czarnota et al . these deposits presumably are associated with the rhizosecretion of sorgoleone , which can constitute as much as 85 % of the exudate dry weight in some cultivars ( bertin et al.2003 ; czarnota et al.2001 , 2003b ; duke 2003 ; rimando et al.2003 ) . the cell - specific localization and prolific output of the sorgoleone biosynthetic pathway renders the use of expressed sequence tag ( est ) analysis an obvious method to isolate genes encoding sorgoleone biosynthetic enzymes ( baerson et al.2008 ) . labeling studies performed by fate and lynn ( 1996 ) first demonstrated that biosynthesis proceeds through the action of an alkylresorcinol synthase ( ars ) , a novel type iii polyketide synthase that acts on fatty acyl - coa starter units ( baerson et al.2008 ) . subsequently , both the predicted 5-n - pentadecatrienyl resorcinol and a 3-methyl ether derivative of sorgoleone were identified in sorghum root extracts , indicating that dihydroxylation of the resorcinol ring is preceded by o - methylation at the 3-hydroxyl position ( baerson et al.2008 ; dayan et al.2003 ; fate and lynn 1996 ) . once released into the rhizosphere , the resulting chemically unstable hydroquinone rapidly oxidizes to the bioactive sorgoleone benzoquinone , where it may persist in soil for extended periods ( czarnota et al.2001 ; einhellig and souza 1992 ; gimsing et al.2009 ; netzly et al.1988 ) . synthesis of sorgoleone from the available palmitoleoyl - coa would , therefore , likely require , at minimum , the participation of a 12 and 15 fatty acid desaturase ( des ) , an ars , 3-o - methyltransferase ( omt ) , and cytochrome p450 ( baerson et al.2008 ) ( fig . 1proposed biosynthestic pathway of the allelochemical , sorgoleone ( baerson et al.2008 ) proposed biosynthestic pathway of the allelochemical , sorgoleone ( baerson et al.2008 ) root hair development has been used as an important model to study the mechanisms of cell patterning and differentiation in higher plants ( schiefelbein 2000 ) . mutational analysis has revealed that several genes in arabidopsis function in the specification of root epidermal cell types . among these , the ttg ( galway et al.1994 ) , gl2 ( masucci and schiefelbein 1996 ) , and wer ( wada et al.1997 ) genes are the best characterized . studies on arabidopsis mutants , such as axr1 and ctr1 , indicate that auxin and ethylene function in root hair initiation . both methyl jasmonate ( meja ) and jasmonic acid ( ja ) have a pronounced effect on promoting root hair formation in arabidopsis . the effect of meja and ja on root hair formation is blocked by ethylene inhibitors ag or aminoethoxyvinylglycine ( avg ) . the stimulatory effects of meja and ja also are diminished in ethylene - insensitive mutants etr1 - 1 and etr1 - 3 ( zhu et al.2006 ) . to the best of our knowledge , the effects of jasmonates on gene expression in sorghum roots have not been reported . the goal of this study was to correlate gene expression levels with the biosynthesis of sorgoleone after exposure to ja or meja for different time periods . such knowledge may be useful for the future development of a bioherbicide by increasing levels of sorgoleone through molecular genetic manipulation . sorghum seeds of the cultivar chalsusu , which has a high sorgoleone content ( uddin et al.2009 ) , were used . seeds were treated with benomyl ( a powdery fungicide , 100 g dissolved in 20 l ) for 4 h , and then rinsed several times in distilled water . seeds ( 25/petri dish ) were placed on the surface of sterile whatman # 1 filter paper ( diam , 90 mm ) saturated with different concentrations of ja or meja placed in sterile petri dishes ( 100 40 mm ) . the dishes were then placed in a growth chamber at 25 c under standard cool white fluorescent tubes with a flux rate of 550 mol s m and a 16-h photoperiod for 1 week . to determine the concentrations of ja and meja that promote maximum sorgoleone biosynthesis , sorghum seeds were imbibed with solution containing different concentrations ( 0 , 0.5 , 1.0 , 5.0 , 10 , 50 , 100 , 250 , and 500 m ) of either ja or meja for 1 week . to examine how gene expression and sorgoleone accumulation in response to meja and ja changed over time , 5-day - old sorghum seedlings were treated once with 5.0 m ja or meja , and then their roots were harvested for analysis at multiple time points ( 0 , 3 , 6 , 12 , 24 , 36 , 48 , 60 , and 72 h after treatment ) . roots samples were stored in sealed clear polyethylene plastic bags at 80 c until use . there were four replicates for each time point , and the entire experiment was repeated twice . sorgoleone was extracted according to the procedures described previously ( czarnota et al.2003b ; netzly and butler 1986 ; netzly et al.1988 ) , except that methanol was used as a solvent instead of methylene chloride ( uddin et al.2010 ) . seedling roots were excised and immersed in methanol ( 1:20 w / v ) for 30 s to extract . the dried extract was dissolved in methanol ( 1 mg ml ) , and the solution was filtered through a poly filter ( pore size , 0.45 m ) . hplc quantification of sorgoleone was performed using the futecs ns-4000 hplc system ( futecs co. ltd . , daejeon , korea ) , with a c18 column ( 250 4.6 mm , particle size 5 m ; rstech , daejeon , korea ) . sorgoleone was detected at 280 nm with a waters tunable absorbance detector after injection of 20 l of the methanol solubilized crude extract sample . the column flow rate was 1 ml min with a 40 min total run time for each sample . the amount of sorgoleone was calculated on the basis of a standard curve obtained from a purified sample . the sorgoleone standard was provided by franck dayan , united states department of agriculture - agricultural research service ( usda - ars ) , natural products utilization research unit . from the meja and ja treatment root samples only the most optimal concentrations of meja and ja ( 5.0 m ) for maximum sorgoleone concentration were selected . total rna was isolated from a root of s. bicolor using tri - reagent ( mrc , usa ) and rneasy plant mini kit ( qiagen , valencia , ca , usa ) . for quantitative real - time polymerase chain reaction ( qrt - pcr ) , 1 g of total rna was reverse - transcribed according to the manufacturer s protocol ( revertraace - a , toyobo , japan ) using an oligo ( dt)20 primer . the cdna mixtures were used as templates for qrt - pcr . for transcript - level analysis by qrt - pcr , rnas from auxin treated sorghum roots were harvested , and single - stranded cdnas were synthesized from the isolated total rna by using the aforementioned protocols . the gene - specific primer sets were designed for qrt - pcr ( table 1 ) . specific primers for des2 ( ef206347 ) , des3 ( ef206348 ) ( pan et al.2007 ) , ars1 ( xm_002441794 ) , ars2 ( xm_002449699 ) ( cook et al.2010 ) , and omt3 ( ef189708 ) ( baerson et al.2008 ) were obtained from ncbi . the pcrs were carried out in triplicates for 40 cycles on a miniopticon ( bio - rad laboratories , hercules , ca , usa ) using the qiagen quantitect sybr green pcr kit . the pcrs were pre - denatured at 95 c for 5 min , then subjected to 40 cycles of amplifications with the following thermal cycles : denaturation for 30 s at 95 c , annealing for 30 s at 55 c ; extension for 30 s at 72 c . identical pcr conditions were used for all targets.table 1primer set for quantitative rt - pcr - mediated amplification of five transcripts that encode enzymes involved in sorgoleone synthesis from sorghumprimer namesequences ( 5 to 3)des2-qfactaccagttcgaccccactccdes2-qrccgccttctcttcctgatttctdes3-qfctgaccatcacctcaacgacacdes3-qrcattgcaccaactgacttcaccars1-qfggagtaccacctctccagcaaars1-qrcagtttgtcgtcctccagaccars2-qfatcttcgtgctcgatgagttgars2-qratttccctccagttccaggttomt3-qfgtttgttgggggtgacatgtttomt3-qrcgtctccagaagcttggtgtct primer set for quantitative rt - pcr - mediated amplification of five transcripts that encode enzymes involved in sorgoleone synthesis from sorghum all data were analyzed using the sas 9.2 software ( sas 2010 ; sas institute inc . , analysis of variance was performed and means were separated by duncan multiple range test ( dmrt ) standard deviations were provided to indicate the variation associated with a particular mean value . the root growth pattern , secondary root , root hair formation , and root growth of sorghum were influenced by jasmonates . photographs of sorghum root revealed variation in rooting patterns among the treatments ( figs . 2 and 3 ) . the number of secondary or branching roots was higher at lower concentrations of jasmonate treatment , and the highest number of secondary roots was observed at 5 m ( fig . ja treatment produced a slightly higher number of secondary roots compared with that induced by meja treatment . greater numbers of healthy root hairs were formed in response to ja and meja treatments compared with those formed in the control . root weight increased slowly with increases in the concentration of jasmonates ( ja and meja ) up to 5.0 m , and then declined slowly with further increasing concentrations of either compound ( fig . 2photographs showing the root growth pattern of sorghum seedlings 7 days after methyl jasmonate ( meja ) treatment . a = control , b = meja ( 0.5 m ) , c = meja ( 1.0 m ) , d = meja ( 5.0 m ) , e = meja ( 10 m ) , f = meja ( 50 m ) , g = meja ( 100 m ) , h = meja ( 250 m ) , i = meja ( 500 m)fig . 3photographs showing the root growth pattern of sorghum seedlings 7 days after jasmonic acid ( ja ) treatment . a = control , b = ja ( 0.5 m ) , c = ja ( 1.0 m ) , d = ja ( 5.0 m ) , e = ja ( 10 m ) , f = ja ( 50 m ) , g = ja ( 100 m ) , h = ja ( 250 m ) , i = ja ( 500 m)fig . 4photomicrograph of sorghum roots showing root hairs 7 day after jasmonate treatment ( bar=90 m ) . a = control , b = methyl jasmonate ( meja ) ( 5 m ) , c = jasmonic acid ( ja ) ( 5 m)fig . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p<0.05 according to a duncan multiple range testfig . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test photographs showing the root growth pattern of sorghum seedlings 7 days after methyl jasmonate ( meja ) treatment . a = control , b = meja ( 0.5 m ) , c = meja ( 1.0 m ) , d = meja ( 5.0 m ) , e = meja ( 10 m ) , f = meja ( 50 m ) , g = meja ( 100 m ) , h = meja ( 250 m ) , i = meja ( 500 m ) photographs showing the root growth pattern of sorghum seedlings 7 days after jasmonic acid ( ja ) treatment . a = control , b = ja ( 0.5 m ) , c = ja ( 1.0 m ) , d = ja ( 5.0 m ) , e = ja ( 10 m ) , f = ja ( 50 m ) , g = ja ( 100 m ) , h = ja ( 250 m ) , i = ja ( 500 m ) photomicrograph of sorghum roots showing root hairs 7 day after jasmonate treatment ( bar=90 m ) . a = control , b = methyl jasmonate ( meja ) ( 5 m ) , c = jasmonic acid ( ja ) ( 5 m ) effects of jasmonates for the production of secondary roots in sorghum roots . each point is the mean of four replicates with an average of 10 seedlings for each replicate . means with the same letter adjacent to the symbols are not different at p<0.05 according to a duncan multiple range test dose - response curves showing the effects of jasmonates on root growth . each point is the mean of four replicates with an average of 10 seedlings for each replicate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test the dose response experiment indicated that sorgoleone content varied widely depending on the concentrations of exogenous jasmonates ( fig . 7 ) . root dry weight and levels of sorgoleone increased with increases in both ja and meja concentrations up to 5.0 m , and then declined with increasing concentrations of either of the jasmonates . compared with untreated controls , sorgoleone levels were 4.1 , 2.3 , and 1.7-fold higher after treatment with 5 , 1.0 , and 0.5 m of ja , respectively ( fig . the level of sorgoleone was highest at a concentration of 5.0 m ja , with a level approximately 10 times lower ( 2.8 mg / g root fresh weight ) in roots exposed to 500 m ja.fig . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test dose - response curves showing the effects of jasmonates on sorgoleone production . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test the concentration - dependent effects of meja on sorgoleone accumulation were almost identical for meja , with 5.0 m meja producing more sorgoleone than any other concentration . compared with an untreated control , sorgoleone levels were 3.4 , 2.1 , and 1.8-fold higher after exposure to 5 , 1.0 , and 0.5 m of exogenous meja , respectively ( fig . exposure to concentrations of meja greater than 5.0 m caused a less dramatic decrease in sorgoleone content than observed for ja ( fig . 7 ) . after exposure to 500 m of meja , the level of sorgoleone was 5.0 mg / g root fresh weight , which is 4.6 times lower than the highest content of sorgoleone observed in roots incubated with 5 m of meja . sorgoleone accumulation was maximal after treatment with 5.0 m of either ja or meja , with the next highest levels of accumulation observed following treatments with jasmonates at 1.0 m and 0.5 m , respectively ( fig . 7 ) . in the case of exposure to 5.0 m meja , increasing exposure time caused sorgoleone levels to increase for up to 72 h after the beginning of the treatment ( fig . 8) . compared with the control at 0 h , 5.0 m meja produced 2.3 times more sorgoleone 72 h after the beginning of the treatment . in the case of exposure to 5.0 m ja , increasing exposure time caused sorgoleone levels to increase for up to 60 h after the beginning of the treatment ( fig . 5.0 m ja produced almost twice as much sorgoleone at 60 h after treatment . sorgoleone levels were highest at 24 h , when they were 1.2 times higher than levels in control plants at 0 h.fig . 8time - courses of sorgoleone production induced by application of jasmonates ( 5.0 m ) to sorghum roots . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test time - courses of sorgoleone production induced by application of jasmonates ( 5.0 m ) to sorghum roots . each point is the mean of four replicates with an average of 10 seedlings for each replicate . ja , jasmonic acid , meja , methyl jasmonate . means with the same letter adjacent to the symbols are not different at p < 0.05 according to a duncan multiple range test the effects of ja and meja on the levels of transcript of five genes involved in sorgoleone biosynthesis were monitored . levels of transcript of the sorgoleone biosynthetic genes , des2 , des3 , ars1 , ars2 , and omt3 increased after exposure to either ja or meja at 5.0 m . the extent to which the levels of transcript were affected by either ja or meja was dependent on the length of time the plants were exposed to jasmonates . shortly after the application of ja , levels of transcripts were comparable to those in control seedlings , although higher transcript levels were noted than in control seedlings at some subsequent time points ( fig . the time after initial exposure when transcript levels were highest differed among the genes . in the case of ars1 , the highest levels of transcript accumulation occurred 48 h after initial exposure . at 48 h levels of ars1 transcript were 1.8-fold higher than in untreated seedlings at 0 h ( fig . 9 ) . levels of ars2 and des2 transcript were highest 36 h after treatment , with increases of 5.2 and 3.3-fold , respectively when compared with untreated seedlings at 0 h ( fig . the largest increase in transcript accumulation was apparent for the des3 gene , 12 h after ja treatment , when des3 transcripts were 3.2-fold more abundant than in untreated seedlings at 0 h ( fig . 9time course of induction of sorgoleone biosynthetic genes in response to jasmonic acid at 5.0 m in the sorghum root . con , control ; ja , jasmonic acid . means with the same letter in both column bars are not different at p < 0.05 according to a duncan multiple range test time course of induction of sorgoleone biosynthetic genes in response to jasmonic acid at 5.0 m in the sorghum root . con , control ; ja , jasmonic acid . means with the same letter in both column bars are not different at p < 0.05 according to a duncan multiple range test transcripts of different sorgoleone biosynthetic genes responded differently to exposure to meja . the times when the largest increase in transcript abundance was apparent after treatment with ja differed among the genes . gene expression was lower than in the untreated control shortly after initial exposure of sorghum seeds to meja . compared with transcript levels in seedlings not treated with meja , transcript accumulation was highest for most of the genes 36 h after meja treatment ( fig . 10 ) , in particular for omt3 ( increased up to 8.1-fold ) , des2 ( increased up to 5.2-fold ) , ars2 ( increased up to 2.8-fold ) , and des3 ( increased up to 2.4-fold ) . levels of des2 transcript were 4.1-fold higher 3 h after treatment with meja than in seedlings that received no meja ( control ) . the highest level of ars1 transcript was observed 24 h after treatment with meja , and was 3.7-fold higher than the level of ars1 transcript in seedlings not treated with meja ( 0 h ) ( fig . 10).fig . 10time course of induction of sorgoleone biosynthetic genes by application of methyl jasmonate at 5.0 m treatment in the sorghum root . con , control ; meja , methyl jasmonate . means with the same letter in both column bars are not different at p<0.05 according to a duncan multiple range test time course of induction of sorgoleone biosynthetic genes by application of methyl jasmonate at 5.0 m treatment in the sorghum root . each value is the mean of 3 replicates . con , control ; meja , methyl jasmonate . means with the same letter in both column bars are not different at p<0.05 according to a duncan multiple range test several possible mechanisms might account for the increase in root hair density ( ma et al.2001 ) . second , increased abundance of trichoblasts with branched root hairs may contribute to an increase in root hair density . third , atrichoblasts may be recruited into trichoblasts during root hair development under certain conditions . our results suggest that the initiation of recruitment of more trichoblast cells into root hairs may be an important cause of meja - induced root hair development . the induction of branched hair formation may be a novel effect of meja on root hair development . investigation of whether jas activate root hair formation via ethylene in arabidopsis revealed that the effects of jas were abolished in the ethylene - insensitive mutants etr1 - 1 and etr1 - 3 , or by inhibitors of ethylene action ( ag ) or biosynthesis inhibitors ( avg ) . furthermore , ibuprofen and sham , which inhibit ja biosynthesis , repressed acc - driven or eto1 - 1-induced root hair formation . collectively , these reports support a role for the interaction between jas and ethylene in the regulation of root hair development in arabidopsis . ( 2003a ) confirmed that sorghum root hairs are physiologically active , and that they contain a complex network of smooth endoplasmic reticulum and possibly golgi bodies . small globules of cytoplasmic exudates also were observed to deposit an oily material between the cell wall and the plasma membrane near the root hair tips . ( 1992 ) indicated that sorgoleone production is quite sensitive to environmental conditions , and it is well documented that sorgoleone production depends mainly on root hair formation ( czarnota et al.2003b ; dayan 2006 ; dolan 2001 ; yang et al.2004 ) . herein we report that root growth pattern , secondary root , root hair formation , and root growth of sorghum are influenced by the use of jasmonates . our study shows that sorghum roots develop many branches ( fig . 4 ) , as well as more root hairs ( fig . 5 ) , when the proper concentrations of jasmonates ( up to 5 m ) are applied . clearly , healthy root hairs in higher numbers than those in the control were formed in response to ja and meja treatment . root weight increased slowly with increased concentrations of jasmonates ( ja and meja ) up to 5.0 m and then declined slowly with further increasing concentrations of these compounds ( fig . zhu et al . ( 2006 ) have reported that jasmonates promote root hair formation in arabidopsis , which agrees with our findings . jasmonates are a class of plant hormones that mediate various aspects of gene and metabolic regulation , defense , stress responses , reproduction , and possibly communication . the role of ja in plants has received considerable attention , and various modes of action for ja and its methyl ester ( meja ) have been proposed ( farmer 2007 ; farmer and ryan 1990 ) . farmer ( 2007 ) , working largely with the model plant arabidopsis thaliana , concluded that the two groups take different routes to the genes involved . ( 2007 ) analyzed eight related gene transcripts of unknown function that are rapidly upregulated in developing arabidopsis stamens treated with jasmonates . by contrast , chini et al . ( 2007 ) characterized a mutant called jasmonate - insensitive3 - 1 ( jai3 - 1 ) . working with different genes , both groups found that over - expression of the full - length transcripts had little or no effect on jasmonate perception . however , over - expression of shortened forms that encode proteins truncated at the carboxyl terminus reduced sensitivity to jasmonates . additionally , although full - length proteins were unstable in vivo when treated with jasmonates , proteins with a truncated carboxyl terminus resisted degradation in response to such treatment . ( 2011 ) reported that induced systemic resistance ( isr ) occurs in response to the application of meja , and systemic acquired resistance ( sar ) occurs after treatment with salicylic acid or benzothiadiazole , whereas some of the arabidopsis genotypes affected in defense signaling show altered numbers of culturable bacteria in their rhizospheres ; however , the effects depend on soil type . genes involved in jasmonate biosynthesis , secondary metabolism , and cell - wall formation , and those encoding stress - protective and defense proteins are up - regulated by meja treatment ( cheong and choi 2003 ) . in this study , five transcripts - des2 , des3 , ars1 , ars2 , and omt3-that encode enzymes involved in sorgoleone biosynthesis were shown to accumulate in response to treatment with ja and meja . levels of omt3 transcript accumulated to levels 8.1-fold higher than in seedlings not treated with meja . this observation is supported by other reported documents ( bonfill et al.2011 ; kim et al.2009 ; korsangruang et al.2010 ; lystvan et al.2010 ; rhee et al.2010 ) , where they investigated the capability of ja and its methyl ester , meja , to induce the accumulation of various secondary metabolites , and showed higher levels of expression for their respective genes and also from previous work ( uddin et al.2011 ) , where they observed that sorgoleone biosynthetic genes are up or down - regulated by auxins . the application of phytohormones to excised sections or organs ( e.g. , root , elongating region of hypocotyl , basal region of hypocotyl , and plumule ) of intact soybean seedlings resulted in the accumulation of mrnas for pghl-4 to levels that are many - fold greater than those found in untreated organs of seedlings ( hagen and guilfoyle 2002 ) . the results obtained using jasmonates to enhance sorgoleone production suggest that sorghum roots benefit from exposure to jasmonates by developing more sorgoleone - rich root hairs , showing different rooting pattern and root weight variation among the jasmonate treatments . transcript accumulation was apparent for all of the seven genes tested , particularly omt3 , for which transcript levels increased up to 8.1-fold after a 36 h exposure to meja compared with omt3 transcript levels in untreated seedlings . in summary , these findings suggested that jasmonates ( ja and meja ) are potent substances for promoting root hair formation and sorgoleone accumulation and up - regulating gene expression in sorghum . further research should focus on the involvement of these products in the regulation of root responses to stress . Output:	this study investigated the roles of jasmonates in the regulation of sorgoleone accumulation and the expression of genes involved in sorgoleone biosynthesis in sorghum roots . both methyl jasmonate ( meja ) and jasmonic acid ( ja ) substantially promoted root hair formation , secondary root development , root weight , and sorgoleone accumulation in sorghum roots . sorgoleone content varied widely depending on the concentration of ja or meja and the duration of their application . root weight and sorgoleone accumulation were highest after the application of ja or meja at a concentration of 5.0 m , and then declined with increasing concentrations of jasmonates . at 5.0 m , ja and meja increased sorgoleone content by 4.1 and 3.4-fold , respectively . transcript accumulation was apparent for all genes , particularly for the o - methyltransferase 3 gene , which increased in expression levels up to 8.1-fold after a 36-h exposure to meja and 3.5-fold after a 48-h exposure to ja . the results of this study pave the way for more effective biosynthesis of sorgoleone , an important and useful allelochemical obtained from a variety of plant species .
PubmedSumm2034	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: human nature is uniquely characterized by the flexibility , complexity , and sophistication with which thought and behavior can be deployed in the service of a goal . this ability is thought to depend on cognitive control , a collection of mechanisms , including perceptual selection , response biasing , and online maintenance of contextual or goal information , by which the human cognitive system adaptively configures itself to optimally perform specific tasks ( miller and cohen , 2001 ; braver et al . , 2002 ) . most of the goals pursued in daily life are emotionally or motivationally meaningful i.e. , to obtain outcomes that are pleasurable or important for survival , and avoid outcomes that are not . it has long been understood that such affective significance is central to determining the goals around which human behavior is organized ; indeed , impairments in affectively driven goal - pursuit may be a critical component of a number of psychiatric disorders , such as depression and schizophrenia ( pessoa , 2008 ) . consequently , the psychological and neurobiological mechanisms by which affective influences modulate cognitive control have become of major interest in recent years and continue to be an important emerging topic of study . much of the experimental research examining how affect modulates cognitive control has involved one of two types of manipulations : emotional manipulations , in which affectively valenced subjective experience is directly induced ( e.g. , through mood inductions or exposure to emotional stimuli ) , or motivational manipulations , where motivational state is altered through the introduction of rewarding or punishing incentives . both types of manipulations are thought to carry affective significance , and both have been hypothesized to impact goal - pursuit and/or cognitive control . however , for the most part , these bodies of research have been carried out independently of one another . in a recent review , pessoa aimed to ameliorate this situation by considering examples of both an emotional manipulation ( threat ) and a motivational manipulation ( reward ) on cognition within a common conceptual framework ( pessoa , 2009 ) . his review suggests that both threat and reward operate in highly similar ways , impacting cognitive performance at both perceptual and executive stages of information processing . however , pessoa acknowledges that emotion and motivation are broad constructs , the impacts of which may not be comprehensively characterized by the phenomena of threat and reward alone . in contrast , and as described further below , other theories of emotion and motivation suggest the possibility of dissociations between the two constructs ( e.g. , liking versus wanting ; berridge , 1996 ) but this remains an understudied issue . thus , the goal of the present paper is to discuss more explicitly existing theoretical accounts regarding the relationship of emotion and motivation to cognitive control , examine how they may relate to one another , and speculate on commonalities and differences in the mechanisms by which they operate . we also suggest future research directions that could be pursued to clarify ambiguity regarding the emotion versus motivation distinction . emotion and motivation are highly related constructs within the domain of affect ( rolls , 2000 ; lang and bradley , 2008 ) , but their influences on cognition generally have not been explicitly considered in relation to one another . when examining the literature regarding the impact of each on cognitive performance , it is important to provide working definitions of relevant terms , so as to begin more carefully examining how these constructs may relate to one another . one review suggests that emotions are best functionally defined as psychological or physiological states that index occurrences of value ( dolan , 2002 ) . as this description suggests , emotion is generally conceptualized as a construct that can be decomposed into multiple subcomponents defining the relation between individual and environment . davidson et al . ( 1990 ) suggest that emotions are comprised of three elements : autonomic reactions , cognitions , and behaviors . more recently , roseman ( 2008 ) asserts that emotion can be thought of as a syndrome of phenomenology ( thought and feeling qualities ) , physiology ( neural , chemical , and other physical responses in the brain and body ) , expressions ( signs of emotion state ) , behaviors ( action tendencies or readinesses ) , and emotivations ( characteristic goals that people want to attain when the emotion is experienced ) . gendron and barrett ( 2009 ) similarly claim that emotions are comprised of subprocesses , including an affective and cognitive ( e.g. , situational construal ) component , and are highly contextualized in nature . common to all of these definitions is the idea that emotions are an affective experience that can be characterized by physiological changes and defined by a cognitive construal of some kind . as states indexing occurrences of value , emotions have been proposed to carry functional value in physiologically preparing the body for action , permitting flexibility of behavioral responses to reinforcing stimuli , facilitating communication and social bonding , and influencing cognitive processes including evaluation , memory encoding , and memory recall ( rolls , 2000 ) . motivations are similar to emotions in that they also serve to define the relation between the individual and the environment ( roseman , 2008 ) , but differ from emotions in being more tightly linked to action and explicit goal associations ; motivated action can be thought of as behavior that is at least partly determined by a desired and hedonically laden end - state ( i.e. , it is goal - directed ) . pessoa ( 2009 ) suggests that motivation can be commonly defined as what makes one work to obtain reward or to avoid punishment . similarly , roseman ( 2008 ) proposes that a motivation is an internal state producing behavior which moves the individual toward desirable reference values or away from undesirable reference values . carver suggests a useful distinction between the two constructs as they relate to goals : while motivation may be the drive toward goal fulfillment , emotion may be emergent from one s sensed rate of progress toward goals ; the difference between one s present status and one s goal state is experienced as affect and may lead to goal reprioritization in order to maximize goal fulfillment ( carver , 2006 ) . similarly , rolls ( 2000 ) suggests that emotions are states elicited by rewarding and punishing reinforcers of behavior . likewise , lang and bradley ( 2008 ) claim appetitive and defense - related brain circuits have evolved to cope with motivationally significant stimuli in the environment ; positive and negative emotion , respectively , are associated with the experience of these brain circuits being activated . thus , according to this general view , emotion can be considered an emergent property of motivationally driven neural activity . however , the lang and bradley view also suggests that emotion is highly characterized by hedonic experience , which accordingly is also tied to the activation of motivational neural circuitry . which motivational system is activated by a stimulus event ( i.e. , appetitive or defensive ) buck ( 2000 ) and laming ( 2000 ) , commenting on d : rolls:2000 ] review of emotion and motivation , argue that emotion and motivation can not be considered separately of one another : laming ( 2000 ) argues that emotion is the subjective experience of being motivated , thus there is no separation between the two ; buck argues that motivation and emotion can not be distinguished from one another if emotion is , as he describes , the manifestation or ( buck , 2000 ) . a contrasting perspective can be drawn from the work of kent berridge , which has highlighted the potential dissociation between activation of motivational circuitry and the neural systems that code for hedonic experience ( berridge , 1996 , 2003 ; berridge and robinson , 1998 , 2003 ; berridge et al . , 2009 ) . he proposes that the hedonic ( i.e. , subjective experiences of pleasure / displeasure ) and motivational ( i.e. , attribution of incentive salience ) facets of reward , shorthanded as liking and wanting respectively , are neurobiologically dissociable : evidence from rodents indicates that hedonic activation may depend on opioid - related circuitry while attribution of incentive salience may depend on the mesolimbic and neostriatal dopamine ( da ) systems . this work suggests that the constructs of emotion and motivation might involve separable neural mechanisms , and as such may have distinct influences on cognitive processing . however , roseman ( 2008 ) has recently suggested key differences : while both may lead to goal - directed action , he proposes several differences between emotion and motivation . roseman argues that motivations are specific , relatively deliberate , and associated with a specific goal . in contrast , emotions are produced by multiple contingencies , are somewhat more impulsive , and are not tightly linked to a particular goal . additionally , he suggests that emotions typically take precedence over motivations : specifically , by engendering emotivations , emotion - specific motivations , that take precedence over non - emotional motivations . these emotivations could potentially be understood ( in terms of carver s conceptualization of emotion ) as a manifestation of goal reprioritization resulting from emotion as an indicator of motivational status . from these working definitions and theoretical accounts of the relationship between emotion and motivation , we suggest that an emotion may be presently considered a construct of multiple processes that together serve to provide an index of value associated with an internal or externally experienced state . while a motivation may be similarly comprised of multiple components , a motivation should be considered a state that produces behavior specifically oriented to carry out a goal that has hedonic value . thus , whereas an emotion may emerge from one s status relative to motivational goals , it may not necessarily be directly relevant to a particular goal . examining the influences of emotion and motivation on cognitive performance may be fundamental to clarifying the relation between these constructs ; currently , however , these investigations have been conducted largely in parallel . the goal of this paper is to integrate these literatures by highlighting some key theoretical accounts of emotional and motivational influences on cognition , and illustrating where empirical evidence suggests these influences may diverge . to facilitate comparison between the emotional and motivational literatures , we have chosen to restrict the focus to studies involving positive emotions and reward incentives . it is , of course , also crucial to explore the relationship between negative emotions and punishment / avoidance - based motivational states , but as positive and negative emotion may be independent of one another ( watson et al . , 1988 ) , the extent to which they share the same mechanisms on cognition remains unclear . gray and braver ( 2002 ) postulate that investigations of emotional influences on cognitive control should fulfill two global aims . the first is to determine whether emotional influences can and do have a selective influence on cognitive control : this must be established first and independently of the nature of these influences . the second aim is to elucidate the mechanisms by which such influences operate . in the present section , we discuss psychological theories regarding the adaptive value of positive emotion on cognition as well as theories regarding the specific mechanisms by which positive emotion takes its effect . it has been suggested that positive emotion might be an adaptive signal indicating safety and security in the environment , giving the organism the freedom to explore and engage in new opportunities ( fredrickson , 2004 ) . building on this postulation , several psychological theories have suggested that positive affect serves to broaden cognition , promote creative problem - solving , and improve cognitive flexibility . foundational work in this area was conducted by isen and daubman ( 1984 ) , who observed that positive affect induction led to broader categorization and facilitated creative problem - solving ( isen et al . , 1987 ) . relatedly , fredrickson ( 2004 ) proposed the broaden - and - build theory of positive emotions , which posits that positive emotions broaden one s repertoire of thought and action , promoting building of intellectual , social , and psychological resources . in the cognitive domain , empirical support for the broaden - and - build theory has come from visual processing and semantic association tasks suggesting a broader scope of attention ( fredrickson and branigan , 2005 ) . dovetailing nicely with this work is carver 's ( 2003 ) coasting hypothesis , which suggests that the security of positive affect might emerge from one s sensed rate of progress toward goals , and result from goal completion at a faster rate than anticipated . under such circumstances , one is free to coast on the goal in question and consider new ideas and/or the pursuit of other goals ( leading to changes in goal prioritization ) . one influential theory , the dopaminergic theory of positive affect ( ashby et al . , ( 1999 ) was developed to address findings that positive emotion is linked to broadened cognition . ashby and colleagues extrapolated from the literature on the neural substrates of reward processing to propose that the psychological effects of positive emotion are specifically linked to increased dopamine ( da ) release ( via the substantia nigra and ventral tegmental area ) in these states . the particular cognitive effects of increased da release during positive affect were postulated to occur through mesocorticolimbic system projections to the anterior cingulate cortex ( acc ) and substantia nigra projections to striatum , with increased da facilitating the ability of acc and striatum to initiate a switch among active task sets , rules , or goal representations maintained in lateral prefrontal cortex ( pfc ) . this facilitation of switching among task - set representations under positive affect enables unusual or non - dominant sets to become active with a greater probability than under neutral affect conditions , which then facilitates creative problem - solving . in connectionist simulations , the account was tested and exhibited an ability to account for certain behavioral performance patterns observed by isen and colleagues under positive affect manipulations ( i.e. , improved performance on creative problem - solving and semantic association tasks ; ashby et al . , 1999 , 2002 ) . dreisbach and goschke ( 2004 ) , dreisbach ( 2006 ) , and muller et al . ( 2007 ) developed a related theoretical framework , which emphasizes that the cognitive flexibility associated with positive affect may have systematic costs in addition to the benefits posited by broadening theories . specifically , dreisbach proposed that changes in dopamine activity triggered by positive affect lead to a shift in the balance between cognitive stability and cognitive flexibility , by increasing the tendency to update to new task goal representations and decreasing the tendency to perseverate in maintaining old ones . empirical evidence from performance in set - shifting and context maintenance paradigms was consistent with this hypothesis , demonstrating positive affect induced facilitation of performance under conditions that depended on flexibility , but impairment under conditions stressing maintenance ( dreisbach and goschke , 2004 ; dreisbach , 2006 ) . a separate theoretical account , put forward by gray ( 2001 ) and gray and braver ( 2002 ) , argued for a hemispherically specialized basis of interactions between positive affect and cognitive control . in this account , an important congruence is noted between prior affective research associating positive emotions with increased activity in the left frontal cortex ( davidson et al . , 1990 ; davidson , 1992 ; davidson and irwin , 1999 ; canli et al . , 2001 ) , and cognitive research linking the left frontal cortex to the active maintenance of verbal information in working memory ( desposito et al . , 1998 ; the primary theoretical claim of the account is that different affective states ( positive versus negative ) should trigger associated behavioral goals in working memory , and that it is adaptive for these goals to be hemispherically segregated such that they can be selectively prioritized by the appropriate affective state . thus , the theory postulates that positive affect states should selectively facilitate verbal working memory a hypothesis that was confirmed experimentally ( gray , 2001 ) . nevertheless , this account is agnostic about why positive and negative affect would be selectively linked with particular stimulus modalities in working memory . in summary , theoretical accounts of the effect of positive emotion on cognition have tended to emphasize influences on cognitive flexibility , potentially by enhancing updating of goal information in working memory . although most work has emphasized the adaptive value of such influences , it has also been suggested to come at a cost to goal maintenance . at the level of neural mechanisms , the focus has been on the dopamine system and pfc , which , as is discussed next , has strong parallels to theoretical accounts regarding how motivation might modulate cognitive control . theoretical accounts of motivation suggest a strong linkage to cognitive control ( simon , 1967 ; carver and scheier , 1998 ; kruglanski et al . , 2002 ) . although early motivational theories suggested a general drive or energization function for motivation ( miller , 1951 ; hull , 1952 ; duffy , 1962 ) , information processing accounts of higher - level cognition have instead emphasized that motivational signals may play a more focused role in the prioritization , updating , and termination of goal representations that provide hierarchical control of behavior ( simon , 1967 ) . over the last 20 years , the intrinsic relationship between motivation and goals has been a central focus of researchers primarily working within the social and individual differences tradition , based on the central claim that motivations are expressed primarily as the activation and representation of specific cognitive and behavioral goals over others ( kruglanski et al . , more recent work has been geared toward demonstrating that goal - directed behavior can be primed and biased by implicit and/or subliminal motivational cues , suggesting a relatively direct route for motivation cognition interactions ( bargh et al . , 2001 ; custers and aarts , 2010 ) . ( 2008b ) have suggested an affective / motivational account of goal - pursuit in which positive motivational signals strengthen goal activation and maintenance , even when this occurs outside of conscious awareness . more recently , psychological theories postulating the role of motivation in activating goals and guiding behavior have begun to be bridged with neuroscience - based studies to more clearly specify the mechanisms by which motivation might influence cognitive control . a primary focus of neuroscience studies on motivation and cognitive control has been to demonstrate that these two processes are integrated within specific brain regions , such as the lateral pfc . early work involving single - unit recording in primates demonstrated that task - related neuronal activity in pfc was modulated by the expected reward value associated with performance ( watanabe , 1996 ; leon and shadlen , 1999 ; watanabe et al . , 2002 ) . in one compelling demonstration , it was found that reward value directly enhanced the fidelity of active maintenance in working memory ( leon and shadlen , 1999 ) . more recent fmri studies carried out in humans have used designs that orthogonally manipulate cognitive control demand and motivational value across a range of task domains , including working memory ( pochon et al . , 2002 ; 2004 ) , context processing ( locke and braver , 2008 ; kouneiher et al . , 2009 ) , task - switching ( savine and braver , 2010 ) , and selective attention ( padmala and pessoa , 2011 ) . these studies have confirmed the presence of specific regions within lateral pfc ( along with effects in other associated regions , such as the acc ) that are sensitive to the interaction of the two factors , consistent with a specific role in integrating motivational and cognitive control functions . the da system also plays a central role in accounts of both motivation and cognitive control . dopamine has long been thought to be a critical component of motivation and reward processing ( wise and rompre , 1989 ; mirenowicz and schultz , 1996 ; robbins and everitt , 1996 ; schultz , 1998 ) . more recent accounts have suggested that da shows phasic , cue - triggered responses to specific events that indicate reward availability ( montague et al . , 1996 ; schultz et al . , 1997 ) and/or high motivational salience ( berridge , 2007 ) . this signal , particularly when a reward is different from anticipated ( i.e. , prediction error ) , may serve as a mechanism for reward - based associative learning ( schultz et al . , 1997 ; schultz , 2002 ; arias - carrion and poppel , 2007 ) . while the role of dopamine as a learning versus salience signal in reward has been debated ( berridge , 2007 ) , both kinds of accounts are compatible with the idea of phasic da involvement in processing motivational incentives and thus consistent with our account . additionally , a separate theoretical account has emphasized that the motivational utility of the current environmental context might be reflected in tonic , rather than phasic , da activation ( niv et al . , 2007 ) . together , these accounts suggest da activity will be increased both by transient cues and sustained contexts that indicate high reward or motivational value . it is worth noting a completely separate literature focused on the influence of da release within pfc , which suggests that the da system provides modulatory role on cognitive control functions . neurophysiological studies in primates show that application of da into pfc sharpens actively maintained stimulus representations ( sawaguchi et al . , 1988 ; sawaguchi and goldman - rakic , 1991 ; arnsten et al . , 1994 ) . in contrast , da antagonists reduce both active maintenance related pfc activity , and also cause behavioral impairments in working memory and cognitive control tasks ( sawaguchi et al . , 1990 ; sawaguchi and goldman - rakic , 1994 ; williams and goldman - rakic , 1995 ) . similar effects have been observed in human pharmacological and fmri studies , with da agonists ( administered systemically ) being associated with improvements in working memory and cognitive control , and leading to associated modulations of pfc activity ( kimberg et al . , 1997 ; gibbs and desposito , 2006 ) . it is striking that the effects of pharmacological manipulations of da in pfc are so similar to the effects that have been observed from motivational manipulations ( leon and shadlen , 1999 ; watanabe et al . , 2002 ) . these linkages between the role of da and pfc in motivation , and the effects of da modulation on pfc - mediated cognitive control functions have prompted the development of theories that explicitly link these two mechanisms . the gating model account , put forward by braver and cohen ( 2000 ) , emphasized the importance of phasic da activity within pfc for the updating and active maintenance of goal representations . specifically , this account suggests that the phasic da responses to cues signaling reward prediction could also be exploited as a means of learning which task - related information should be actively maintained in pfc , and when to update such information . simulation studies demonstrated that a system could in fact learn appropriate updating and maintenance of task context or goal information based on reward prediction cues . thus , this account suggests the possibility of a linkage between the reward / motivational and cognitive control functions mediated by da this point was made even more explicitly in the recent dual mechanisms of control framework ( braver et al . , 2007 ) , which specifically suggests that signals of reward motivation will bias cognitive control toward a proactive mode , in which task cues trigger sustained goal activation and maintenance in the service of preparation for anticipated control demands , via the aforementioned da pfc interaction . proactive control is distinguished from reactive control , in which the same task cues only trigger transient control related processes , rather than sustained active maintenance , under conditions of low da activity , such as when the environmental context is perceived to have reduced motivational value . this account has been supported by recent experimental evidence from an fmri study of motivational influences on task - switching ( savine and braver , 2010 ) . task trials that had high reward value were associated within increased activity in both lateral pfc and the midbrain da system . moreover , the increased cue - related pfc activation on high reward trials was associated with behavioral measures of improved task preparation , supporting the idea of a reward - related shift toward proactive control . in summary , investigations of motivation and cognitive control have primarily focused on the role of goal representations in psychological models , and on interactions of the da system in pfc in neuroscience studies . one mechanistic account suggests that da activity triggered by reward cues can serve as an updating and prioritization signal , that modulates active maintenance of goal information within pfc . the potential similarity of the proposed mechanisms underlying both positive affect and reward motivation influences on cognitive control , according to the ashby model ( i.e. , cognitive changes due to increased dopamine release to cortical areas such as the pfc and acc ) , is striking , but has not yet been directly confirmed by experimental studies . emotion and motivation are closely related constructs , but it is still not clear how to relate their influences on cognitive control . some theories do not clearly distinguish between the two , while others more explicitly state that emotion and motivation , while highly related , remain distinct constructs . given the state of current evidence , this is still a question calling for clarification . most proposals regarding affective influences on cognitive control have not drawn a clean distinction between emotion and motivation . this includes gray 's ( 2001 ) hemispheric specialization hypothesis , which explicitly does not distinguish between positive / negative emotion and approach / avoidance motivation , and ashby et al . 's ( 1999 ) dopaminergic theory of positive affect , which posits a relation between da and positive emotion because of da s involvement in reward processing . aarts et al . ( 2008b ) go even further , by explicitly suggesting an equivalence between positive emotion and reward motivation in terms of their effects on goal - pursuit and executive control . in their model , positive affective signals occurring in temporal proximity to activation of a cognitive goal should strengthen the maintenance , accessibility , and pursuit of that goal , regardless of whether those affective signals relate to goal attainment ( i.e. , whether or not they serve as direct reward motivation cues ) . despite these claims , the relationship between positive affect , approach motivation , and da system activity is somewhat tenuous . experimental studies have shown that affective valence ( positive / negative ) and motivational direction ( approach / avoidance ) can be dissociated ( harmon - jones , 2003 ; harmon - jones and gable , 2009 ) . likewise , in carver 's ( 2006 ) theoretical model , positive and negative affect emerge from detection of the rate of progress toward fulfilling either approach or avoidance - related goals : positive emotions result from above - anticipated goal progress ; negative emotions result from below - anticipated goal progress . finally , in berridge s neuroscience - based account , dopamine is only critical for the transfer of motivational salience , or stimulus - triggered wanting to new cues or events , and does not generate hedonic experience , or liking ( which is thought to be represented in the ventral pallidum via neurochemicals such as opiates and endocannabinoids ; berridge , 2007 ; berridge et al . , 2009 ) . thus , without supportive direct evidence , links between positive affect , approach motivation , and da system activity should not be assumed . deconfounding emotional and motivational influences from one another is a challenge for exploring and clarifying these relationships . we argue that , as a starting point , emotional and motivational manipulations need to be operationalized ( i.e. , with exposure to emotional stimuli during or just prior to the task , or motivational performance - contingent incentives , respectively ) and examined with performance - independent measures . examining psychophysiological signatures of these influences [ e.g. , startle reflex , facial electromyography ( emg ) ] that have been linked to valence but not motivational orientation of affective experiences ( cacioppo et al . , 1986 ; lang et al . , 1990 ) may be useful in dissociating these influences from one another . although these influences have yet to be directly disentangled , independent empirical work from the emotion and motivation literatures indirectly hints that these are dissociable influences . while several theories have proposed that positive emotion can lead to cognitive broadening and flexibility ( isen et al . , 1987 ; ashby et al . , 1999 ; fredrickson , 2004 ; rowe et al . , 2007 ) and exploration of alternate goals ( carver , 2003 ) , growing evidence suggests that reward incentives enhance goal maintenance / representation and influence proactive cognitive control ( savine and braver , 2010 ; padmala and pessoa , 2011 ) . empirical evidence is mixed , suggesting that positive emotion and reward may have similar effects on some cognitive processes , such as task - switching ( reducing switch costs ; yan - mei and de - jun , 2008 ; savine et al . , 2010 ) , opposite effects on other processes , such as selective attention ( rowe et al . , 2007 ; padmala and pessoa , 2011 ) and mixed results on yet other processes such as goal maintenance ( dreisbach and goschke , 2004 ; dreisbach , 2006 ; aarts et al . how these diverging effects relate has yet to be systematically clarified , since positive affect and reward motivation have not yet been directly examined and compared . nevertheless , there is at least one example from the literature in which the same experimental paradigm the ax continuous performance task ( ax - cpt ; cohen and servan - schreiber , 1992 ; servan - schreiber et al . , 1996 ; braver et al . , 2001 ) has been utilized ( in separate studies ) to explore the effects of positive affect and reward motivation on cognitive control . the ax - cpt is a potentially advantageous paradigm for this purpose , because it permits selective examination of goal maintenance capability and proactive control . however , surprisingly , the results of the positive affect and reward motivation studies appear to be somewhat different . under positive affect induction via valenced pictures displayed prior to each trial , participants showed evidence of reduced maintenance capability relative to neutral affect ( dreisbach , 2006 ) . conversely , under manipulations of reward motivation , participants showed evidence of enhanced maintenance capability relative to baseline ( locke and braver , 2008 ) . in both studies , the results were interpreted as arising from modulation of da activity in pfc , but it is not clear if the two different manipulations actually led to similar or distinct effects within these brain systems . thus , direct comparisons are clearly needed , with monitoring of neural activity , to determine whether there is a potential dissociation between positive affect and reward motivation effects on goal maintenance . one possibility that is worth considering is that da activity in pfc underlies both positive affect - related reduction in maintenance capability and reward - related increases in maintenance capability , but that the two effects reflect dissociable temporal dynamics of da influence . in particular , it is well - accepted that da activity should be considered in terms of both tonic and phasic components , which can interact with each other ( grace , 1991 ) . further , prior research has more strongly linked facilitation of goal maintenance with tonic da activation in pfc , primarily via d1 receptors ; in contrast , phasic da activity in pfc may promote updating and cognitive flexibility , potentially via d2 receptors ( cohen et al . , 2002 ; oreilly and frank , 2006 ; durstewitz and seamans , 2008 ) . thus , distinctions between reward motivation and positive affect may reflect a distinction in the balance between tonic versus phasic da activation and/or a d1 versus d2 dominated state . however , the relationship between emotional and motivational processes and differential temporal dynamics of da activity has not been directly shown and has yet to be investigated . the constructs of emotion and motivation are closely interrelated , and typically their influences on cognition have not been explicitly separated from one another ( e.g. , pessoa , 2009 ) . however , in the light of accumulating evidence suggesting they may be dissociable , their influences on cognition must be clarified . as alluded to above , there are unresolved questions in this domain that provide promising routes for future investigation . here a first , straightforward suggestion is for studies to be conducted that directly test and compare the effects of positive affect manipulations with those involving reward motivation within a single sample , and using closely matched experimental designs . of course , this suggestion does beg the question of what exactly differentiates a positive affect manipulation from one involving reward motivation , especially since some studies advertised as examining positive affect have actually used manipulations involving delivery of rewards ( isen et al . although this question is one that may involve a deeper discussion that is outside of the scope of this article , we want to highlight that this issue has not really been adequately considered by researchers working in this area . nevertheless , it is at least possible to operationalize a distinction in which positive affect is manipulated by influences such as mood inductions or strongly valenced stimuli , whereas reward motivation is manipulated by varying the incentives provided for task - performance . with such an approach , it would be possible to directly test the hypothesis that positive emotion promotes cognitive flexibility while reward incentives promote greater goal maintenance . as mentioned previously , cognitive paradigms that specifically probe active maintenance and proactive control , such as the ax - cpt , could be used with emotional and motivational experimental manipulations to probe the possibility of dissociable versus common behavioral effects and/or their associated physiological bases . other paradigms should also be examined , such as the remote associates and global / local paradigms , that have been frequently and fruitfully examined to demonstrate increased cognitive flexibility under positive affect ( isen and daubman , 1984 ; fredrickson and branigan , 2005 ; rowe et al . , 2007 ) , but have not previously been studied with respect to reward motivation . finally , the use of a common paradigm would be useful for exploring other hypotheses relating to positive affect and/or reward motivation such as carver s coasting model . unfortunately , this model has not previously been studied within cognitive experimental paradigms , so further work in how to operationalize maintained goal - pursuit versus goal - switching would need to occur ( e.g. , via exploration of potentially approach paradigms such as volitional task - switching ) . a second promising direction for future research is a more direct examination of the role of neurotransmitter systems in mediating affective and motivational influences on cognitive control . this is especially important with regards to the effects of positive emotion on cognition , since theories such as ashby s posit a neurotransmitter mechanism without direct evidence for its involvement . study of the role of neurotransmitter systems in positive affect is somewhat challenging , since detecting positive affect in animal models is difficult ( although potentially not impossible , e.g. , berridge , 2000 ) , as is direct monitoring of neurotransmitter activity in humans . however , studies that examine state - related changes in neurotransmitter receptor binding ( e.g. , using pet radioligand - labeling methods and dynamic imaging approaches ) do provide a promising avenue for this research , especially for the monitoring of da system activation ( egerton et al . , 2009 ) . in particular , receptor binding studies may provide a powerful means of directly testing whether increases in positive affect are associated with increases in da system activity . pharmacological manipulations provide another method by which to examine the link between neurotransmitter activity and cognitive change . pharmacological challenge studies involving agents that influence da activity have been used fruitfully in a range of domains to understand the role of this system in cognitive functions such as working memory ( e.g. , cools et al . , 2007 ) , as well as in mediating basic aspects of reward processing ( e.g. , pessiglione et al . , 2006 ) . thus , targeted studies are needed that directly investigate how pharmacological manipulation of this system impacts motivational versus affective effects on cognitive control . although dopamine has received much attention for its involvement in reward and cognitive control processes , other neurotransmitters such as norepinephrine ( ne ) may also play crucial roles . ne activity , historically linked to physiological arousal ( foote et al . , 1980 ) , is becoming increasingly appreciated as a complex and specific mediator in the control of behavior as well ( aston - jones and cohen , 2005 ) . the locus coeruleus ( lc)-ne system may play a critical role in the regulation of exploitation ( i.e. , optimizing current task - performance ) and exploration ( i.e. , disengagement in search of alternative behaviors ) tendencies during task - oriented behavior phasic and tonic ne activity may respectively reflect these control tendencies ; aston - jones and cohen , 2005 ) . it may be that exploration is highly analogous to carver 's ( 2003 ) idea of coasting or that the exploitation exploration balance is analogous to the maintenance - flexibility balance discussed with regard to dopamine and control ( dreisbach and goschke , 2004 ; muller et al . , 2007 ) , but connections between these bodies of work have yet to be made . in particular , it is still not well understood what the relative roles of dopamine and norepinephrine are in emotional and motivational interactions with cognition . exciting new evidence suggests that pupil dilation can be used as a non - invasive marker of lc - ne activity in humans , with pupil dilation changes indexing shifts in the exploitation exploration balance during performance of cognitive control tasks ( gilzenrat et al . , 2010 ) . thus , pupillometric methods might be one promising avenue for exploring the role of the lc - ne system in affective versus motivational influences in cognitive control , along with the psychopharmacological and neurotransmitter imaging methods described above . more generally , the use of psychophysiological measures may provide another approach by which possible dissociations between emotional and motivational influences on cognition can be investigated . pupil dilation has been well - established to index fairly specific changes in cognitive demand and effort ( beatty , 1982 ; granholm et al . , 1996 ; beatty and lucero - wagoner , 2000 ) and thus may provide a measure of cognitive control , independent from behavior , that can finely index changes in temporal control dynamics . for example , pupil dilation has been successfully utilized to examine the temporal dynamics of goal maintenance and proactive versus reactive control within the ax - cpt paradigm ( related to developmental changes ; e.g. , chatham et al . , 2009 ) . additionally , pupil dilation is sensitive to emotional variables as well , and may reflect emotional arousal ( bradley et al . , 2008 ) . thus , pupillometric methods might be exceptionally well - suited for examining interactions of affective / motivational systems and cognitive control , although such interactions have almost never been explored ( e.g. , satterthwaite et al . , 2007 ) . skin conductance ( scr ) , startle probes , and emg are all well - established autonomic indicators of affective state ( bradley et al . , 2001 ) . thus , comparison of the effects of affective versus motivational manipulations on these indicators might help to reveal potential dissimilarities in autonomic profile , as well as whether some indicators serve as better predictors of the cognitive control effects of such manipulations . another means by which to probe for distinctions between emotion and motivation might be to more clearly assess the role of subjective experience . subjective experience is thought to be a core ingredient to the construct of emotion ( barrett et al . , 2007 ) , but it remains unclear whether it should be considered key to the construct of motivation as well . recent evidence suggests that subliminally presented reward cues can lead to similar effects on behavior and cognitive control as stimuli that are consciously perceived ( aarts et al . 2010 ) , suggesting that subjective awareness of motivational value is not critical . in the emotion realm , subliminal information has been shown to influence affective preference ( zajonc , 1980 ) , and non - emotional facial changes ( i.e. , to a smile - like position ) have been associated with changes in emotional response ( strack et al . , 1988 ) . however , subliminal presentation of positively valenced stimuli seems to only impact motivated behaviors such as the amount consumed and willingness - to - pay for rewards , but not subjectively experienced affective responses such as positive mood or reward liking ( winkielman et al . , 2005 ) . thus , it is not clear whether subliminal manipulations of positive affect would impact cognitive control in the same manner as manipulations that are subjectively experienced . as a final consideration , the relationship between emotion and motivation could be explored by examining the effect these manipulations have on each other . some evidence suggests that reward incentives promote positive mood ( meloy et al . , 2006 ) , but such research is relatively sparse and , to our knowledge , effects of emotion on the effectiveness of a motivational manipulation remains unknown . while emotion and motivation have been assumed to be related , they have largely been investigated independently with relation to cognitive control . current theories suggest that the influences of positive emotion and reward incentives may depend on a common neuroanatomy : both may increase midbrain dopaminergic activity ( i.e. , ventral tegmental area ) , which projects to control - associated areas , such as the pfc and the acc . the pfc in particular has received attention as a potential integration site for task and affective information in both emotional and motivational lines of research . despite these parallels , perhaps most fundamentally , the neurobiological mechanism , independent of reward , by which positive emotion influences cognition remain unclear . specifically , there is as of yet no direct evidence that supports a model in which positive emotion , independent of other components of reward , is associated with dopamine activity . clarifying this mechanism will be key to establishing whether positive emotion and reward motivation influences on cognitive control are common or distinct . second , further exploration of differential contributions of tonic and phasic activity in both the da system and other relevant neurotransmitter systems , such as lc - ne , could help characterize both emotional and motivational influences on cognition , especially in probing further the hypothesis that positive affect and reward might be associated with more reactive and proactive ( and/or exploratory versus exploitative ) modes of cognitive processing , respectively . third , inclusion of both psychophysiological measures and assessment / manipulation of subjective experience may be beneficial in theoretically distinguishing constructs of emotion from motivation and improving understanding of how they impact cognition . fourth , effects of emotion and motivation on one another should be probed to help clarify their relationship . emotions may be characterized as emergent from one s current status relative to one s optimal goal status ( carver , 2006 ; lang and bradley , 2008 ) . thus , while emotion and motivation are closely related , emotions as an evaluator of goal status serve a slightly different functional purpose than motivations , and consequently may be more fluid in relation to any given goal . accordingly , they may have a more variable influence on cognitive control relative to motivational influences , which serve to optimize goal - relevant processing directly . clarifying these constructs will help unite disparate lines of research within a common theoretical framework , and provide a more nuanced picture of these interactions and their complexities . by doing so , this research effort may help to significantly advance the emerging field of affect cognition interactions . the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest . Output:	it is becoming increasingly appreciated that affective influences can contribute strongly to goal - oriented cognition and behavior . however , much work is still needed to properly characterize these influences and the mechanisms by which they contribute to cognitive processing . an important question concerns the nature of emotional manipulations ( i.e. , direct induction of affectively valenced subjective experience ) versus motivational manipulations ( e.g. , delivery of performance - contingent rewards and punishments ) and their impact on cognitive control . empirical evidence suggests that both kinds of manipulations can influence cognitive control in a systematic fashion , but investigations of both have largely been conducted independently of one another . likewise , some theoretical accounts suggest that emotion and motivation may modulate cognitive control via common neural mechanisms , while others suggest the possibility of dissociable influences . here , we provide an analysis and synthesis of these various accounts , suggesting potentially fruitful new research directions to test competing hypotheses .
PubmedSumm2035	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: glucocorticosteroids have been used in an attempt to minimize or prevent post - operative edema after oral and maxillofacial surgery . dexamethasone seems to be the most suitable because it has the highest anti - inflammatory activity , no mineralocorticoid activity and a longest available half - life of 36 - 54 h. psychosis is a generic psychiatric term for a mental state often described as involving a loss of contact with reality . people experiencing psychosis may report hallucinations or delusional beliefs and they may exhibit personality changes and disorganized thinking . we report three maxillofacial cases who experienced post - operative psychosis after peri - operative dexamethasone therapy , as a result of which , the patients had a prolonged stay in the surgical intensive care unit . - operatively , as per protocol , she received intravenous ( iv ) dexamethasone 16 mg on the day of surgery at 6 am . surgery lasted for 4 h. after extubation , she was transferred to the post - operative care unit ( pacu ) awake and calm . on arriving at pacu , she was given another dose of dexamethasone , 8 mg . iv midazolam , propofol and iv fentanyl were administered for sedation , but the patient remained agitated and confused . over the next 3 h , iv fentanyl 100 g , propofol 40 iv midazolam 2 mg and intramuscular ( i m ) haloperidol 4 mg were given . an iv dexmedetomidine infusion at 0.4 microgm / kg / min tapered to 0.2 microgm / kg / min , provided sedation for 6 h , after which it was stopped . as the patient continued to be calm over the next 12 h , she was transferred to the ward . she continued to be stable and was discharged on the post - operative day 5 . a 58-year - old , female patient , presented for surgery for wide local excision of squamous cell carcinoma of maxilla and closure with split skin graft . pre - operatively , as per protocol , she received iv dexamethasone 16 mg on the day of surgery at 6 am . surgery lasted for 2 h. after extubation , she was awake and calm . on arriving at pacu , another dose of 8 mg of dexamethasone was administered . an iv dexmedetomedine infusion providing sedation was administered for 3 h at 0.4 g / kg / h . the dexmedetomidine infusion was restarted for next 6 h , initially at 0.4 g / kg / h tapered to 0.2 g / kg / h and then stopped . as the patient remained calm over the next 12 h , she was transferred to the ward . she continued to be psychologically stable and was discharged on the post - operative day 6 . an 80-year - old , female patient , weighing 35 kg presented with biopsy proven squamous cell carcinoma of the left side of the mandible . she was taken up for wide excision with segmental mandibulectomy , modified radical neck dissection and reconstruction with pectoralis major myocutaneous flap . pre - operatively , as per protocol , she received iv dexamethasone 8 mg on the day of surgery at 6 am . surgery lasted for 4 h. she was electively placed on endotracheal tube ( ett ) with t - piece in the post - operative period in view of extensive surgery . in the pacu , another dose of 8 mg was administered . fourteen hours after surgery , she became restless and she pulled out her ett and ryle 's tube . after the surgery , she was placed on dexmedetomidine infusion for 12 h , initially at 0.3 /kg / h and then tapered to 0.1/kg / h ( the infusion was stopped intermittently whenever the patient developed bradycardia or hypotension and whenever the patient remained calm ) . iv midazolam 1 mg and i m haloperidol 2.5 mg were continued 8 hourly for 2 days , after which the rest of post - operative period was uneventful . she was extubated 16 h after the surgery and then shifted back to the ward . she continued to be psychologically stable and was discharged on the post - operative day 8 . - operatively , as per protocol , she received intravenous ( iv ) dexamethasone 16 mg on the day of surgery at 6 am . surgery lasted for 4 h. after extubation , she was transferred to the post - operative care unit ( pacu ) awake and calm . on arriving at pacu , she was given another dose of dexamethasone , 8 mg . iv midazolam , propofol and iv fentanyl were administered for sedation , but the patient remained agitated and confused . over the next 3 h , iv fentanyl 100 g , propofol 40 iv midazolam 2 mg and intramuscular ( i m ) haloperidol 4 mg were given . an iv dexmedetomidine infusion at 0.4 microgm / kg / min tapered to 0.2 microgm / kg / min , provided sedation for 6 h , after which it was stopped . as the patient continued to be calm over the next 12 h , she was transferred to the ward . she continued to be stable and was discharged on the post - operative day 5 . a 58-year - old , female patient , presented for surgery for wide local excision of squamous cell carcinoma of maxilla and closure with split skin graft . pre - operatively , as per protocol , she received iv dexamethasone 16 mg on the day of surgery at 6 am . surgery lasted for 2 h. after extubation , she was awake and calm . on arriving at pacu , another dose of 8 mg of dexamethasone was administered . an iv dexmedetomedine infusion providing sedation was administered for 3 h at 0.4 g / kg / h . the dexmedetomidine infusion was restarted for next 6 h , initially at 0.4 g / kg / h tapered to 0.2 g / kg / h and then stopped . as the patient remained calm over the next 12 h , she was transferred to the ward . she continued to be psychologically stable and was discharged on the post - operative day 6 . an 80-year - old , female patient , weighing 35 kg presented with biopsy proven squamous cell carcinoma of the left side of the mandible . she was taken up for wide excision with segmental mandibulectomy , modified radical neck dissection and reconstruction with pectoralis major myocutaneous flap . pre - operatively , as per protocol , she received iv dexamethasone 8 mg on the day of surgery at 6 am . surgery lasted for 4 h. she was electively placed on endotracheal tube ( ett ) with t - piece in the post - operative period in view of extensive surgery . in the pacu , another dose of 8 mg was administered . fourteen hours after surgery , she became restless and she pulled out her ett and ryle 's tube . after the surgery , she was placed on dexmedetomidine infusion for 12 h , initially at 0.3 /kg / h and then tapered to 0.1/kg / h ( the infusion was stopped intermittently whenever the patient developed bradycardia or hypotension and whenever the patient remained calm ) . iv midazolam 1 mg and i m haloperidol 2.5 mg were continued 8 hourly for 2 days , after which the rest of post - operative period was uneventful . she was extubated 16 h after the surgery and then shifted back to the ward . she continued to be psychologically stable and was discharged on the post - operative day 8 . glucocorticosteroids are used extensively for minimizing and preventing post - operative oedema after oral and maxillofacial surgery . dexamethasone seems to be the most suitable because it has the highest anti - inflammatory activity , no mineralocorticoid activity and a longest available half - life of 36 to 54 h. dexamethasone has also been in use in the peri - operative period for post - operative nausea and vomiting pain management and in neurosurgery . even though , efficacious for preventing and minimizing edema in oral and maxillofacial surgeries , there have been reports attributing post - operative psychosis to peri - operative dexamethasone [ table 1 ] . in fact in one of the reports a single dose has been implicated for the development of post - operative psychosis . while the incidence of post - operative delirium is 36.8% , acute steroid related psychiatric disorder frequency is 5% . in view of such a high incidence , it may be imperative that , with every case where peri - operative dexamethasone is administered , a post - operative psychotic reaction should be anticipated . in patients who are being post - operatively ventilated , it may be necessary to use restraints to prevent self - extubation and a pre - operative consent for the same would be required . the episodes of psychosis can prolong the hospital stay or may result in readmission and increase the total cost for the patient . early recognition and once there is a psychotic event , the stay of the patient is prolonged in the intensive care units thereby increasing the costs . the exact dose of dexamethasone that can cause a psychotic reaction is unknown . since it has been reported that a single dose can cause post - operative psychosis [ table 1 ] all patients receiving dexamethasone have to be monitored carefully post - operatively special care has to be taken in patients who have a previous history of psychiatric disorders . awareness and the ability to diagnose post - operative psychosis following use of peri - operative dexamethasone are crucial to provide better care and effective treatment . Output:	a broad array of behavioral symptoms , including psychosis , can transpire post - operatively following a variety of surgeries . it is difficult to diagnose the exact cause of post - operative psychosis . we report three cases , which developed psychosis post - operatively after undergoing major oral and maxillofacial surgeries . all the three patients were administered dexamethasone peri - operatively . dexamethasone is used to prevent or reduce post - operative edema . the exact dose of dexamethasone , which can cause psychosis , is unknown . it is important to raise awareness about this potential complication so that measures for management can be put in place in anticipation of such an event .
PubmedSumm2036	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: haemophilus influenzae type b ( hib ) is responsible for most cases of invasive h. influenzae disease , including meningitis and septicemia , in unvaccinated children . with the decline in the incidence of invasive hib disease as a result of the hib vaccine program , nontypeable h. influenzae ( nthi ) has become an important cause of invasive h. influenzae disease in vaccinated children . nthi is part of the normal flora in the upper respiratory tract of children and commonly causes local respiratory tract diseases , including otitis media , sinusitis , conjunctivitis , and bronchitis [ 2 , 3 ] . meningitis and septicemia have been reported rarely in children with underlying predisposing conditions , including head trauma , cerebrospinal fluid ( csf ) leakage , and immune compromise , in neonates associated with colonization of the maternal genital tract [ 7 , 8 ] , and more rarely in previously healthy children [ 911 ] . according to a japanese nationwide surveillance system , the frequency with which nthi meningitis in children occurs seems to vary , depending on the populations surveyed : 4 ( 0.7% ) of the 535 patients with h. influenzae meningitis were nthi strains ; 2 ( 4.9% ) of the 41 pediatric patients with h. influenzae meningitis were nthi strains . we report a case of a previously healthy 2-year - old girl who developed meningitis and septicemia caused by nthi biotype , and wish to emphasize that nthi must be borne in mind as a potential pathogen that can cause invasive disease , even in previously healthy children . on 7 september 2008 a previously healthy 2-year - old girl was admitted to the hospital with a 12-h history of fever and vomiting . she had no rhinorrhea or cough before her hospital stay . she had been born at 40 weeks of gestation , and her birth weight was 3062 g. subsequent mental and physical development had been normal . she had been immunized with various vaccines as appropriate according to the immunization schedule for children ( bcg , diphtheria , tetanus , pertussis , poliomyelitis , measles , rubella , and varicella ) . she lived with her parents and 3-year - old brother , and attended a nursery school . on admission , she had a body temperature of 39.2c , heart rate of 122 beats / min , respiratory rate of 40/min , and an instant capillary refill time without cold peripheries . laboratory data on admission were : white blood cell ( wbc ) count , 13400/l ( neutrophils 91% , monocytes 2% , lymphocytes 7% ) ; serum aspartate aminotransferase ( alt ) , 29 iu / l ; serum alanine aminotransferase ( ast ) , 17 iu / l ; serum lactate dehydrogenase ( ldh ) , 252 iu / l ; serum c - reactive protein ( crp ) , 1.5 mg / dl . she was initially treated with intravenous fluid and antipyretics , with her condition monitored closely . on the second day of admission , she had a body temperature of 39.3c , heart rate of 145 beats / min , and respiratory rate of 45/min . she became disoriented and the glasgow coma scale showed an eye opening ( e ) score of 4 , verbal response ( v ) score of 2 , and best motor response ( m ) score of 5 . no anterior fontanelle bulging or hemiparesis were observed , and kernig s sign and babinski s sign were negative . the laboratory data on september 8 were : wbc count , 15300/l ( neutrophils 82% , monocytes 9% , lymphocytes 8% ) ; alt , 30 iu / l ; ast , 16 iu / l ; ldh , 307 iu / l ; fasting blood glucose , 144 mg / dl ; crp , 15.0 mg / dl ; csf cell count , 3146 cells/l ( polymorphonuclear leucocytes , 78% ) ; csf protein , 51 mg / dl ; csf glucose , 35 mg / dl . gram staining of the csf revealed the presence of small , gram - negative , pleomorphic rods resembling h. influenzae . however , a csf test kit designed to detect the capsular antigens of bacteria causing meningitis ( pastrorex meningitis ; bio - rad laboratories , tokyo , japan ) gave a negative result for hib antigen . on the basis of the clinical and laboratory findings , diagnosis of bacterial meningitis was made , and empiric treatment with ceftriaxone sodium hydrate ( ctrx ) ( 120 mg / kg / day , 12-hourly ) and meropenem hydrate ( mepm ) ( 140 mg / kg / day , 8-hourly ) was started . to avoid neurological sequelae , dexamethasone ( 0.60 mg / kg / day , 6-hourly ) the isolates were subsequently identified as h. influenzae by 16s rrna gene sequencing , followed by a basic local alignment search tool ( blast ) search in the laboratory of ubukata kimiko at the kitasato institute for life sciences kitasato university . on serotyping , no agglutination was observed with antisera against the capsular serotypes a through f , leading to categorization of the strain as nontypeable . on biotyping , the organism failed to produce indole and to decarboxylate ornithine , but was able to hydrolyze urea , leading to its categorization as biotype . h. influenzae biotype could be distinguished from h. aegyptius by its fermentation of xylose or by its growth on tryptic soy agar containing x and v factors . all of the csf , blood , and throat swab cultures yielded beta - lactamase - negative nthi biotype . the minimum inhibitory concentrations ( mic ) of several antibiotics for the organism are listed in table 1.table 1minimum inhibitory concentration of several antibiotics for nontypeable haemophilus influenzae biotype isolated from the cerebrospinal fluidantibioticmic ( g / ml)ampicillin0.25cefditoren<0.03cefcapene<0.03cefotaxime<0.03panipenem1telithromycin8clarithromycin<0.03meropenem<0.03ceftriaxone<0.03chloramphenicol1levofloxacin<0.5 minimum inhibitory concentration of several antibiotics for nontypeable haemophilus influenzae biotype isolated from the cerebrospinal fluid a repeat csf culture on september 9 was also positive . blood culture was not repeated . on september 12 , , ctrx was stopped when the serum crp had remained within the normal range for 7 days . on september 28 , because of the unusual finding of nthi meningitis , further investigations were performed on the child . the results revealed normal levels of immunoglobulin g ( igg ) , igg subtypes , iga , igm , cd4 , cd8 , and of the blood complement components . cranial and paranasal sinus computed tomography and magnetic resonance imaging of the head were normal . dexa dexamethasone , ctrx ceftriaxone , mepm meropenem , wbc white blood cell , crp c - reactive protein clinical course of a 2-year - old girl with nontypeable haemophilus influenzae meningitis . dexa dexamethasone , ctrx ceftriaxone , mepm meropenem , wbc white blood cell , crp c - reactive protein we report a rare case of meningitis and septicemia caused by nthi biotype . this case demonstrates that nthi should be borne in mind as a potential pathogen that can cause meningitis and septicemia , even in previously healthy children . almost all cases of h. influenzae meningitis in unvaccinated children are caused by hib or biotype strains . in contrast with hib , nthi is primarily recognized as a commensal organism in the upper respiratory tract . however , nthi biotype seems to have the potential to cause invasive disease such as meningitis [ 4 , 9 , 10 ] and septicemia [ 7 , 9 ] . therefore , children presenting with invasive nthi disease should be promptly subjected to further evaluation and diagnostic imaging for identifying predisposing factors such as head trauma , csf leakage , and immune compromise . concerning the possible route of transmission of nthi biotype , it seems likely that the meningitis and septicemia in our patient were caused by the strain isolated from her throat culture , because all strains isolated from the csf , blood , and throat belonged to identical biotypes and the mic distributions were almost identical . a subgroup of nthi strains is spontaneous capsule - deficient progeny of hib strains called b strains . the b strains might be generated by loss of their capsulation ability through homologous recombination . the frequency of b strains in hib is 0.10.3% [ 14 , 18 , 19 ] . the b strains can not be ruled out for our patient s strain , because the slide agglutination test against the capsular serotypes are not able to differentiate between nthi and b strains . since the 1980s , when hib conjugate vaccines were introduced in many foreign countries , the incidence of hib meningitis has decreased dramatically . as a consequence , nthi has become relatively more important as a pathogen for meningitis . in december 2008 , hib conjugate vaccine was introduced in japan . since the introduction of hib conjugate vaccine , investigation of capsular serotypes is necessary in all cases of h. influenzae meningitis , because hib vaccine does not protect against meningitis due to nontypeable strains . we conclude that nthi biotype must be borne in mind as a potential pathogen that can cause meningitis and septicemia , even in previously healthy children . Output:	nontypeable haemophilus influenzae ( nthi ) commonly colonizes the upper respiratory tract of children and causes otitis media , sinusitis , and bronchitis . invasive nthi diseases such as meningitis and septicemia have rarely been reported , especially in children with underlying predisposing conditions such as head trauma and immune compromise . however , we report a previously healthy 2-year - old girl who developed meningitis and septicemia caused by nthi biotype . she was treated with dexamethasone , meropenem , and ceftriaxone , and recovered uneventfully . we wish to emphasize that nthi should be borne in mind as a potential pathogen that can cause meningitis and septicemia , even in previously healthy children .
PubmedSumm2037	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: diabetes mellitus is a major cause of erectile dysfunction ( ed ) , and the prevalence of ed is more common and the symptoms of ed occur earlier in diabetic men than in the general population . the suggested mechanisms for diabetic ed include damage to the cavernous endothelial and smooth muscle cells at both structural and functional levels , peripheral autonomic neuropathy , and endocrine dysfunction , such as hypogonadism [ 3 - 7 ] . it is well known that men with diabetic ed respond poorly to the currently available oral phosphodiesterase-5 ( pde5 ) inhibitors compared with nondiabetic men . the reduced responsiveness to the pde5 inhibitors in men with diabetes may be related to the severity of functional and structural derangements in the corporal vasculature as well as the degree of peripheral neuropathy . nerve injury - induced protein 1 , ninjurin 1 , is a cell surface protein related to peripheral nerve injury that is known to be profoundly up - regulated in the distal segment and proximal neuroma of the transected sciatic nerve . it was also reported that ninjurin 1 is responsible for the initiation and progression of multiple sclerosis , an autoimmune inflammatory disease of the central nervous system ( cns ) characterized by demyelination and axonal damage . moreover , ninjurin 1 plays an important role in blood vessel homeostasis during the embryonic period . a recent study reported that ninjurin 1 resulted in the regression of the hyaloids vascular system , a capillary vascular network responsible for the growth and maturation of the lens during early ocular development , through induction of angiopoietin-2 ( ang2 ) , an endogenous antagonist of angiopoietin-1 ( ang1 ) involved in destabilization of the blood vessels and endothelial cell apoptosis , and of wnt7b , a critical mediator of cell death . inhibition of ninjurin 1 by systemic administration of neutralizing antibody delays regression of the hyaloids vascular system . these findings suggest the critical function of ninjurin 1 in both neuropathy and blood vessel regression . therefore , it is meaningful to examine the expression of ninjurin 1 in diabetic ed , a disease characterized by neuropathy and vasculopathy . thus , in the present study , we determined the differential expression of ninjurin 1 in both penile tissue of streptozotocin ( stz)-induced diabetic mice with ed in vivo and in primary cultured mouse cavernous endothelial cells ( mcecs ) in vitro . specific - pathogen - free c57bl/6j mice were purchased from orient bio ( seongnam , korea ) . the experiments performed were approved by the institutional animal care and use subcommittee of our university . diabetes was induced in 8-week - old c57bl/6j mice by intraperitoneal injections of stz ( 50 mg / kg ) for 5 days consecutively as previously described . at 8 weeks after the induction of diabetes , the diabetic animals and their age - matched controls were used for western blot analysis , histologic examination , and physiologic erection study . fasting and postprandial blood glucose levels were determined with an accu - check blood glucose meter ( roche diagnostics , mannheim , germany ) before the mice were killed . the mice from the control and diabetic groups were anesthetized with ketamine ( 100 mg / kg ) and xylazine ( 5 mg / kg ) intramuscularly . stimulation parameters were 5 v at a frequency of 12 hz , a pulse width of 1 ms , and a duration of 1 minute . during tumescence , the maximal intracavernous pressure ( icp ) the total icp was determined by the area under the curve from the beginning of cavernous nerve stimulation to a point 20 s after stimulus termination . systemic blood pressure was measured by using a noninvasive tail - cuff system ( visitech systems , apex , nc , usa ) . the ratios of maximal icp and total icp ( area under the curve ) to mean systolic blood pressure ( msbp ) were calculated to adjust for variations in systemic blood pressure , as previously described . briefly , the corpus cavernosum tissue was harvested from 8-week - old c57bl/6j mice and transferred into sterile vials containing hank 's balanced salt solution ( gibco , carlsbad , ca , usa ) and was washed two times in phosphate - buffered saline ( pbs ) . the corpus cavernosum tissue was cut into two or three pieces and the samples were then plated on matrigel stock solution ( becton dickinson , mountain view , ca , usa)-coated 60-mm cell culture dishes . the matrigel was polymerized with a 5-minute incubation period at 37 , and 3 ml of complement medium 199 ( gibco ) , supplemented with 20% fetal bovine serum , 1% penicillin / streptomycin , 0.5 mg / ml heparin , and 5 ng / ml vascular endothelial growth factor - a , was added to the cell culture dish , and the dishes were then incubated at 37 with 5% co2 . after the cells were confluent and spread on the whole bottom of the dish ( about 2 to 3 weeks after the start of culture ) , only sprouting cells were used for subcultivation . briefly , cells were cultured on sterile cover glasses ( marienfeld laboratory , lauda - knigshofen , germany ) that were placed on the bottom of 24-well plates . individual chambers were incubated with antibody to platelet / endothelial cell adhesion molecule 1 ( 1:50 ; pecam-1 , an endothelial cell marker ; chemicon , temecula , ca , usa ) , cd90 ( 1:50 ; a fibroblast marker ; r&d systems inc . , minneapolis , mn , usa ) , or myosin ( 1:100 ; a smooth muscle cell marker ; sigma - aldrich , st . , ca , usa ) was applied to the chamber and nuclei were labeled . to examine the effect of the high - glucose condition with or without insulin treatment on ninjurin 1 expression , the primary cultured mcecs were serum - starved for 24 hours and were then exposed to following conditions for 48 hours : normal glucose condition ( 5 mmol ) , high - glucose condition ( 30 mmol ) , or high - glucose condition ( 30 mmol)+insulin treatment ( 1 nm ) . equal amounts of protein ( 50 g per lane ) were electrophoresed on sodium dodecylsulfate - polyacrylamide gels ( 12% ) , transferred to nitrocellulose membranes , and probed with antibodies to ninjurin 1 ( bd biosciences ; franklin lakes , nj , usa ) or -actin ( 1:6,000 ; abcam , cambridge , uk ) . for fluorescence microscopy ( n=6 per group ) , the penis tissue was fixed in 4% paraformaldehyde for 24 hours at 4 , and frozen tissue sections ( 8 m thick ) were incubated with antibodies to pecam-1 ( 1:50 ; chemicon ) , neurofilament ( 1:100 ; an axonal maker , sigma - aldrich ) , or ninjurin 1 ( 1:50 ; santa cruz biotechnology , santa cruz , ca , usa ) at 4 overnight . after several washes with pbs , the sections were incubated with tetramethyl rhodamine isothiocyanate- or fluorescein isothiocyanate - conjugated secondary antibodies ( zymed laboratories , south san francisco , ca , usa ) for 2 hours at room temperature . signals were visualized , and digital images were obtained with a confocal microscope ( fv1000 , olympus co. , tokyo , japan ) under identical exposure settings . the group comparisons were made by mann - whitney rank - sum tests or kruskal - wallis tests . specific - pathogen - free c57bl/6j mice were purchased from orient bio ( seongnam , korea ) . the experiments performed were approved by the institutional animal care and use subcommittee of our university . diabetes was induced in 8-week - old c57bl/6j mice by intraperitoneal injections of stz ( 50 mg / kg ) for 5 days consecutively as previously described . at 8 weeks after the induction of diabetes , the diabetic animals and their age - matched controls were used for western blot analysis , histologic examination , and physiologic erection study . fasting and postprandial blood glucose levels were determined with an accu - check blood glucose meter ( roche diagnostics , mannheim , germany ) before the mice were killed . the mice from the control and diabetic groups were anesthetized with ketamine ( 100 mg / kg ) and xylazine ( 5 mg / kg ) intramuscularly . stimulation parameters were 5 v at a frequency of 12 hz , a pulse width of 1 ms , and a duration of 1 minute . during tumescence , the maximal intracavernous pressure ( icp ) the total icp was determined by the area under the curve from the beginning of cavernous nerve stimulation to a point 20 s after stimulus termination . systemic blood pressure was measured by using a noninvasive tail - cuff system ( visitech systems , apex , nc , usa ) . the ratios of maximal icp and total icp ( area under the curve ) to mean systolic blood pressure ( msbp ) were calculated to adjust for variations in systemic blood pressure , as previously described . briefly , the corpus cavernosum tissue was harvested from 8-week - old c57bl/6j mice and transferred into sterile vials containing hank 's balanced salt solution ( gibco , carlsbad , ca , usa ) and was washed two times in phosphate - buffered saline ( pbs ) . the corpus cavernosum tissue was cut into two or three pieces and the samples were then plated on matrigel stock solution ( becton dickinson , mountain view , ca , usa)-coated 60-mm cell culture dishes . the matrigel was polymerized with a 5-minute incubation period at 37 , and 3 ml of complement medium 199 ( gibco ) , supplemented with 20% fetal bovine serum , 1% penicillin / streptomycin , 0.5 mg / ml heparin , and 5 ng / ml vascular endothelial growth factor - a , was added to the cell culture dish , and the dishes were then incubated at 37 with 5% co2 . after the cells were confluent and spread on the whole bottom of the dish ( about 2 to 3 weeks after the start of culture ) , only sprouting cells were used for subcultivation . briefly , cells were cultured on sterile cover glasses ( marienfeld laboratory , lauda - knigshofen , germany ) that were placed on the bottom of 24-well plates . individual chambers were incubated with antibody to platelet / endothelial cell adhesion molecule 1 ( 1:50 ; pecam-1 , an endothelial cell marker ; chemicon , temecula , ca , usa ) , cd90 ( 1:50 ; a fibroblast marker ; r&d systems inc . , minneapolis , mn , usa ) , or myosin ( 1:100 ; a smooth muscle cell marker ; sigma - aldrich , st . burlingame , ca , usa ) was applied to the chamber and nuclei were labeled . to examine the effect of the high - glucose condition with or without insulin treatment on ninjurin 1 expression , the primary cultured mcecs were serum - starved for 24 hours and were then exposed to following conditions for 48 hours : normal glucose condition ( 5 mmol ) , high - glucose condition ( 30 mmol ) , or high - glucose condition ( 30 mmol)+insulin treatment ( 1 nm ) . primary cultured mcecs incubated under each condition were used for western blot analysis . equal amounts of protein ( 50 g per lane ) were electrophoresed on sodium dodecylsulfate - polyacrylamide gels ( 12% ) , transferred to nitrocellulose membranes , and probed with antibodies to ninjurin 1 ( bd biosciences ; franklin lakes , nj , usa ) or -actin ( 1:6,000 ; abcam , cambridge , uk ) . for fluorescence microscopy ( n=6 per group ) , the penis tissue was fixed in 4% paraformaldehyde for 24 hours at 4 , and frozen tissue sections ( 8 m thick ) were incubated with antibodies to pecam-1 ( 1:50 ; chemicon ) , neurofilament ( 1:100 ; an axonal maker , sigma - aldrich ) , or ninjurin 1 ( 1:50 ; santa cruz biotechnology , santa cruz , ca , usa ) at 4 overnight . after several washes with pbs , the sections were incubated with tetramethyl rhodamine isothiocyanate- or fluorescein isothiocyanate - conjugated secondary antibodies ( zymed laboratories , south san francisco , ca , usa ) for 2 hours at room temperature . signals were visualized , and digital images were obtained with a confocal microscope ( fv1000 , olympus co. , tokyo , japan ) under identical exposure settings . the group comparisons were made by mann - whitney rank - sum tests or kruskal - wallis tests . metabolic and physiologic variables , including body weight , blood glucose concentrations , and msbp , are summarized in table 1 . eight weeks after diabetes was induced by intraperitoneal injections of stz , the fasting and postprandial blood glucose concentrations of the diabetic mice were significantly higher than those of the control mice . body weight was significantly lower in the diabetic mice than in the controls ( table 1 ) . erectile function parameters , such as the ratios of maximal icp and total icp to msbp , which were recorded at 5 v , were significantly lower in the diabetic mice than in the controls ( fig . 1 ) . no significant difference in systemic blood pressure was noted between the groups ( table 1 ) . we performed fluorescent immunohistochemistry and western blotting to evaluate the expression of ninjurin 1 in the penile tissue of diabetic mice . western blot analysis revealed a higher ninjurin 1 protein expression in the penile tissue of diabetic mice than in that of controls ( fig . 2 ) . immunohistochemical staining also showed an increase in ninjurin 1 expression in both corpus cavernosum tissue and the dorsal nerve bundle of diabetic mice compared with that in controls , whereas cavernous endothelial content ( pecam-1 ) and axonal content ( neurofilament ) in the dorsal nerve bundle were decreased by diabetes ( figs . immunofluorescent double - staining revealed that the majority of ninjurin 1 protein expression was overlapping with cavernous endothelial cells and with the dorsal nerve bundle ( figs . 3 , 4 ) . when we characterized the primary cultured cells , the majority of cells ( more than 95% ) showed positive staining for the endothelial cell marker ( pecam-1 ) , but did not show positive staining for the smooth muscle cell marker myosin or the fibroblast marker cd90 ( fig . we determined the effect of the high - glucose condition on ninjurin 1 expression in mcecs by western blot . ninjurin 1 expression was significantly increased by the high - glucose condition , and the levels were returned to baseline levels in mcecs exposed to the high - glucose condition and treated with insulin ( figs . metabolic and physiologic variables , including body weight , blood glucose concentrations , and msbp , are summarized in table 1 . eight weeks after diabetes was induced by intraperitoneal injections of stz , the fasting and postprandial blood glucose concentrations of the diabetic mice were significantly higher than those of the control mice . body weight was significantly lower in the diabetic mice than in the controls ( table 1 ) . erectile function parameters , such as the ratios of maximal icp and total icp to msbp , which were recorded at 5 v , were significantly lower in the diabetic mice than in the controls ( fig . 1 ) . no significant difference in systemic blood pressure was noted between the groups ( table 1 ) . we performed fluorescent immunohistochemistry and western blotting to evaluate the expression of ninjurin 1 in the penile tissue of diabetic mice . western blot analysis revealed a higher ninjurin 1 protein expression in the penile tissue of diabetic mice than in that of controls ( fig . 2 ) . immunohistochemical staining also showed an increase in ninjurin 1 expression in both corpus cavernosum tissue and the dorsal nerve bundle of diabetic mice compared with that in controls , whereas cavernous endothelial content ( pecam-1 ) and axonal content ( neurofilament ) in the dorsal nerve bundle were decreased by diabetes ( figs . 3 , 4 ) . immunofluorescent double - staining revealed that the majority of ninjurin 1 protein expression was overlapping with cavernous endothelial cells and with the dorsal nerve bundle ( figs . when we characterized the primary cultured cells , the majority of cells ( more than 95% ) showed positive staining for the endothelial cell marker ( pecam-1 ) , but did not show positive staining for the smooth muscle cell marker myosin or the fibroblast marker cd90 ( fig . we determined the effect of the high - glucose condition on ninjurin 1 expression in mcecs by western blot . ninjurin 1 expression was significantly increased by the high - glucose condition , and the levels were returned to baseline levels in mcecs exposed to the high - glucose condition and treated with insulin ( figs . we report here that the expression of ninjurin 1 protein was up - regulated in both cavernous endothelial cells and the dorsal nerve bundle of the diabetic mice . in the in vitro study in mcecs , ninjurin 1 expression was also increased by the high - glucose condition and was returned to baseline levels by treatment with insulin . peripheral neuropathy of the autonomic nerve and endothelial dysfunction are important causes of diabetic ed . the reduction in neuronal nitric oxide synthase - containing nerve fiber and endothelial nos activity , impaired penile angiogenesis , and the subsequent impairment in neurogenic- and endothelium - mediated smooth muscle relaxation are major contributing factors to diabetic ed . therefore , therapies aimed at restoring both nerve and endothelial function may be a promising therapeutic strategy in patients with ed associated with diabetes . in the present study , western blot analysis revealed higher ninjurin 1 expression in the penile tissue of diabetic mice than in that of age - matched controls . moreover , immunohistochemical staining showed higher ninjurin 1 expression in the dorsal nerve bundle of diabetic mice than in that of controls , whereas axonal content , as determined by neurofilament immunohistochemistry , was decreased by diabetes . previous study also reported an increase in ninjurin 1 expression after peripheral nerve injury , with the peak level of expression occurring 7 to 14 days after injury . several lines of evidence suggest that blood - borne immune cells play an important role in the onset and progression of cns - related disease , such as multiple sclerosis , an autoimmune inflammatory disease in the cns . a recent study reported in an experimental autoimmune encephalomyelitis model , an animal model for multiple sclerosis , that ninjurin 1 was highly expressed in myeloid cells ( macrophage / monocyte and neutrophils ) , which enhances penetration of these cells into the blood - brain barrier and induces neuroinflammation and nerve damage . the authors from this study suggested that ninjurin 1 could be a novel therapeutic target for neuroinflammatory disease related with immune cells . besides diabetic neuropathy , cavernous nerve injury is a major cause of radical prostatectomy - induced ed . cavernous nerve injury induces local inflammation and immune responses and finally contributes to the development of ed [ 22 - 24 ] . therefore , it is necessary to examine the effects of the blockade of ninjurin 1 on erectile function in animal models for diabetes or cavernous nerve injury . ninjurin 1 is also known to play a crucial role in vascular homeostasis during the embryonic period . in the present study , the expression of ninjurin 1 protein was significantly increased in both cavernous endothelial cells in vivo and primary cultured mcecs exposed to a high - glucose condition in vitro . ang1 is the ligand of the tie2 receptor tyrosine kinase and plays an important role in angiogenesis and maturation of blood vessels . in contrast , ang2 is an antagonist for ang1 on the tie2 receptor and is known to induce endothelial apoptosis through inhibition of survival signal , such as the phosphatidylinositol 3-kinase / akt pathway . a recent study reported that ninjurin 1 is a critical mediator for the regression of the hyaloids vascular system during the developmental period by regulation and ang1/ang2 balance , i.e. , decrease in ang1 expression and increase in ang2 expression , whereas ninjurin 1 neutralizing antibody inhibited regression of the hyaloids vascular system . collectively , these findings suggest that ninjurin 1 is involved in both neuropathy and angiopathy , which are important for the pathogenesis of diabetic ed . therefore , further studies are needed to test whether inhibition of the ninjurin 1 pathway can restore erectile function by reestablishing penile neural and microvascular structures . we for the first time have reported an up - regulation of ninjurin 1 protein in the penis of diabetic mice . the expression of ninjurin 1 protein was significantly induced in both cavernous endothelial cells and the dorsal nerve bundle of the diabetic mice in vivo and in mcecs exposed to the high - glucose condition in vitro . with regard to the role of ninjurin 1 in neuropathy and vascular regression , it would be interesting to examine the effects of inhibition of ninjurin 1 on erectile function in animal models of ed of vascular or neurogenic causes . Output:	purposeendothelial dysfunction and peripheral neuropathy are important mechanisms responsible for diabetes - induced erectile dysfunction ( ed ) . nerve injury - induced protein 1 ( ninjurin 1 ) is known to be related to neuroinflammatory processes and is also reported to induce vascular regression during the developmental period . in the present study , we determined the differential expression of ninjurin 1 in penile tissue of streptozotocin ( stz)-induced diabetic mice with ed.materials and methodsdiabetes was induced in 8-week - old c57bl/6j mice by intraperitoneal injections of stz ( 50 mg / kg for 5 days ) . eight weeks later , erectile function was measured by electrical stimulation of the cavernous nerve ( n=6 per group ) . the penis was then harvested for immunohistochemical analysis and western blot analysis for ninjurin 1 ( n=4 per group ) . we also determined ninjurin 1 expression in primary cultured mouse cavernous endothelial cells ( mcecs ) incubated under the following conditions : normal glucose condition ( 5 mm ) , high - glucose condition ( 30 mm ) , and high - glucose condition ( 30 mm)+insulin ( 1 nm).resultsthe expression of ninjurin 1 protein was significantly higher in both cavernous endothelial cells and the dorsal nerve bundle of diabetic mice than in those of controls . in the in vitro study in mcecs , ninjurin 1 expression was also significantly increased by the high - glucose condition and was returned to baseline levels by treatment with insulin.conclusionsregarding the role of ninjurin 1 in neuropathy and vascular regression , it would be interesting to examine the effects of inhibition of ninjurin 1 on erectile function in animal models of ed with a vascular or neurogenic cause .
PubmedSumm2038	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: indeed , the common pathological feature of chronic inflammation ( e.g. , chronic colitis ) and solid cancer involves a massive infiltration of immune cells into the sites . the pathological changes in solid cancers include recruitment and modifying of various types of dysregulated immune cells and endothelial cells to form a tumor microenvironment . a variety of chemokines and cytokines are produced by cancer cells and surrounding stromal cells and recruit leukocytes from the circulation to local sites according to their chemokine gradient . cancer - associated fibroblasts ( cafs ) constitute majority of the tumor stromal cells and play a critical role in tumor development . most of cafs are also recruited from bone marrow via chemokine signaling as well as immune cells . cancer cells regulate and modify these immune cells to escape from host side immune system . a growing body of evidence supports that cancer initiation and progression essentially depend on escape from host immunosurveillance . immune evasion involves a shift of immune responses , including imbalance in th1/th2 responses and enhancement of immunosuppressive cells such as myeloid - derived suppressor cells ( mdscs ) , regulatory t cells , m2 macrophages ( tumor - associated macrophages ) , and type 2 nkt cells . mdscs are a heterogeneous population of immune cells characterized by the ability to suppress cytotoxic functions of t cells and nk cells . mdscs originate from myeloid progenitor cells and are thought to be immature cells that do not differentiate into granulocytes , macrophages , or dendritic cells ( dcs ) . pathological conditions such as infection , trauma , autoimmune diseases , and cancer trigger expansion of mdscs in bone marrow and spleen . mdscs then accumulate in the peripheral blood , tumor , lymphoid organs , and parenchymal organs . in the past decade , mdscs have been thought essential especially in solid cancers and one of key drivers of not only cancer - associated immune evasion but also tumor progression and metastasis by establishing tumor microenvironment . indeed , the number of circulating mdscs in the peripheral blood correlates well with clinical cancer stage and metastatic tumor burden in patients [ 6 , 7 ] . mdscs also play a key role in gaining chemoresistant phenotype in cancer [ 8 , 9 ] . this review summarizes and discusses the recruitment mechanisms and immunosuppressive functions of mdscs and the potential strategies to target cancer - associated mdscs . mdscs are composed of heterogeneous immature myeloid cells that arise from bone marrow progenitor cells , at different stages of differentiation from early myeloid cells to more differentiated macrophages , granulocytes , or dendritic cells . circulating cd11bgr1 cells are arrested and accumulate in the splenic marginal zones and migrate to the red pulp and proliferate [ 10 , 11 ] , suggesting that cd11bgr1 cells in peripheral blood may represent both proliferated mdscs and precursors for mdscs . in mice , mdscs are broadly characterized by cd11bgr-1 , and mdscs are classified to two subsets as either granulocytic ( polymorphonuclear ) mdscs ( g - mdscs ) or mononuclear mdscs ( m - mdscs ) ( figure 1 ) . g - mdscs are defined as cd11bly6gly6c and m - mdscs as cd11bly6cly6 g . although the pattern of g - mdsc and m - mdsc subsets differs between tumors and organs , over 80% of mdscs are g - mdscs , whereas less than 10% of mdscs are m - mdscs in most of experimental models [ 1214 ] . however , substantial neutrophils also express both cd11b and ly6 g , causing difficulty to discriminate g - mdscs from neutrophils . in addition to reactive oxygen species ( ros ) and arginase 1 ( arg1 ) , m - csfr and cd244 have been proposed as phenotypes of g - mdscs . in human , phenotypes of human mdscs circulating cd33 cells , cd33hla - dr , or cd33hla - drlin are described to be mdscs in patients with renal cell cancer , colorectal cancer , or hepatocellular carcinoma , respectively [ 1618 ] . cd11bcd33 cells are reported as mdscs in peripheral blood of patients with non - small cell lung cancer . yu et al . suggested cd45cd13cd33cd14cd15 mdscs in tumors and peripheral blood of breast cancer patients . in general , human mdscs are defined as cd11b - positive , cd33-positive , hla - dr - negative or low , and lineage markers ( lin ) ( cd3 , cd14 , cd19 , cd56)-negative . zhang et al . described circulating cd11bcd33hla - drlin mdscs and their detailed profiles as cd13cd39cd115cd117cd124pd - l1cd14cd15cd66b in patients with colorectal cancer . similarly to mice , human mdscs can be phenotypically classified as granulocytic and monocytic population ( figure 1 ) . g - mdscs express a granulocytic marker cd15 or cd66b in addition to cd11b and cd33 in head and neck cancer , non - small cell lung cancer , pancreas cancer , bladder cancer , renal cell cancer , and breast cancer [ 2227 ] . on the other hand , human m - mdscs have been described as cd14hla - dr(cd33lin ) in hepatocellular carcinoma , metastatic prostate cancer , renal cell cancer , and malignant melanoma [ 2831 ] . m - mdscs have been generally described as cd11bcd14hla - dr or cd11bcd14cd15 ( or cd66b ) [ 2225 ] . vasquez - dunddel et al . defined m - mdscs in tumors , draining lymph nodes , and peripheral blood as cd11bcd33cd14hla - drcd34arg1ros in head and neck squamous cancer . direct cell - to - cell contact is required for such immunosuppressive activities of mdscs . l - arginine is a substrate for inducible nitric oxide synthase ( inos ) and arginase 1 ( arg1 ) . arg1 and reactive oxygen species ( ros ) are upregulated in activated g - mdscs , while arg1 and inos are highly expressed in activated m - mdscs . the upregulation of either arg1 or inos results in l - arginine shortage , leading to consequent inhibition of t cell proliferation through multiple mechanisms such as reduction of cd3 -chain expression and ifn-/il-2 secretion by t cells [ 4 , 13 , 34 , 35 ] . high levels of ros in g - mdscs can induce nitrosylation of the t cell receptor ( tcr ) during direct cell - to - cell communication , which contributes to the inhibition of antigen - specific t cell activation [ 13 , 36 , 37 ] . ros production by g - mdscs is known to be induced by several tumor - derived factors such as tgf , il-6 , il-10 , and gm - csf . the suppressive function of g - mdscs depends on arg1 and ros , whereas that of m - mdscs requires signal transducer and activator of transcription 1 ( stat1 ) and inos . in activated g - mdscs , stat3 is highly activated , which results in increased expression levels of ros via upregulation of nadph oxidase ( nox2 ) but not no production . on the other hand , stat1 and inos are highly upregulated in m - mdscs , resulting in increased levels of no but not ros production . in addition , stat6 signaling pathway is involved in upregulation of arg1 and tgf- through activation of il-4 and il-13 , leading to immunosuppressive activity [ 39 , 40 ] . however , the immunosuppressive mechanisms overlap between g - mdscs and m - mdscs in human cancers . inos is also upregulated in g - mdscs in a variety of human cancer [ 24 , 41 , 42 ] . cd14hla - dr m - mdscs express nadph oxidase component gp91 ( phox ) and produce high level of ros in human non - small cell lung cancer . these m - mdscs inhibit t cell proliferation and ifn- secretion in a cell - contact - dependent manner . cysteine is essential for t cell activation but can not be synthesized by t cells . antigen - presenting dendritic cells and macrophages can deliver cysteine by converting methionine and cystine to cysteine . mdscs also import extracellular cystine for converting it to cysteine but they do not export cysteine , leading to lack of cystine for dendritic cells and macrophage . recently , another subset of mdscs , cancer - induced fibrocytes , is proposed . this novel subset of mdscs shows phenotypic and functional characters of fibrocytes but mediates immune suppression by inhibiting t cell proliferation via indoleamine oxidase ( ido ) in human metastatic pediatric sarcomas . cd14hla - dr m - mdscs inhibit nk cell cytotoxicity and cytokine secretion in a cell - contact - dependent manner in human hepatocellular carcinoma . the inhibition of nk cells is independent of arginase activity . on the other hand , mdsc - mediated immunosuppressive activities against nk cells furthermore , mdscs can skew macrophage - derived cytokine profiles from type 1 to type 2 putatively through toll - like receptor 4 signaling pathway . indeed , increased mdsc levels in peripheral blood and tumor are closely associated with the infiltration of cd163 m2 macrophages in human esophageal cancer . although mechanisms of induction and trafficking mdscs are still elusive , multiple tumor - derived factors are suggested in preclinical models and cancer patients . these tumor - derived factors include granulocyte - macrophage colony - stimulating factor ( gm - csf ) , granulocyte colony - stimulating factor ( g - scf ) , macrophage colony - stimulating factor ( m - csf ) , il-1 , il-6 , vegf , il-13 , il-10 , prostaglandin e2 ( pge2 ) , tnf , bombina variegate peptide 8 ( bv8 , also known as prokineticin 2 or prok2 ) , and stem cell factor ( scf ) [ 5161 ] . most of these tumor - derived factors activate signaling pathways which involve stat3 or janus kinase ( jak ) in myeloid precursor cells . among these factors , gm - csf and il-6 may be most powerful in inducing mdscs from bone marrow hematopoietic progenitors . indeed , gm - csf induces the kit mdsc precursors in spleen of tumor - bearing mice . gm - csf is also required for recruitment of mdscs to the tumor microenvironment [ 64 , 65 ] . the stat3 activation in myeloid precursor cells enhances mdsc proliferation and expansion in bone marrow and also promotes the production of calcium binding proteins s100a8 and s100a9 which form heterodimers and inhibit differentiation to dendritic cells , contributing expansion and accumulation of mdscs . s100a8/a9 bind to cell surface glycoprotein receptors on mdscs and promote mdsc activation and migration through the nf-b signaling pathway . mdscs also synthesize and secrete s100a8/a9 , resulting in an autocrine feedback loop that sustains accumulation of mdscs . bv8 mobilizes immature myeloid cells from hematopoietic progenitor cells , and anti - bv8 treatment decreases mobilization of cd11bgr1 cells from bone marrow . tumor - derived pge2 promotes differentiation of cd11bgr1 cells from bone marrow stem cells through ep2 signaling . pge2 stimulates mdscs to produce high levels of arg1 and inos in tumor and spleen through ep4 signaling pathway , leading to t cell suppressive functions [ 35 , 68 ] . in human ovarian cancer , positive feedback loop between pge2 and cyclooxygenase-2 ( cox-2 ) facilitates the differentiation of human cd1a dcs toward stable m - mdscs ( cd14cd11bcd33cd34 ) . ccaat - enhancer - binding protein ( c / ebp ) also contributes to mdscs expansion and activation of the immunoregulatory activity of mdscs . additionally , human hepatic stellate cells ( hscs ) may have capacity to transform monocytes in peripheral blood into mdscs in a cd44-dependent manner . as aforementioned , several chemokine systems have been suggested to be involved in recruitment of mdscs to the primary cancer or the premetastatic niche . tumor - derived cxcr2-ligands ( il-8 , cxcl1 , cxcl2 , and cxcl5 ) attract mdscs to the tumor microenvironment [ 7173 ] and inhibition of cxcr2 profoundly suppresses gr-1 leukocyte migration into tumor . cxcr2 is expressed on most of circulating g - mdscs but not on m - mdscs and is prerequisite for g - mdscs to be recruited to tumor microenvironment . tumor - derived inflammatory mediator pge2 promotes cxcl12 production via ep2/4 receptor signaling and thus accumulates cxcr4-expressing mdscs in human ovarian cancer . on the other hand , ccr2 m - mdscs are recruited by tumor - derived ccl2 in mice and human [ 7678 ] . s100a8 and s100a9 not only activate mdscs but also attract mdscs by binding receptor for advanced glycation end products ( rage ) on mdscs . recruited mdscs secrete s100a8/a9 , resulting in further accumulation of mdscs in human breast cancer . g - mdscs promote tumor angiogenesis via secreting bv8 which is regulated by stat3 activation . mdscs also promote tumor - associated angiogenesis by secreting matrix metalloproteinase-9 ( mmp-9 ) . accumulating evidences mdscs facilitate cancer cell invasion and intravasation by secreting multiple proteolytic enzymes including mmps which are necessary for extracellular matrix degradation and disruption of endothelial cadherins , adhesion proteins or basement membrane of vessels [ 8183 ] . epithelial - mesenchymal transition ( emt ) is one of the steps for dissemination of cancer cells . when cancer cells undergo emt , cancer cells lose epithelial markers and gain mesenchymal phenotypes . g - mdscs induce emt in cancer cells using tgf- , epidermal growth factor ( egf ) , and hepatocyte growth factor ( hgf ) . on the other hand , mdscs also contribute to mesenchymal - epithelial transition ( met ) of cancer cells by secreting versican . mir-101 subsequently decreases the corepressor gene c - terminal binding protein-2 ( ctbp2 ) , which results in increased cancer cell stemness in ovarian cancer . on the contrary to prometastatic functions of mdscs , gr-1 myeloid cells generate a metastasis - resistant microenvironment in distant organs through the induction of thrombospondin-1 ( tsp-1 ) by tumor - derived prosaposin in mice bearing metastasis - incompetent tumors . this may represent not only heterogeneity of mdscs but also functional plasticity of mdscs , and the activities of mdscs might depend on a type of primary tumor . in addition , cd11bly6 g myeloid - derived cells differentiate into tumor - entrained neutrophils ( tens ) , which inhibit formation of premetastatic niche in the lungs . most of preclinical and clinical evidences indicate the association of mdscs with poor prognostic outcome as well as immunosuppressive effect in tumor microenvironment both in primary and metastatic tumor regardless of the heterogeneity of mdscs particularly in human cancer patients . the cancer - supportive activities provide a rationale for therapeutic approaches to target cancer - associated mdscs . the strategies targeting mdscs would be roughly divided into two approaches : ( 1 ) directly harnessing the mechanisms of mdsc expansion , recruitment , or immunosuppressive processes ( figure 2 ) to target the protumorigenic / prometastatic activities of mdscs and ( 2 ) sensitizing other anticancer agents through suppressing mdsc accumulation . several potential strategies for targeting mdscs can be proposed for each step of mdsc accumulation and the activities ( figure 2 ) . first potential treatment strategy is inhibiting tumor - derived or mdsc - derived factors which expand and mobilize mdscs from bone marrow or spleen . several neutralizing antibodies or inhibitors against tumor - derived factors or those receptors such as gm - csf , gm - csf receptor ( gm - csfr ) , m - csf , m - csf receptor ( m - csfr ) [ 88 , 89 ] , g - csf , vegf - a , or stem cell factor ( scf or kit ) however , anti - vegf - a monoclonal antibody , bevacizumab , could not reduce the accumulation of mdscs in human renal cell cancer . anti - il-6 receptor monoclonal antibody neutralizes tumor - derived il-6 and thus suppresses expansion of cancer - associated mdscs . also , directly inhibiting mdsc proliferation from hematopoietic precursor cells in bone marrow or spleen would be an approach to suppress mdsc expansion as well as decreasing circulating mdscs . some chemotherapy drugs such as gemcitabine and 5-fluorouracil decrease circulating mdscs [ 94 , 95 ] . however , one study shows that combination of gemcitabine and capecitabine does not affect the levels of mdscs in patients with advanced pancreatic cancer . an aminobisphosphonate zoledronic acid ( zoledronate ) reduces mobilization of mdscs through suppressing bone marrow progenitor - derived and tumor - derived mmp-9 [ 97 , 98 ] although one study reports that zoledronic acid impairs tumor - associated macrophage proliferation but not mdscs . sunitinib is a multikinase inhibitor which inhibits vegfr1 - 3 , platelet - derived growth factor receptor- ( pdgfr- ) and , stem cell factor receptor ( c - kit ) , flt3 , and ret . vemurafenib is a highly specific inhibitor of mutant b - raf and suppresses the release of tumor - derived soluble factors involved in mdsc generation . peroxisome proliferator - activated receptor- ( ppar ) is an anti - inflammatory molecule expressed in myeloid - lineage . of course , it is promising to inhibit accumulation of mdscs by targeting aforementioned tumor - derived factors or tumor - related inflammatory mediators or by using chemotherapeutic agents or monoclonal antibodies which are thought to suppress expansion of mdscs . however , some limitations should be considered particularly in clinical studies , that is , cause or result issue . the tumor - derived factors or inflammatory mediators can facilitate tumor progression by themselves . and the chemotherapeutic agents or monoclonal antibodies which are thought to inhibit mdsc accumulation also target cancer cells themselves , consequently leading to reduction of mdscs . accordingly , the attenuation of mdsc accumulation may , at least partly , reflect the reduction of the tumor burden caused by the therapeutic agents . a second therapeutic strategy would be inhibiting mdsc recruitment from the circulation to the tumor microenvironment or to the peripheral lymphoid organs . tumor - derived cxcr2-ligands , cxcl1 , cxcl2 , and cxcl5 , have been known to promote recruitment of mdscs to the site of tumor or to the premetastatic niche [ 12 , 7173 , 104 ] . since cxcr2 is mainly expressed on g - mdscs , cxcr2 inhibition may impair the recruitment of g - mdscs . accordingly , it is warranted to clarify the organ - specific preferences of mdsc types . other antagonists or inhibitors for chemokine systems such as anti - cxcr4 antibody and ccl2 neutralizing antibody would have potential to inhibit mdsc accumulation [ 4 , 75 , 105 ] . csf1r antagonist can suppress the infiltration of mdscs and tumor - associated macrophages ( tams ) to the site of tumor [ 88 , 89 ] . recently , one study suggested mdsc - specific peptides ( h6 and g3 ) and generated peptide - fc fusion proteins ( peptibodies ) which bind and affect both g - mdscs and m - mdscs . these peptibodies specifically depleted mdscs without affecting proinflammatory immune cell types such as mature dendritic cells . since these peptibodies target only mdscs , they have potential to solve aforementioned cause or result issue of mdsc - targeting therapies . cyclooxygenase-2 ( cox-2 ) inhibitors suppress activation of mdscs through ccl2 , cxcl12 , or pge2 inhibition and increase cytotoxic t lymphocytes ( ctls ) [ 11 , 75 , 107 ] . phosphodiesterase-5 ( pde-5 ) inhibitors ( sildenafil , tadalafil , and vardenafil ) are currently applied for clinical use in nonmalignant diseases ( e.g. , pulmonary hypertension ) . pde-5 inhibitors increase infiltration of activated ctls into tumor and tumor - induced t cell proliferation . pde-5 inhibitors restore -chain expression in tcr by decreasing mdscs [ 109 , 110 ] . importantly , pde-5 inhibitors downregulate expression of arg1 , inos , and il-4 in mdscs , which results in restoration of cytotoxic activities of t cells . synthetic triterpenoid c-28 methyl ester of 2-cyano-3,12-dioxooleana-1,9,-dien-28-oic acid ( cddo - me , bardoxolone methyl ) abrogates the immunosuppressive activities of mdscs and restored immune responses in both preclinical murine model and patients with renal cell carcinoma . a fourth strategy would be promoting the differentiation of mdscs into mature , nonsuppressive cells . all - trans retinoic acid ( atra ) , a vitamin a derivative , induces mdsc differentiation into mature myeloid cells by upregulating glutathione synthase in mdscs . removal of immature myeloid cells from dc fractions by atra restored the ability of dcs to stimulate antigen - specific t cell activity . in patients with metastatic renal cell carcinoma , atra administration decreased circulating cd33hla - drlin mdscs , which leads to improved myeloid / lymphoid dc ratio and antigen - specific t cell response . immune suppressive cd34 progenitors ( mdscs ) are increased in peripheral blood and tumors of patients with head and neck squamous cell cancer ( hnscc ) . these cd34 mdscs isolated from peripheral blood of hnscc patients can be differentiated into phenotypically and functionally dc - like cells in vitro , and 1,25-dihydroxyvitamin d3 ( 1,25(oh)2d3 ) accelerates differentiation of mdscs into the dc - like cells . indeed , 25-hydroxyvitamin d3 reduced the circulating cd34 mdscs in hnscc patients which resulted in increased levels of plasma il-12 and ifn- and t cell proliferation . although 25-hydroxyvitamin d3 alone failed to improve clinical outcome , it may have a potential to sensitize other chemotherapeutic agents by reducing mdscs . one study shows that tumor necrosis factor- ( tnf- ) blocks differentiation and augments suppressive activity of mdscs in chronic inflammatory settings and that administration of a tnf- antagonist , etanercept , reduces mdscs ' suppressive activity and promotes their maturation into dendritic cells and macrophages . however , this study is based on chronic inflammation model and remains to be confirmed in tumor - bearing model as well . several preclinical studies have shown that mdscs play a critical role in refractoriness against anticancer drugs such as bevacizumab , cyclophosphamide , anthracycline , and sunitinib [ 6 , 8 , 9 , 118 , 119 ] ( table 1 ) . therefore , an anti - vegf monoclonal antibody , bevacizumab , and a multikinase inhibitor , sunitinib , upregulate bv8 production from mdscs , unexpectedly leading to tumor - associated angiogenesis . indeed , anti - bv8 antibody treatment restores antiangiogenic effects and shows additive cytotoxic effect on those of anti - vegf antibody therapy . anthracycline and cyclophosphamide ( ac ) regimen enhances production of tumor necrosis factor- ( tnf- ) from tumor stromal cells and endothelial cells . cxcl1/2 promote recruitment of g - mdscs which secrete s100a8/9 , which results in amplifying the cxcl1/2-s100a8/9 loop and causing increased cancer cell survival , metastasis , and chemoresistance . accordingly interestingly , tumor - derived g - csf induced mdscs may contribute to radioresistant feature in uterine cervical cancer . anti - gr-1 neutralizing antibody or mdsc depletion by splenectomy restored radiosensitivity in cervical cancer xenografts . collectively , several chemotherapeutic agents or radiotherapy unexpectedly stimulates secretion of tumor - derived factor , which leads to expansion of mdscs critically causing refractoriness against the treatments . to break this vicious cycle by inhibiting recruitment of mdscs may sensitize these chemotherapeutic agents or radiotherapy and prevent the refractoriness against the anti - cancer therapies . combination of antigen - specific peptide and atra significantly increases and prolongs antigen - specific t cell response . in patients with extensive stage small cell lung cancer , atra improved the immune response to vaccination by significant reduction of cd33hla - drlin mdscs and consequent restored antigen - specific t cell response . her2/neu is a self - antigen with poor immunogenicity which is overexpressed in human cancer cells such as breast cancer . gemcitabine treatment with her2/neu vaccine and antiglucocorticoid - induced tumor necrosis factor receptor ( tnfr ) family - related receptor ( gitr ) antibody is effective in her2/neu - positive tumor , in which gemcitabine attenuates the tumor immune suppression through dramatic reduction of mdscs . sunitinib , a small molecule inhibitor of multiple receptor tyrosine kinases , reduces mdscs in peripheral blood of patients with renal cell carcinoma . the addition of sunitinib to low - dose radiation additionally suppressed tumor growth in glioblastoma ; however , the mechanisms of the tumor suppressive effect in this combination therapy are still unclear . the programmed death 1 ( pd1 ) receptor is an important immune checkpoint which is expressed on t cells , and pd1 signaling inhibits t cell receptor - mediated activation . recently , clinical evidences have shown limited efficacy of anti - pd1 monoclonal antibody in a minority of cancer patients . rhabdomyosarcoma attenuates the efficacy of anti - pd1 therapy by inducing expansion of cxcr2cd11bly6 g g - mdscs in preclinical model . anti - cxcr2 monoclonal antibody therapy improved significant antitumor effects of anti - pd1 treatment . these findings suggest that inhibition of cxcr2 g - mdsc tumor trafficking may sensitize and enhance anti - pd1 monoclonal antibody therapy in other cancers . collectively , targeting mdscs is a promising strategy to improve or increase immunotherapies against cancers . one interesting preclinical study indicates the application of mdscs as a vehicle to deliver radioisotope - coupled attenuated variant of listeria monocytogenes to tumor sites . on the other hand , as various studies show that circulating mdsc levels are correlated with tumor stages [ 21 , 128 ] , mdscs would be a good marker for a potential relapse of tumor or a surrogate marker which represents treatment efficacy . for instance , an elevated activity of circulating mdscs indicated by increased no production is suggested to be an early marker of incomplete treatment and a potential relapse of cancer in non - small cell lung cancer . also , mdsc levels in peripheral blood may predict chemotherapy failure in patients with pancreatic adenocarcinoma . immune evasion is a hallmark of cancer , and cancer - induced mdscs play an essential role in tumor progression through tumor - associated immunosuppression . therefore , it is a promising approach to break the vicious cycle in which mdscs are expanded and recruited to support tumor progression . several approaches have been sought , for example , inhibition of tumor - derived factors , suppression of generation and expansion of mdscs from hematopoietic progenitors , blockade of mdsc trafficking and infiltration , harnessing immune suppressive activities of mdscs , or facilitating differentiation of mdscs into mature nonsuppressive cells . furthermore , targeting mdscs may enhance and sensitize the effects of chemotherapies , vaccine therapies , or radiotherapies . however , mdscs are composed of heterogenous populations which are probably in different stages of differentiation and may have different activity in individuals with cancer . importantly , markers of mdscs in human are much more complicated than in mice , leading to further heterogeneity and difficulty . mdsc biology including these limitations remains to be elucidated for development of mdsc - targeting therapy , patient selection , and decision of treatment strategy . Output:	development of solid cancer depends on escape from host immunosurveillance . various types of immune cells contribute to tumor - induced immune suppression , including tumor associated macrophages , regulatory t cells , type 2 nkt cells , and myeloid - derived suppressor cells ( mdscs ) . growing body of evidences shows that mdscs play pivotal roles among these immunosuppressive cells in multiple steps of cancer progression . mdscs are immature myeloid cells that arise from myeloid progenitor cells and comprise a heterogeneous immune cell population . mdscs are characterized by the ability to suppress both adaptive and innate immunities mainly through direct inhibition of the cytotoxic functions of t cells and nk cells . in clinical settings , the number of circulating mdscs is associated with clinical stages and response to treatment in several cancers . moreover , mdscs are reported to contribute to chemoresistant phenotype . collectively , targeting mdscs could potentially provide a rationale for novel treatment strategies in cancer . this review summarizes recent understandings of mdscs in cancer and discusses promissing clinical approaches in cancer patients .
PubmedSumm2039	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: a 49-year - old woman presented with breathlessness on exertion with gradual onset over the previous one month ; there was no other significant past medical history . cardiac and respiratory examinations were normal , and the 12-lead electrocardiogram ( ecg ) was inconspicuous . two - dimensional echocardiography showed an anomalous left main coronary artery ( lmca ) arising from the main pulmonary artery ( mpa ) , moderate left ventricular dysfunction , an ejection fraction of 40% , and mild mitral regurgitation . the coronary angiogram revealed anomalous origin of the left coronary artery from the pulmonary artery ( alcapa ) . multislice computed tomography ( ct ) coronary angiography revealed an alcapa from the posterior wall of the mpa , which bifurcated into the left anterior descending ( lad ) artery and the left circumflex artery . the right coronary artery ( rca ) was tortuous , dilated , and a rising from the aorta ( fig . the patient was otherwise healthy and had an uneventful life history ( from childhood to adulthood ) , without any evidence of arrhythmic or syncopal attacks . the patient underwent coronary artery bypass grafting ( cabg ) with closure of the lmca opening from inside the mpa . after sternotomy and opening of the pericardium , we observed the rca arising from the aorta , which was hugely dilated , tortuous , and the lmca was arising from the lateral aspect of the mpa ( fig . it was not possible to mobilize the lmca for reimplantation to the aorta , therefore we planned a cabg with closure of the left main orifice of the lmca . the left internal mammary artery ( lima ) was harvested and cardiopulmonary bypass ( cpb ) was established with right atrial and aortic cannulation . after aortic cross clamping , cardioplegia was given by two methods : root cardioplegia was given in the aorta for the rca and followed by left main ostial cardioplegia after opening the mpa ( fig . the lima was grafted to the lad and the reversed saphenous vein was grafted to the second obtuse marginal artery . the left main ostium was closed with 6 - 0 prolene sutures from inside the mpa . aortic cross clamp was released , cabg completed , the patient was weaned from cpb , and the chest was closed . the patient was transferred to the intensive care unit with minimal inotropes , stable hemodynamics , and an isoelectric ecg . on postoperative day 2 , the patient had atrial fibrillation with a fast ventricular rate , which required amiodarone infusion , oral beta - blockers , and oral digoxin for rate control . after 72 hours ( i.e. , postoperative day 5 ) , the patient converted to normal sinus rhythm and was maintained with oral amiodarone and beta - blockers . on postoperative day 8 , a ct coronary angiogram was performed , which showed blood flowing through the bypass grafts and no communication between the mpa and lmca ( fig . first described by brooks in 1886 , alcapa has an incidence of 1 in 300,000 live births ( 0 . the anomaly , if left untreated , has a mortality rate approaching 90% in the first year of life . due to extensive intercoronary collaterals with the large dominant rca , alcapa occurs from either an abnormal septation of the conotruncus or from the persistence of the pulmonary buds combined with involution of the aortic buds that form the coronary arteries . it usually occurs as an isolated entity , but may be associated with atrial septal defect , ventricular septal defect , and coarctation of the aorta . alcapa is well tolerated in utero and in infancy because the pulmonary arterial pressure equals the systemic pressure , which leads to antegrade flow in the anomalous left coronary artery ( lca ) and the normal rca . development of the extensive collateral circulation between the rca and lca during this time period determines the extent of myocardial injury and the outcome . patients with alcapa who survive into adulthood have an enormous enlargement of the rca and extensive intercoronary collaterals . variable clinical presentation of alcapa is seen in this adult cohort , as patients may be asymptomatic or present with mitral insufficiency , ischemic cardiomyopathy , and cardiac failure or with malignant ventricular dysrhythmias . coronary angiography is the traditional standard method used for diagnosis ; however , cardiac ct and cardiac magnetic resonance imaging have been increasingly employed for the diagnosis of alcapa . echocardiography serves as an important tool to aid in diagnosis , because it visualizes the abnormal origin of the lca , dilated rca , retrograde filling of the anomalous coronary , abnormal diastolic flow in the mpa , and abnormal septal or epicardial color flow signals from the collateral vessels . examples of surgical correction methods include direct reimplantation of the left coronary into the aorta , baffle repair , interposition of the free subclavian artery , and use of the internal thoracic artery as a bypass graft . establishment of a dual coronary system is a widely used corrective surgery to repair alcapa in adult patients . all variants of cabg incorporate ligation of the alcapa at the pulmonary origin in order to avoid competitive flow . administration of cardioplegia for myocardial protection is a crucial element common to all surgical techniques . in this case , we gave cardioplegia in the aortic root and direct ostial cardioplegia for the left coronary system . management of alcapa in adults is controversial and may consist of ligation of the anomalous left coronary artery , cabg , or direct reimplantation . a high index of suspicion and appropriate diagnostic modalities should allow rapid diagnosis of alcapa in any patient seen with dilated congestive cardiac failure . recently , operations that reestablish two coronary arterial circulations have dramatically improved surgical results for an otherwise fatal disease . close and long - term follow - up of this patient is necessary to better understand the corrected natural history after surgical treatment of alcapa . we report a case of a 49-year - old woman with alcapa who presented with dyspnea on exertion . management involved cabg to the lad and obtuse marginal arteries , closure of the lmca ostium , and reestablishment of the dual coronary artery system . Output:	anomalous origin of the left coronary artery from the pulmonary artery ( alcapa ) is an extremely rare , potentially fatal , congenital anomaly with a high mortality rate in the first year of life . it occurs rarely in adulthood and may appear with malignant ventricular a rrhythmia or sudden death . we report a case of a 49-year - old woman with alcapa who presented with dyspnea on exertion . management was coronary artery bypass grafting to the left anterior descending artery and obtuse marginal arteries , closure of the left main coronary artery ostium , and reestablishment of the dual coronary artery system .
PubmedSumm2040	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: nemazee had been studying the deletion of autoreactive b cells in transgenic mice whose b cells all expressed receptors specific for a particular mhc molecule . when this transgene was bred onto mice that expressed that mhc protein , as expected , these mice lacked autoreactive b cells in the periphery . but to nemazee 's surprise , a large pool of autoreactive b cells persisted in the bone marrow ( 3 ) . to pinpoint the exact step at which the cells were escaping deletion , nemazee and his team engineered strains in which the autoantigen was targeted to different anatomical locations . expression of autoantigen in just the liver caused a near complete deletion of anti - mhc b cells , which were not replaced by non - autoreactive counterparts . but when autoantigen was also present in the bone marrow , there were many more peripheral b cells that were no longer self - reactive . nemazee discovered that these cells had escaped by switching antibody receptors they had new light chains derived from a rearranged version of the endogenous gene . the alterations took place in immature b cells , as nemazee only detected the rearrangements in bone marrow b cells . light chain replacement was simultaneously demonstrated in a different transgenic model by martin weigert and his team at the fox chase cancer center ( philadelphia , pa ) . while studying lupus , weigert had designed mice that carried a transgene for an anti - dna antibody . younger mice had fewer b cells , suggesting that autoreactive clones had been deleted . but adult mice had normal numbers of b cells . weigert and nemazee published their seminal results in a series of papers in the journal of experimental medicine in 1993 ( 46 ) . the studies were at first controversial , but numerous reports from other groups have since bolstered the case for receptor editing as a major mechanism for tolerance ( 7 ) . it is now estimated that at least 25% of b cells in the repertoire have undergone receptor editing ( 8) . nemazee 's anti - mhc transgenic mouse model has since been modified such that transgenes insert specifically into the heavy and light chain loci ( 9 ) . repairing its receptors rather than throwing away the whole cell might be the most efficient way to generate a diverse , non - self - reactive repertoire . Output:	in 1993 , david nemazee and martin weigert independently showed that autoreactive b cells could proofread , alter , and reexpress modified receptors to become nonautoreactive . this process , called receptor editing , has since gained prominence as the main mechanism of b cell tolerance .
PubmedSumm2041	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: iron deficiency , the common cause of anemia , underlies 115,000 maternal deaths worldwide per year.1 iron is a major constituent of hemoglobin , the oxygen - carrying pigment of the red blood cells . its level varies with different trimesters of pregnancy , and this , in turn , is a function of the net effect of absorption , utilization , and body losses . there is an increased demand for iron during pregnancy as a result of fetal and placental growth as well as expansion of blood volume . it has been shown that iron stores at conception strongly predict the maternal iron status and the risk of anemia later in pregnancy.2,3 several biomarkers have been used to assess the iron status in individuals . these include hemoglobin , serum ferritin , zinc protoporphyrrin , total iron - binding capacity , and transferrin saturation.4 serum ferritin is an iron - storage glycoprotein whose concentration is a true reflection of body iron stores in the absence of inflammatory / chronic infectious changes.4,5 the serum ferritin level is considered the best in assessing iron deficiency in pregnancy5 and together with hemoglobin level is regarded as the most efficient combination of indicators for monitoring the change in the iron status of a population.6 it is known that hemoglobin and iron concentrations vary during pregnancy , immediate postpartum , throughout the puerperium , and sometimes up to 12 weeks postpartum . when these parameters are below the normal threshold , complications such as iron - deficiency anemia may arise . iron - deficiency anemia during pregnancy has been associated with increased risks of low birth weight , prematurity , and maternal morbidity.7 studies carried out in africa and other regions of the world have identified iron deficiency as a common cause of anemia.8 it was reported in a study performed in pregnant women from zimbabwe , people s republic of china , india , and mexico ( between 1996 and 2008 ) that 43%73% of the subjects were iron deficient , of which 7%33% had iron - deficiency anemia.8 puerperal hematological status is important for optimal maternal health for breastfeeding and restoration of the maternal iron and hemoglobin status before subsequent conception . the prevalence of iron deficiency during the puerperium has not been documented in this area before this study . the aim of this study was to determine the rate and predictors of low serum ferritin levels during the puerperium among apparently healthy women in enugu , southeast nigeria . enugu state is a mainland state in southeastern nigeria , with a land area of ~8,727.1 km ( 3,369.6 mile2 ) located between the latitude 60 301n and longitude 70 301e . the state shares borders with abia and imo states to the south , ebonyi state to the east , benue state to the northeast , kogi state to the northwest , and anambra state to the west . enugu state lies partly within the semitropical rain forest belt of the south , and its vegetation changes gradually from the tropical rain forest to open woodlands and then to savannah . the 2006 population census estimated the enugu state population to be 3,257,298.9 the females constitute 50.1% of the population , and the population of women of reproductive age ( 1549 years ) was estimated to be 716,600.9 the study took place in the university of nigeria teaching hospital , ituku - ozalla , and enugu state university teaching hospital , park lane , both of which are tertiary health care centers that offer multidisciplinary services at primary and tertiary levels . in addition to serving as referral centers to other southeastern states , these institutions are also training and research centers . the study population included women with uncomplicated pregnancies who delivered singleton babies at term in the two study centers . exclusion criteria included women with hematological disorders like bleeding disorders , sickle cell anemia , or such conditions requiring blood transfusion or iron infusion ; women with other comorbidities like renal diseases , pre - eclampsia / eclampsia , diabetes mellitus , hiv infections ; and women with fever during the peripartum period . pregnant women who booked for antenatal care in the two centers were told about the study by the researcher / assistants during the antenatal clinic visits . the importance of serial blood sample collection and the need for compliance to follow - up were explained to the antenatal attendees . only those women whose pregnancies had reached term were given consent forms to sign and bring to the hospital while coming for delivery , if they consented . on admission for delivery at the study centers , the counseling on the proposed study was repeated . then , review of their antenatal records , preliminary history taking , and physical examination were done to exclude confounders . the actual enrollment took place 48 hours postdelivery after exclusion of those with postpartum hemorrhage requiring blood transfusion or iron infusion . forty - eight hours after delivery , a venous blood sample was collected in plain bottles to assay the serum ferritin levels . also , a blood sample was collected for hemoglobin estimation at 48 hours after delivery . this included the continuation of the routine hematinics given to them during pregnancy for at least 2 weeks postpartum , which consists tablets of ferrous sulfate ( 200 mg ) twice daily and folic acid once daily for 6 weeks . women with normal vaginal delivery were discharged 24 hours after delivery but stayed for 48 hours sample collection at no extra cost during the study period . follow - up was facilitated by obtaining the phone numbers from the patients and their partners or spouses in order to remind them of their scheduled appointment 23 days before time . participants were told to report any morbidity as they arose during the puerperium before the scheduled postnatal clinic visits at 6 weeks . at the 6-week postpartum visit , clinical evaluation of the subjects was carried out to assess the general well - being and if there was any form of morbidity necessitating medical intervention since discharge from the hospital . information regarding the commencement of menses or persistence of lochia discharge was also elicited . finally , venous blood samples were taken for hemoglobin and serum ferritin assay . this was performed by elisa technique using elisa machine stat fax-2600 ( awareness tech inc . , the sera now in well - labeled cryovials were stored at 80 in a freezer till a complete batch for one kit of reagent was obtained . prior to the biochemical analysis of each batch , the samples were thawed and kept at room temperature . the kit used was assay max human ferritin elisa kit , and it ran between 90 and 96 samples per batch . the assay employed the quantitative sandwich enzyme immuno assay technique that measured the ferritin levels in < 4 hours at room temperature . the samples of sera were made to undergo a series of incubations and washed at different intervals using the reagent and buffer solutions till optimal color density developed . the absorbance was then read immediately on a microplate reader at a wavelength of 450 nm . the proforma contained the patients sociodemographic variables , breastfeeding information , gestational age at delivery , estimated blood loss at delivery , weight of the baby , compliance with and duration of intake of routine hematinics , levels of serum ferritin and hemoglobin 48 hours and 6 weeks after delivery , and information regarding the return of menses . the principal investigator ( emegoakor fcj ) together with the trained research assistants collected samples for the 48-hour postdelivery hemoglobin and serum ferritin estimation for the two centers . two registrars from each of the study centers were trained on the study protocol , as research assistants , to help in the recruitment of patients and sample collection . all the samples were taken to university of nigeria teaching hospital for the biochemical analysis . the data collected were analyzed using the statistical package for social sciences ( spss ) computer software version 20.0 for windows . the normal range of serum ferritin for the study laboratory was 15200 ng / ml . simple descriptive statistics was used to calculate the mean serum ferritin 48 hours after delivery and at 6 weeks . categorical variables were compared using chi - square and fisher s exact tests where appropriate . logistic regression was done to determine the predictors of low serum ferritin levels at 6 weeks postpartum . the presence of low serum ferritin ( ie , < 15 ng / ml ) was coded as 1 while normal ferritin was coded as 0 ; independent predictor variables included in the model were parity , age , use of antenatal hematinics , educational status , anemia at 48 hours postpartum , low serum ferritin at 48 hours postpartum , anemia at term , birth interval , and employment status . the results of logistic regression were reported as adjusted odds ratios ( aor ) and 95% confidence intervals ( ci ) for odds ratios . the p - value for significance ethical consent was obtained from the research ethics committee of the university of nigeria teaching hospital . enugu state is a mainland state in southeastern nigeria , with a land area of ~8,727.1 km ( 3,369.6 mile2 ) located between the latitude 60 301n and longitude 70 301e . the state shares borders with abia and imo states to the south , ebonyi state to the east , benue state to the northeast , kogi state to the northwest , and anambra state to the west . enugu state lies partly within the semitropical rain forest belt of the south , and its vegetation changes gradually from the tropical rain forest to open woodlands and then to savannah . the 2006 population census estimated the enugu state population to be 3,257,298.9 the females constitute 50.1% of the population , and the population of women of reproductive age ( 1549 years ) was estimated to be 716,600.9 the study took place in the university of nigeria teaching hospital , ituku - ozalla , and enugu state university teaching hospital , park lane , both of which are tertiary health care centers that offer multidisciplinary services at primary and tertiary levels . in addition to serving as referral centers to other southeastern states , these institutions are also training and research centers . the study population included women with uncomplicated pregnancies who delivered singleton babies at term in the two study centers . exclusion criteria included women with hematological disorders like bleeding disorders , sickle cell anemia , or such conditions requiring blood transfusion or iron infusion ; women with other comorbidities like renal diseases , pre - eclampsia / eclampsia , diabetes mellitus , hiv infections ; and women with fever during the peripartum period . pregnant women who booked for antenatal care in the two centers were told about the study by the researcher / assistants during the antenatal clinic visits . the importance of serial blood sample collection and the need for compliance to follow - up were explained to the antenatal attendees . only those women whose pregnancies had reached term were given consent forms to sign and bring to the hospital while coming for delivery , if they consented . term was defined as 37 completed weeks to 41 + 6 weeks . on admission for delivery at the study centers , then , review of their antenatal records , preliminary history taking , and physical examination were done to exclude confounders . the actual enrollment took place 48 hours postdelivery after exclusion of those with postpartum hemorrhage requiring blood transfusion or iron infusion . forty - eight hours after delivery , a venous blood sample was collected in plain bottles to assay the serum ferritin levels . also , a blood sample was collected for hemoglobin estimation at 48 hours after delivery . this included the continuation of the routine hematinics given to them during pregnancy for at least 2 weeks postpartum , which consists tablets of ferrous sulfate ( 200 mg ) twice daily and folic acid once daily for 6 weeks . women with normal vaginal delivery were discharged 24 hours after delivery but stayed for 48 hours sample collection at no extra cost during the study period . follow - up was facilitated by obtaining the phone numbers from the patients and their partners or spouses in order to remind them of their scheduled appointment 23 days before time . participants were told to report any morbidity as they arose during the puerperium before the scheduled postnatal clinic visits at 6 weeks . at the 6-week postpartum visit , clinical evaluation of the subjects was carried out to assess the general well - being and if there was any form of morbidity necessitating medical intervention since discharge from the hospital . this was performed by elisa technique using elisa machine stat fax-2600 ( awareness tech inc . , the sera now in well - labeled cryovials were stored at 80 in a freezer till a complete batch for one kit of reagent was obtained . prior to the biochemical analysis of each batch , the samples were thawed and kept at room temperature . the kit used was assay max human ferritin elisa kit , and it ran between 90 and 96 samples per batch . the assay employed the quantitative sandwich enzyme immuno assay technique that measured the ferritin levels in < 4 hours at room temperature . the samples of sera were made to undergo a series of incubations and washed at different intervals using the reagent and buffer solutions till optimal color density developed . the absorbance was then read immediately on a microplate reader at a wavelength of 450 nm . the proforma contained the patients sociodemographic variables , breastfeeding information , gestational age at delivery , estimated blood loss at delivery , weight of the baby , compliance with and duration of intake of routine hematinics , levels of serum ferritin and hemoglobin 48 hours and 6 weeks after delivery , and information regarding the return of menses . the principal investigator ( emegoakor fcj ) together with the trained research assistants collected samples for the 48-hour postdelivery hemoglobin and serum ferritin estimation for the two centers . two registrars from each of the study centers were trained on the study protocol , as research assistants , to help in the recruitment of patients and sample collection . all the samples were taken to university of nigeria teaching hospital for the biochemical analysis . the data collected were analyzed using the statistical package for social sciences ( spss ) computer software version 20.0 for windows . the normal range of serum ferritin for the study laboratory was 15200 ng / ml . simple descriptive statistics was used to calculate the mean serum ferritin 48 hours after delivery and at 6 weeks . categorical variables were compared using chi - square and fisher s exact tests where appropriate . logistic regression was done to determine the predictors of low serum ferritin levels at 6 weeks postpartum . the presence of low serum ferritin ( ie , < 15 ng / ml ) was coded as 1 while normal ferritin was coded as 0 ; independent predictor variables included in the model were parity , age , use of antenatal hematinics , educational status , anemia at 48 hours postpartum , low serum ferritin at 48 hours postpartum , anemia at term , birth interval , and employment status . the results of logistic regression were reported as adjusted odds ratios ( aor ) and 95% confidence intervals ( ci ) for odds ratios . the p - value for significance ethical consent was obtained from the research ethics committee of the university of nigeria teaching hospital . of the 220 women who were recruited during the period of the study , 202 completed the study and were used for analysis . ninety - five percentage of the women had at least secondary education and at least 64% were employed . forty - eight hours postdelivery , 29.2% women had low ferritin levels and this decreased to 12.4% at 6 weeks postpartum . the mean serum ferritin levels at 48 hours and 6 weeks were 27.8218.41 g / l ( range : 585 g / l ) and 36.1221.53 g / l ( range : 1097 g / l ) , respectively . there was a statistically significant difference between the mean serum ferritin levels at 48 hours and 6 weeks postpartum ( t = 11.73 ; p - value < 0.001 ) . there was a significant positive correlation between hemoglobin level at 48 hours postpartum and serum ferritin level at 6 weeks postpartum ( r=0.39 , p<0.001 ) . there was , also , a significant positive correlation between serum ferritin level at 48 hours postdelivery and at 6 weeks postpartum ( r=0.89 , p<0.001 ) . table 2 summarizes the logistic regression to determine the predictors of serum ferritin levels at 6 weeks postpartum . it showed that low ferritin levels at 48 hours postpartum increased the likelihood of a low ferritin level at 6 weeks postpartum ( or = 78.1 , 95% ci = 8.8 , 108.3 ) . multiparity increased the likelihood of low ferritin levels at 6 weeks , ( or = 63.7 , 95% ci = 3.18 , 127.5 ) . the intake of antenatal hematinics for < 12 weeks increased the likelihood of low ferritin levels at 6 weeks postpartum , ( aor = 2.80 , 95% ci = 1.74 , 5.42 ) . maternal age < 20 years increased the likelihood of low ferritin levels at 6 weeks postpartum , ( aor = 0.7 , 95% ci = 0.53 , 0.93 ) . anemia at 48 hours postpartum increased the likelihood of a low ferritin level at 6 weeks postpartum , ( aor = 61.7 , 95% ci = 3.3 , 116.6 ) . this work is the second part of a study on the hematological status of healthy parturient women in enugu . available local studies on iron stores and other hematological parameters were performed during pregnancy and none aimed at the iron stores with regard to the postpartum period . therefore , most of the comparisons made here relied more on data published outside our immediate environment . values of serum ferritin levels obtained at 6 weeks in our study were in agreement with the values that were obtained by mara et al among women who received no iron supplementation.10 however , our results show lower ferritin levels than what were obtained among women who received iron supplementation in the studies by mara et al10 and milman et al.11 however , both studies differed slightly from our study in that they assayed serum ferritin at 8 weeks postpartum . since the return to the pre - pregnancy state is a continuous process and does not abruptly end at 6 weeks postpartum , the 2-week difference in the timing of the measurements could account for the higher values obtained in the quoted studies . unlike in our study , these researchers selected women who were of a relatively better nutritional status , which impacts positively on the iron stores even prior to conception . it is known that iron stores at conception are a strong predictor of the maternal iron level status and the risk of anemia later during pregnancy.2 also , the compromised prenatal iron status limits the amount of iron that would return to maternal iron stores following delivery.12 the women in the abovementioned studies were not just supplemented early but monitoring for adherence was instituted . our results suggest that the women with uncomplicated pregnancies on an average do not have a high prevalence of iron depletion in the study centers . this is based on the previously documented fact that a ferritin concentration < 15 g / l indicates iron depletion and levels 15 g / l implies the presence of iron stores.5,13 also , in women of reproductive age , a serum ferritin level < 15 g / l has shown a specificity of 98% and sensitivity of 75% for iron deficiency , as defined by no stainable bone marrow iron.14 our results , therefore , agree with previous studies on the ferritin level of pregnant and non - pregnant subjects , which concluded that iron deficiency did not have a high prevalence in nigeria.15,16 of importance , in this study , however , is the observation of different degrees of iron deficiency and sideropenia in which some women who had no anemia were found with iron deficiency . this may stem from the fact that iron deficiency occurs in stages , and the women observed with sideropenia may be in the early stage where serum ferritin is significantly reduced with no obvious change in the hemoglobin level.5,17 other stages of iron deficiency are the stage of iron - deficient erythropoiesis and finally iron - deficiency anemia.5,18 similar observations were also noted by previous researchers despite the use of iron / folate supplements.10,19 our results also show that antenatal and delivery factors were strong predictors of low serum ferritin levels in the puerperium . the fact that increasing duration of the use of antenatal hematinics reduced the likelihood of sideropenia at 6 weeks postpartum is not surprising . this is because antenatal hematinics help to replenish iron stores apart from replacing the iron lost during delivery . on its part , blood loss at delivery significantly correlated negatively with serum ferritin levels at 24 hours and 6 weeks . this is in keeping with previous studies that observed blood loss at delivery to be a strong predictor / risk factor for postpartum anemia and iron deficiency.2023 it is known that each ml of blood lost from the body results in a loss of 0.5 mg of iron and that each gram of hemoglobin contains 3.47 g of iron.24 this implies that the more the blood loss at delivery , the higher the incidence of iron deficiency and anemia at 6 weeks . other factors like anemia at term and 48 hours after delivery were predictive iron deficiencies at 6 weeks like was observed in previous studies12,20,25 this is expected considering that uncorrected prepartum iron deficiency anemia is already a risk factor for postpartum iron deficiency and anemia.2628 again , prepartum iron insufficiency and anemia make women vulnerable to the effects of blood loss at delivery thereby limiting the amount of iron that will return to the iron stores at 6 weeks.12,22,29 some sociodemographic characteristics such as age and parity were observed to predict low ferritin levels at 6 weeks . an increase in parity increased the likelihood of low serum ferritin levels and an increase in age reduced the likelihood of low ferritin levels at 6 weeks . the effect of high parity on serum ferritin may be due to an inadequate replacement of iron utilized in pregnancy ( and pregnancy has a high demand for iron as previously documented ) . therefore , the more the number of pregnancies carried to term , the more the iron status is depleted , worse if the birth interval is short . on its part , the effect of age on serum ferritin level seen in our study may possibly be due to the less demand of iron on growth and development seen in older than younger people as also observed in a previous study.30 a similar effect of parity and age on serum ferritin levels was also noted in a previous study.31 the limitations of this study include the fact that it was hospital based , which limits the external validity of our findings . factors such as nutritional status / dietary habits as well as possible parasitic ( hookworm ) infestations which may affect iron stores were not sought or controlled . nigeria comprises an ethnically diverse population while this study was conducted on igbo women only . a multicenter study of puerperal women belonging to the different ethnic groups will throw more light on the possible differences , if any , on the patterns of hematological changes after childbirth . changes in ferritin levels during puerperium in this cohort of women followed the same trend as observed in other parts of the world . however , there was a significant occurrence of low ferritin levels during the puerperium in the study centers , and this was associated mainly with pregnancy and delivery factors . efforts to improve the iron stores in parturient women could benefit from improving the compliance with antenatal hematinics and optimizing hemoglobin and iron levels before delivery . Output:	backgroundlow serum ferritin levels signify low iron stores and this could predispose to iron deficiency anemia.objectiveto determine the rate and predictors of low serum ferritin levels during the puerperium in enugu , southeast nigeria.study designa hospital - based prospective longitudinal study involving parturient women who delivered singleton fetuses at term . venous blood samples were collected to determine the serum ferritin concentration at 48 hours and 6 weeks postpartum . data analysis involved descriptive and inferential statistics at 95% confidence interval ( ci ) using statistical package for social sciences ( spss ) computer software version 20.0.resultstwo-hundred and two women who carried singleton pregnancies to term were studied . the mean serum ferritin levels at 48 hours and 6 weeks were 27.8218.41 g / l and 36.1221.53 g / l , respectively . forty - eight hours postdelivery , 29.2% had low ferritin levels and this decreased to 12.4% at 6 weeks postpartum . there was a significant positive correlation between the serum ferritin level at 48 hours postdelivery and the serum ferritin level at 6 weeks postpartum ( r=0.89 , p<0.001 ) . predictors of the low ferritin level at 6 weeks included age < 20 years ( odds ratio [ or ] = 0.70 , 95% ci = 0.53 , 0.93 ) , multiparity ( or = 63.7 , 95% ci = 3.18 , 127.5 ) , anemia at 48 hours postpartum ( or = 61.7 , 95% ci = 13.27 , 116.6 ) , a low ferritin level at 48 hours ( or = 78.1 , 95% ci = 8.8 , 108.3 ) , and intake of antenatal hematinics for <3 months ( or = 0.04 , 95% ci = 0.01 , 0.20).conclusionthere was a significant occurrence of low ferritin levels during the puerperium in the study centers , and this was associated mainly with pregnancy and delivery factors . efforts to improve the iron stores in parturient women could benefit from early booking and compliance with antenatal hematinics and optimizing hemoglobin and iron levels before delivery .
PubmedSumm2042	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: peroxisome proliferator activator receptor gamma ( ppar ) is a member of the nuclear receptor superfamily and is a ligand - activated transcription factor that regulates genes responsible for some very important biological functions including : cell growth , differentiation and metabolism and acts as dietary lipid sensor . the ppar gene is located on the human chromosome 3p25 - 24 and consists of nine exons and as a result of alternative splicing and promoter usage produces four isoforms ; ppar1 , ppar2 , ppar3 and ppar4 . primer extension studies have confirmed that ppar4 mrna is present in adipose tissue suggesting that the ppar4 isoform has a role in adipocyte biology . ppar plays critical roles in regulating lipid and carbohydrate metabolism ( energy balance ) , adipocyte differentiation , proliferation and insulin sensitivity and create a relationship between environmental factors and metabolic processes of the organism . ppar also plays roles in macrophages by enhancing foam cell formation and suppressing inflammatory cytokine production . since ppar is a nuclear transcription factor regulating multiple genes involved in energy production , glucose and lipid metabolism it may be a promising candidate gene for several major common diseases including cardiovascular disease , cancer , diabetes , inflammation and polymorphisms in this receptor may influence the pathology of these diseases . the ppar gene has been reported to be associated with various metabolic disorders including diabetes , obesity , dyslipidemia , insulin resistance , metabolic syndrome and coronary artery disease ( cad ) . several ppar2 gene single nucleotide polymorphisms ( snps ) have been reported to be associated with diabetes , obesity , dyslipidemia , insulin resistance , metabolic syndrome and cad . several genetic variations of the ppar gene have been found in the activation function domain 1 , deoxyribonucleic acid binding domain and ligand binding domain ( lbd ) of the receptor and confer some conformational changes in protein structure that may affect transcriptional activity . among several genetic variants of the ppar gene , two polymorphisms pro12ala of the exon b ( rs1801282 ) and the c1431 t silent substitution ( rs3856806 ) in the 6 exon are the most frequently occurring snps and have been associated with various diseases and extensive studies have been undertaken to assess the effects of these polymorphisms on many aspects of human physiology . the c1431 t polymorphism is also known as the c161 t or his477his was identified in 1998 by meirhaeghe et al . , and has been studied in relation to bone metabolism , metabolic syndrome , cad , obesity and glucose intolerance . the pro12ala protein polymorphism is due to a cca - to - gca missense mutation and was identified by yen et al . it is associated with type-2 diabetes mellitus , insulin resistance , obesity and metabolic disorders . in addition , numerous studies have demonstrated rare alleles of these two common variants play a role in the complex pathogenetic mechanism of major diseases supposed to be useful markers to evaluate the connection between ppar and metabolic derived disorders . given the pivotal roles of ppar in regulating metabolism several studies in various ethnic populations including , caucasians , mexican - americans , african - americans , asian , eastern asian , european , hispanic and non - hispanic have been conducted to determine allelic frequencies and genotypic distribution . there have been no studies of the distribution of c1431 t polymorphism in an iranian population and there is some controversy about the frequency of pro12ala polymorphism . therefore , the present study was designed to explore the distribution of these two common variants of ppar gene in iranian population and compare the finding with other populations . a number of 160 healthy individuals were analyzed to evaluate the frequency of ppar alleles and genotypes in a healthy iranian population . a total of 160 healthy volunteer subjects comprised of 70 males and 90 females , mainly people of iranian descent ( primarily persian ) , were selected randomly from all parts of mashhad as a second largest city in iran and enrolled in this study . a health questionnaire , including questions such as name , ethnicity , family history , age , sex and dietary habits , was provided to each participant . blood was taken in accordance with the world health organization protocol for blood donation and the healthy state of the all the participants was determined by medical history , physical examination and blood chemistry tests . subjects were excluded from the study if they had a history of congestive heart disease , liver and/or renal disease , endocrinological abnormalities and alcohol consumption or were under medications that altered blood pressure , glucose or lipid metabolism . the clinical and biochemical characteristics of the all the individuals enrolled into the study were normal at baseline . the protocol was approved by the ethics committee of the mashhad university of medical science and informed consent was obtained from all participants . genomic deoxyribonucleic acid ( dna ) was extracted from whole blood using the flexigene dna isolation kit ( qiagen ) . c1431 t and pro12ala polymorphisms of the ppar gene were determined by a predesigned taqman snp genotyping assay ( applied biosystems ) . oligonucleotides used for allelic discrimination assays for pro12ala and c1431 t were as follows : context sequences for pro/12ala ( [ vic / fam ] ) ( applied biosystems i d : c_1129864_10 ) : aactctgggagattctcctattgac[c / g ] cagaaagcgattccttcactgatacpro12 probe context sequences for c1431 t ( [ vic / fam ] ) ( applied biosystems i d : c_11922961_30 ) : context sequences for pro/12ala ( [ vic / fam ] ) ( applied biosystems i d : c_1129864_10 ) : aactctgggagattctcctattgac[c / g ] cagaaagcgattccttcactgatacpro12 probe context sequences for c1431 t ( [ vic / fam ] ) ( applied biosystems i d : c_11922961_30 ) : acctcagacagattgtcacggaaca[c / t ] gtgcagctactgcaggtgatcaagac1431 probe the reaction was performed in 25 l final volume with real - time polymerase chain reaction ( pcr ) using 96-well plates on an abi 7500 real time pcr system ( applied biosystems ) . the pcr conditions were 95c for 10 min and 40 cycles of 92c for 15 s and 60c for 1 min . individual genotypes identification was analyzed by sds software version 1.3 ( applied biosystems ) . for genotyping quality control , duplicate samples and negative controls were included to ensure the accuracy . allele frequencies of two snps were computed using genotype data obtained from these healthy controls and were compared with those reported in other populations . the statistical significance of the differences between genotypic distributions in populations was tested by the chi - square ( ) or fisher 's exact test using the spss 16.0 statistical package . allelic and genotypic frequencies for both polymorphisms of ppar gene were in hwe ( p 0.05 ) [ table 1 ] . the genotypic distribution of the c1431 t ppar polymorphism were 0.869 for cc , 0.119 for ct and 0.013 for tt and allelic frequencies were 0.93 for c and 0.07 for t respectively [ table 1 ] . for another pro12ala variant of ppar , genotypic distributions and allelic frequencies were , 0.813 for cc , 0.181 for cg and 0.06 for gg and 0.903 for c and 0.097 for g respectively [ table 1 ] . differences in allelic frequencies and genotype distribution of the polymorphisms between iranian population and those reported for other populations are shown in tables 2 and 3 respectively . allelic and genotypes frequency of studied population comparison of allelic and genotypic frequencies of ppar c1431 t variant between our population and others comparison of allelic and genotypic frequencies of ppar pro12ala variant between our population and others the present study represents the examination of the ppar gene c1431 t and pro12ala polymorphisms distribution in iranian healthy population and comparison with other populations . several studies have demonstrated these two common polymorphisms of ppar gene have a relationship with some major diseases . this current study is part of another study dealing with patients with metabolic syndrome and cads ( unpublished data ) ; many studies have showed these two common variants of ppar are associated with decreased risk of cad and ms in various populations . on the other hand , c1431 t polymorphism has not been studied in our population before , so this is why we selected these two common variants to evaluate in our population . allelic and genotypic frequencies of ppar c1431 t polymorphism in our population showed significant differences from other populations listed in table 2 except for two reports from european , one from italy , two in african - american , and one from sub - saharan african population . allelic but not genotypic frequencies in one report from european , one from caucasian , one from hispanic and one from indian showed significant differences from what we found in our iranian population . in all the populations the frequency of rare allele homozygote 's ( tt ) was low ( 0 - 7% ) , but there was one exception for a chinese population that showed more than 12% frequency that it might be due to small sample size . in general , chinese and other asian populations showed a higher frequency than others . in this case , our data was different from those of asian populations and was similar to those in australian , european and caucasian . with respect to pro12ala polymorphism , comparison of allelic and genotypic frequencies between our data and other studies listed in table 3 shows no significant differences with most of them . however there were some significant differences with two reports from china ( p = 0.00 ) , one from korea ( p = 0.041 ) , one from asia ( p = 0.00 ) , and a few reports from europe including , italy ( p = 0.003 ) , slovenia ( p = 0.017 ) , the czech republic ( p = 0.004 ) , france ( p = 0.00 ) and in an african - american population ( p = 0.00 ) as well . one report from europe and one within a caucasian population showed significance differences in allelic but not genotypic frequencies with our results . as shown in table 3 , the cc ( ( pro / pro ) genotype was the predominant form in all populations . data are presented in table 3 shows some inconsistency among different studies in the distribution of the g ( 12ala ) allele that might be due to the difference in racial and ethnic groups . the highest distribution of the g ( 12ala ) allele belongs to european and caucasian populations and the least is relate to chinese and african - american populations . most of the population in iran is genetically close to caucasian and their ancestors were the aryans who had migrated from central asia to iran . the main reason for the significant differences between our data and other populations results from iran as a country with an ethnically diverse population including pars , turk , kurd , tajik , turkmen , baloch and special religious such as muslims , zoroastrians , jews , christians and assyrians . however , iran is located along the ancient silk road and connected asia to europe and during the course of history , iranian population has encountered foreigners including macedonians , arabs , turks and mongols on various occasion . therefore , the population living in this country might be mixed due to contacting with others and the immigration of some people from neighboring nations . several studies have been undertaken on the association between the pro12ala and c1431 t mutations with numerous diseases such as cardiovascular disease , cancer , diabetes , inflammation on various populations and the results showed controversy . the controversial findings related to these polymorphisms may be attributable to genetically differences in populations . therefore , it is necessary to study in different populations to reach a general consensus in this field . an important limitation in our study further association study with a larger population base is needed to confirm our results and may be useful in understanding these snps roles in pathology of the main disease in the future . statistical differences in the distribution of two common polymorphisms of ppar gene between iranian population and others showed the importance of studying these snps in relation to some major diseases . regards to iranian different genetic with some other populations in asia and other continents , it seems that our study confirms this difference concerning the two common ppar polymorphisms . Output:	background : peroxisome proliferator activator receptor gamma ( ppar ) is a nuclear transcription factor regulating multiple genes involved in cell growth , differentiation , carbohydrate and lipid metabolism and energy production . several genetic variations in the ppar gene have been identified to be associated with diabetes , obesity , dyslipidemia , insulin resistance , metabolic syndrome and coronary artery disease . the present study was designed to explore the distribution of two common single nucleotide polymorphisms of the ppar gene ( c1431 t and pro12ala ) in an iranian population.materials and methods : genotype frequencies for these two polymorphisms were compared for 160 healthy iranian individuals with reports from other populations . the genotyping was performed using real - time polymerase chain reaction.results:the genotype distribution of the c1431 t ppar polymorphism was 0.869 for the cc genotype , 0.119 for the ct genotype and 0.013 for uncommon tt genotype . allelic frequencies were 0.93 for c and 0.07 for t allele respectively . for the pro12ala polymorphism of ppar gene , genotypic distributions and allelic frequencies were , 0.813 for cc , 0.181 for cg and 0.06 for gg and 0.903 for c and 0.097 for g respectively . allelic and genotypic frequencies for both polymorphisms of ppar gene were in hardy - weinberg equilibrium.conclusions:iran is a country with an ethnically diverse population and a comparison of allelic and genotypic frequencies of ppar c1431 t and pro12ala polymorphisms between our population and others showed significant differences .
PubmedSumm2043	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: over the past few years , semiconductor nanowires ( nws ) have become a large research field due to their interesting electrical and optical properties due to quantum effects , large surface to volume ratio , and capability for bottom - up assembly . nw - based memory devices , lasers , single - molecule sensors , and solar cells have been demonstrated in recent years . additionally , one outstanding feature of the nw growth is that the crystal structure can be different from the stable bulk crystal structure , with possible striking effects on their optical properties . the stable bulk crystal structure for most iii v binary semiconductor materials is cubic zincblende ( zb ) , while synthesis of the hexagonal wurtzite ( wz ) structure in bulk gaas requires high pressure and temperature . however , nws can adopt both zb and wz crystal structures , and previous research has demonstrated the control of wz and zb crystal structure in gaas nws grown by metal organic chemical vapor deposition ( mocvd ) . v nws is [ 111 ] with a stacking sequence of abcabc , which could also be denoted as 3c using the ramsdell notation . the equivalent growth direction in the hexagonal case is [ 000.1 ] , with the stacking sequence ababab for wz , or 2h in the ramsdell notation . 3c ( zb ) and 2h ( wz ) thus differ only in the stacking sequence in the [ 111]/[000.1 ] direction and exhibit a small difference in cohesive energies , with 3c being the highest , hence the stable phase in bulk . however , with the large surface to volume ratio of nws , compared to bulk , the relative contribution of lateral surface energies to the total free energy of formed nws has to be considered and is a key parameter in growth models attempting to explain formation of polytypes in iii nws with a structure different from their bulk counterpart should be treated as metastable , and the growth is kinetically controlled . treating nws as metastable implies that their phase could change to a more stable one upon perturbation . in addition , if it is the large surface to volume ratio that enables growth of metastable nws , will the structure be maintained under large radial overgrowth , where the surface to volume ratio decreases ? phase transformation of grown gaas nws has been observed previously , where 2h structure epitaxially buried in planar 3c overgrowth gradually adopted the structure of the burying layer . phase transformation has also been observed in ingaas nws , using an electron beam to deliver a high energy dose to the structure , enough to transform 2h to a mixed 3c/2h structure . in the case of epitaxial burying , one layer at a time of the metastable 2h is gradually surrounded with a layer of the stable 3c structure . with a lateral mismatch of the top layer burying the nw , a linear defect is created at the interface between the burying layer and nw , which could be viewed as a shockley partial dislocation in the 3c matrix . once the dislocation has formed along at least two sides of the nw , it could propagate into the nw without changing its length . as the surrounding 3c layer has a higher cohesive energy , the total energy associated with the linear defect is decreased as the dislocation propagates through the nw and changes its structure to 3c . by completely burying the structure in a 3c matrix , the low energy surfaces of 2h can not stabilize the structure any longer , and the structure is forced into 3c . to further develop the model to apply to free - standing structures , contributions from the surface energies in addition , the dislocation line between a buried structure and its surrounding matrix compared to the dislocation line formed between free - standing structures has different geometries , which could have a big impact on the propagation of the dislocation line . in this work , we investigated what happens when adjacent wz nws radially merge with each other , for different amounts of radial overgrowth and merging . the approach resembles the epitaxial lateral overgrowth ( elo ) technique , which has been used since the 1960s to reduce threading dislocation densities in growth of lattice - mismatched thin films . elo is widely used to grow high quality gan films , but is not as common for non - nitride iii - vs . using a mocvd system , we first grew gaas 2h nws on 3c gaas substrate , and by radial overgrowth in a second step increased the nw diameter . during radial growth , the shell will adopt the same crystal structure as the core , which has been observed in several nanowire core shell material combinations . with increasing radial overgrowth time , more and more of the initial 2h nws radially merge with each other . for the longest growth times , the initial gaas 2h nws have merged to form a nearly continuous film . the crystal structure of samples with different amounts of radial overgrowth and merging was investigated . we used high resolution xrd and high resolution transmission electron microscopy ( hrtem ) for structural investigations . although one might expect to find a 2h film after the radial overgrowth , our observations show that this is not the case . once individual nws merge together , the crystal structure of the merged nws is transformed progressively from the initial pure 2h structure to a mixed 2h/3c structure . we use a rather simple combinatorial approach to explain the main features in the resulting crystal structure of radially merged nws . comparing energy contributions from differences in cohesive energy , surface energies and dislocation line energy , only a small energy difference for transforming 2h compared to 3c is found , which justifies the use of a combinatorial model . gaas 2h nws were grown on gaas ( 111)b substrates using mocvd and 70 nm size - selected au seed particles with a surface density of 1 m with 2h growth parameters as reported previously . a pregrowth annealing step was performed in a 100 mbar mocvd reactor under hydrogen ( h2 ) and arsine ( ash3 ) for 10 min at 650 c to remove the surface oxides and form catalyst droplets . after annealing , the temperature was reduced to the growth temperature of 550 c , and tmga was introduced to initiate the growth . the molar fractions of tmga and ash3 flow were 1.46 10 and 5.5 10 , respectively ( v / iii ratio of 4 ) , with a total gas flow of 6 slm . after 2h nw growth , the samples were cooled down under h2 only to prevent formation of a 3c neck region . after cooldown , the samples were removed from the mocvd system in order to remove the au - alloy seed particles by etching in a cyanide based etching solution ( tfac from transene ) . removing the au seed particles is essential to limit axial elongation of the nws during subsequent radial overgrowth . for this step , the samples were reintroduced into the mocvd system and annealed under a h2/ash3 environment at 650 c for 10 min to remove surface oxides formed as the nws were exposed to air . the temperature was then reduced to 630 c , and tmga was introduced to initiate radial overgrowth . the growth time ranged from 5 to 240 min with molar fractions of tmga and ash3 of 1.5 10 and 2.2 10 , respectively ( v / iii ratio 150 ) . ( a ) growth of wz gaas nws , ( b ) removal of au particles , ( c ) radial overgrowth of gaas nws , ( d ) merging of nws in order to form the designed structure . for structural investigations of large ensembles of nws and parasitic substrate growth , we performed high - resolution x - ray diffraction ( xrd ) measurements on as - grown samples . two laboratory diffractometers with cu anode and channel - cut monochromators were used together with one - dimensional detectors to record reciprocal space maps ( rsms ) . the measurements were performed in the scattering plane spanned by the [ 111 ] and [ 112 ] directions of the substrate . symmetric rsms around the 3c gaas ( 333 ) reciprocal lattice point and 2h gaas ( 000.6 ) as well as asymmetric rsms between 3c gaas substrate ( 224 ) bragg peak and ( 331 ) twinned gaas bragg peak were recorded , which also covers the 2h gaas ( 101.5 ) . the occurrence of the 3c ( 331 ) from twinned nws in the same azimuth as the 3c substrate ( 224 ) is due to the 180 rotation around [ 111 ] caused by twin defects . to characterize the local crystal structure and the quality at the interface between merged nws , we used focused ion beam ( fib ) to prepare cross sectional lamellae samples for transmission electron microscopy ( tem ) . using fib could potentially induce defects , but our method is gentle enough to preserve high crystalline quality of the prepared sample . high resolution tem ( hrtem ) , dark field tem ( dftem ) , scanning tem ( stem ) , and selected area electron diffraction ( saed ) were used for analysis . the grown nws have an average length of 2 m and average diameter of 100 nm prior to radial overgrowth , with 2h structure including low densities of stacking defects in the range of < 1020 m , as revealed by hrtem ; see figure 2a c . after the etching process , most of the au catalyst particles on the tops of the nws were removed , but the nws were insignificantly affected by the etchant ( see detail in supporting information , figure s1 ) . ( a c ) tem and hrtem images of a 2h structured nw before overgrowth imaged in the 112.0 zone axis . ( d i ) sem images of nws with radial overgrowth for different times . with 30 tilting angle : ( d ) 5 min ; ( e ) 20 min ; ( f ) 60 min ; top view sem images of nws radial overgrowth with different time : ( g ) 120 min ; ( h ) 180 min ; ( i ) 240 min . the insets shown in ( g ) and ( h ) are sem images taken at a 30 tilting angle , scale bars are 2000 nm . figure 2d i shows the nw overgrowth at different stages ranging from 5 to 240 min overgrowth time . after the first 5 min , we observed mainly an increase in the volume of the pyramidal base of the nws ; see figure 2d . the 2h nws are terminated by low energy { 101.0 } facets , which gives a low possibility of nucleation directly at the side facets . the pyramidal base consists of higher index facets , which have a higher surface energy due to the increased density of dangling bonds , resulting in an increased rate of nucleation of new layers . after 20 min ( see figure 2e ) , the base starts to form { 101.0 } facets , and the morphology evolution of the nws changes with radial overgrowth time of 60 min ( compare figure 2e , f ) . at the kink site between the base and the nw side facets , the higher coordination number is expected to have an even higher rate of nucleation . nucleation at the kink site followed by growth of layers along the nw [ 000.1 ] direction could explain the homogeneous shell at 60 min with steps at the top of the nw . as the growth time is increased further , the area between the nws becomes smaller , and the nws eventually merge . after 240 min of growth , most of the nws have merged with each other , and a nearly continuous film is formed . in parallel with the radial growth of the nws , we observe continuous growth on the substrate surface . as the parasitic substrate growth proceeds , some of the lower parts of the initial 2h nws are buried , which explains the apparently thinner base in figure 2f compared to 2d and 2e . however , most parts of the nws are radially merged before being buried in parasitic substrate growth , as seen in the insets in figure 2g , h . average nw length and diameter and parasitic substrate growth thickness are plotted in supporting information , figure s3 . the nws never become fully buried in the parasitic substrate growth , as the openings between the nws become too small to allow adatoms to reach the bottom of the pits between them . instead , growth on the nw top facets increases with decreased openings between the nws . using high resolution xrd to characterize the crystal structure of the samples , the measurements collect diffraction data from several mm area of the sample , and signals from the substrate and grown structures can not be separated directly . however , as the 2h structure only originates from the nws , the nw signal can be distinguished from the substrate . as 3c occurs in two different twin directions , two sets of 3c diffraction peaks occur ; one originates from the substrate and possible 3c formed during the growth in the same twin direction , and the other set solely from 3c formed during the growth with the other twin direction . figure 3 displays rsms from the sample with 120 min radial overgrowth time . in the symmetric rsm , the 3c gaas ( 333 ) peak at q[111 ] = 5.777 and 2h gaas ( 000.6 ) at q[111 ] both the substrate and twinned 3c contribute to the signal of the 3c ( 333 ) bragg peak and can not be distinguished . between the 2h and 3c peaks , a weaker signal is visible at q[111 ] = 5.758 , which originates from gaas 4h . x - ray diffraction reciprocal space maps ( rsms ) of the sample with 120 min radial overgrowth time . panel ( a ) shows a symmetric rsm around the 3c ( 333 ) and 2h ( 000.6 ) bragg peaks , ( b ) shows asymmetric rsm including zb substrate ( 224 ) , ( 331 ) twin 3c , 2h ( 101.5 ) , and 4h ( 101.9 ) , ( 101.10 ) , and ( 101.11 ) bragg reflections . in ( c ) and ( d ) , details of the twin 3c and 2h peaks are shown . with roman numerals , the positions of possible detector streak ( i , white line ) , broadening due to tilt distribution ( ii , red line ) and finite nw radius ( iii , black line ) are marked . linecuts from the rsms are shown in ( e ) and ( f ) for the symmetric and asymmetric case , respectively . linecuts from rsms recorded in both ( 331 ) and ( 224 ) azimuths are overlaid in ( e ) and ( f ) . in the asymmetric rsms in figure 3b , the peaks are attributed to the 3c ( 224 ) bragg peak , which originates from the substrate and grown 3c material with same twin orientation as the substrate as the peak with highest intensity and the 3c ( 331 ) bragg peak from 3c grown with opposite twin - orientation as the second strongest peak . in between the two 3c peaks , the 2h ( 101.5 ) is visible next to a weak 4h gaas ( 101.10 ) peak . on either side of the 3c peaks , two more 4h peaks are visible , 4h gaas ( 101.9 ) and 4h gaas ( 101.11 ) . for asymmetric rsms for all six growth shell times figure 3c , d shows details in the rsm around the 2h ( 101.5 ) and 3c ( 331 ) bragg peaks . the position of possible detector streak is marked with white ( i ) , broadening due to tilt distribution with red ( ii ) and broadening due to finite radius of the nanowires as black ( iii ) . in figure 3e , f , line cuts from the symmetric and asymmetric rsms for both ( 331 ) and ( 224 ) are shown . using the bragg peak integrated intensity , we estimate the change in scattering volume . first , line cuts along [ 111 ] are made in the asymmetric rsms ; see figure 4a . to each peak , a voigt shape curve was fitted after removal of background signal , and the curves integrated intensity was normalized with respect to the substrate peak intensity . as a consistency check , peaks from both azimuths on each sample were used . using different azimuths , the difference in structure factors for ( 224 ) and ( 331 ) as well as the difference in illuminated area the peak intensities for the 2h , 4h ( 101.10 ) and twinned 3c peaks are plotted . as seen , the 2h initially increases , and the largest 2h volume was found for the sample with 60 min radial overgrowth time . for the twinned 3c , the volume increases for the full radial overgrowth series . for the 4h ( 101.10 ) peak intensity , the scattering intensity follows a similar trend as for 2h , but with the strongest scattering signal at 120 min radial overgrowth time . ( a ) linecuts along q_[111 ] from asymmetric rsm with substrate peak at ( 224 ) plotted with offsets . the position for different bragg peaks decreasing intensities for the 4h and 2h peaks are visible for the longer radial overgrowth times . ( b ) integrated intensities for the 2h ( red , square ) , 4h ( triangles , green ) , and the 3c ( blue , circles ) peaks arising from the twinned zb . the trend lines are guides for the eyes only . by preparing a cross sectional lamella sample by fib , the crystal structure along the nws and the parasitic substrate growth were investigated after 120 min radial growth . figure 5a , b shows merged sections in bright - field ( bf ) and dark - field ( df ) tem images , respectively . the dark field is taken around ( 111 ) and ( 002 ) reflection for the two different 3c twin directions , and ( 000.1 ) for the 2h . the top part includes two free - standing nws , which have pure 2h crystal structure with no visible stacking faults . beneath this section , the merged region includes two crystal orientations with twin boundaries perpendicular to the nw growth direction , and the barcode - like pattern of twin regions span the majority of the formerly individual nanowires . the parasitic substrate growth results in 3c crystal structure which is heavily twinned including boundaries not perpendicular to the nw growth direction ( diffraction pattern and additional tem image in supporting information , figure s5 ) . the crystal structure of the merged region is confirmed by the saed pattern shown in the inset of figure 5b . the saed pattern indicates two dominant 3c crystals twinned in the nw growth direction only . ( a ) bftem images from an fib lamella with two partly merged nanowires . ( b ) dftem image of the same region as in ( a ) highlighting the crystal structure in different parts marked with different colors . the free - standing nanowires have 2h crystal structure with no visible stacking faults , while the merged regions have 3c crystal structure . the substrate growth has 3c crystal structure which is heavily twinned including boundaries not perpendicular to the growth direction . the inset in ( b ) shows saed from the merged region , which indicates two dominant 3c crystals twinned in the growth direction only . figure 6 shows the tem image of a fib lamella with two merged nws after 120 min radial overgrowth in which a large section of 4h was found . the corresponding saed patterns of the 2h and 4h phases are shown in figure 6b , c , respectively . the independent gaas nw segment before the merging exhibits a pure 2h crystal structure with almost no stacking faults , which is clear from the saed pattern in figure 6b . the appearance of a ( 000.1 ) reflection indicated by a small arrow in figure 6c and two times shorter distance between ( 000.0 ) and ( 000.1 ) reflection compared to saed pattern of 2h in figure 6b confirms the existence of 4h crystal structure . the hrtem image shown in figure 6d and the fft shown in the inset the length of the 4h segment is approximately 200 nm , which is consistent with the observation in xrd ( see below ) . ( b ) saed from the region in the blue circle in ( a ) showing 2h structure , and ( c ) saed from the region in the green circle in ( a ) showing the 4h crystal structure . ( d ) hrtem image from the region marked with rectangular in ( a ) showing a band of 4h crystal structure merging the two nanowires . the inset in ( d ) shows a fft from the left nanowire , which is a typical 4h diffractogram . xrd performed on nanostructures is affected by finite size effects , which in the case of nws are usually dominated by their small diameter , causing a broadening in the [ 112 ] direction . nws also tend to not grow exactly perpendicular to the substrate surface , but a small mosaicity due to a random nw tilt distribution causes a broadening in the same direction as the finite width in the symmetric rsms . with both finite size and tilt contributing to the same broadening in symmetric rsms , it is not possible to distinguish one from the other . since the effects scale differently for different reflections as well as show up in slightly different directions in asymmetric rsms , a distinction of the two effects is possible in the asymmetric rsms . the finite size contribution is always inversely proportional to the width of the nws , while the broadening due to tilt distribution increases linearly with q. in the asymmetric scans , the tilt distribution and finite width broadens the peak in different directions , which is indicated in figure 3c , d . for the 120 min radial overgrowth time shown in figure 3 , the broadening due to tilt distribution is larger than that due to finite nw width . the tilt distribution found is on the order of 0.1 deg , which is on the same order of magnitude as for other nw samples , such inas 2h and insb 2h on inas 3c and gap tilt distribution is a general feature in nw samples and is a limiting factor when performing high resolution xrd on such samples . another finite size effect present in the asymmetric rsms is the broadening of the diffraction peaks in the [ 111 ] direction , caused by short segments of the respective polytype in the nw growth direction . the inverse width of the diffraction peaks is proportional to the average segment length and for the 4h peaks segment lengths of 100200 nm were estimated . worth noting is that no 4h was found in the reference nws prior to radial overgrowth . for the 3c and 2h peaks , the widths are limited by the instrumental resolution and length estimations are therefore not possible . with a very high crystal quality as seen in both the hrtem analysis and xrd , we are able to accurately calculate the lattice parameters of both gaas 2h and 4h using the asymmetric rsms . the accuracy of the measurements is similar to the case of inp 2h , limited mainly by experimental setup and beam broadening due to measurement geometries . table 1 summarizes the calculated lattice parameters with relative differences with respect to ideal tetrahedral structures , using geometrically converted cubic 3c lattice parameters . both the 2h and 4h unit cells are elongated in the nw growth direction and contracted in the radial direction compared to 3c . from the sem images shown in figure 2 , a continuous increase in 2h volume would be expected for the full radial overgrowth time series , since the radii of the nws continuously increase with time . however , from the xrd data , a decrease in 2h is found for radial overgrowth times longer than 60 min . intuitively , one could argue that the decrease in 2h is due to epitaxial burying in a 3c substrate layer , similar to what has been observed in refs ( 18 and 19 ) , which would also be supported by the continuous increase in 3c diffraction signal with increasing growth time . however , comparing the volume of not - yet - buried nws for 5 and 180 min radial overgrowth , the volume is 2 orders of magnitude larger for the longer growth time , yet the 2h diffraction signal is weaker . hence , the burying of the nws is not sufficient to explain the drastic decrease in the 2h signal . in addition , epitaxial burying of the 2h nws in a 3c matrix would not explain the appearance of 4h segments , estimated to be 100200 nm long in the samples with the most 4h , as an epitaxial burying driven phase transformation would force the structure to adopt the burying matrix structure , in this case 3c . interestingly , the largest increase in 4h scattering intensity is found as the 2h scattering signal starts to decrease . the decreasing 2h signal coincides with the onset of the radial merging of adjacent nws , as seen in the inset in figure 2g , h . as reported by ng et al . , a high energy electron beam could potentially induce crystal phase transformation , which could affect the results obtained by tem . however , the agreement of the xrd and tem results suggests that the crystal phase transformation is independent of measurement technique . in addition , reference nws and the parts not yet merged showed only pure 2h structure under similar tem analysis ; therefore , the possibility of electron beam induced transformation is discarded . instead of explaining the phase transformation by epitaxial burying , we propose that the transformation could be explained by a nw nw interaction . to understand the process of the merging better we come back to the stacking sequences of the different crystallographic phases . the stacking along the hexagonal [ 000.1 ] direction of the 2h crystal structure can be described as ababab , while for 3c crystal structure it is described as abcabc along the equivalent [ 111 ] direction . each capital letter denotes a pair of layers consisting of one group v and group iii atom , a bilayer , at three distinct lateral positions . as the nws grow in [ 111]/[000.1 ] , the position of as in each new bilayer will determine whether the layer is in cubic or hexagonal position . the stacking sequences of 2h can also equivalently be described as bcbcbc , acacac , etc . ( nw 1 ) retains the stacking order as ababab and the second 2h structured nw ( nw 2 ) retains the stacking order as bababa. when the bottom bilayer of nw 1 reaches the periphery of nw 2 , the as atoms in the bottom bilayer of nw 1 is not in lateral registry with that of nw 2 . in principle now two scenarios compete with each other . either this mismatch is kept at the cost of having a line defect at the interface of the merging nws , or in order to release this mismatch , the line defect may propagate through one of the nws to accommodate the corresponding as plane . for such a gliding process , three different translation vectors ( the burger vectors , b ) are possible , 1/3[101.0 ] , 1/3[110.0 ] , and 1/3[011.0 ] ; see figure 7 . this edge dislocation could be seen as a shockley partial dislocation , and when propagating through the crystal changes the configuration of the as from hexagonal to cubic position ( or vice versa ) . ( a ) schematic of two miss - matched 2h nws viewed in the { 000.1}/{111 } direction . the mismatch causes a translation in the [ 110.0 ] direction of the as atoms in nw 1 closest to the interface . ( b d ) schematics displaying propagation of a defect through one of the nanowires . changing a 2h gaas to 3c gaas would to a first approximation result in a lower total energy for the crystal , as the cohesive energy is 24 mev / iii v pair higher for 3c . changing from 2h { 101.0}-type facets to 3c { 112}-type facets increases the surface energy due to the increase of dangling bonds , with a net energy difference of 3.06 mev / nm . in addition to the change in cohesive energy and surface energies , the dislocation line itself has an energy per unit length . the dislocation line energy originates from the strain field around it and the distorted bonds at the dislocation core . detailed calculations on defects in gaas 2h are lacking , but as an approximation the dislocation line energy could be calculated as edis = gb , where g is the shear modulus and b is the magnitude of the burgers vector . using the shear modulus of gaas 3c , as the shear modulus of 2h is unknown , the dislocation line energy is estimated to be 10.74 ev / nm . the 2h nws are hexagonal and have { 101.0 } type facets , which are perpendicular to the burgers vector , and the nws all have facets parallel to each other . for two nws merging along the full length of a side facet , as in figure 7 , the dislocation line has initially the same length as the side facet of the nws . for the dislocation line to propagate into one of the two nws by only changing the position of one as , it has to increase its length ; see figure 7b d . the minimum increase in length would then be b / cos(30 ) , if the propagation occurs at a nw facet . in total similarly , if the dislocation propagates into a 3c structure , the total energy increases by 2.88 ev for the first as moved . the difference between the two values is less than thermal energy at the present growth temperature . with the dislocation line energy as the dominant factor for the energies related to the phase transformation , the initial structures of the two merging nws are of minor importance the probability of either one to transform is roughly equal . comparing our phase transformation with the case of epitaxial burying described by patriarche et al . , the dislocation is of the same type , and both transformations are triggered by the interaction between two crystals with as atoms at different lateral positions . however , in the case of epitaxial burying where the burying matrix completely surrounds the nw , the nw has to adopt the structure of the burying matrix or keep the dislocation at the periphery of the nw . to let the dislocation propagate through the nw makes the dislocation shorter , and eventually disappear , which is the thermodynamically most favorable case , and the final structure will be easy to predict the one of the burying matrix . in our case , with a nw nw interaction , the dislocation could propagate into either crystal , and the final structure is much harder to predict . as the dislocation line energy depends on the shear modulus of the crystal and the burgers vector , changing growth conditions would not change the dynamics of the crystal phase transformation . changing the growth temperature could however change the propagation rate of the dislocation , as the peierls energy barrier sets an activation energy for the propagation . hypothetically , by reducing growth temperature enough , propagation of the defect could be kinetically hindered such that it stays at the interface between the nws , which then would retain their initial structure . however , since growth temperature in this study is high enough to ensure reasonable growth rates , we assume that the defect propagation is not kinetically hindered . assuming equal probability for the dislocation to propagate in either nw , a combinatorial approach gives the probabilities of different structures . assuming that a dislocation can propagate with the same probability into either nw as two nws merge , each merging bilayer gives rise to two equally likely outcomes . for merging of two nws with 10 bilayers each there will be 2 possible outcomes . if we involve a third 10 bilayer nw in the merging process , there will be ( 2 ) outcomes . to estimate the amount of 3c , 2h , and 4h in each outcome , we define a 3c segment to be of at least six bilayers ( two unit cells ) , 2h as six bilayers ( three unit cells ) and 4h as eight bilayers ( two unit cells ) . if the 10 bilayer long segment does not include any of the above segments , it is considered to be a mixed structure . using a simple matlab script , we simulate the merging of two , three , and four 2h nws with each 10 bilayers ( where the 10 bilayer limit is chosen to limit computation time ) . a schematic view on the merging process the simulations shows that the merging of just two nws gives a mixed fraction of around 80%that is , most of the possible outcomes have only segments consisting of less than six bilayers of any considered polytype . 2h is still the most common polytype , with 1516% , depending on the initial stacking sequences of the nws and how many nws merge . both 3c and 4h show up with lower amounts ( 36% and 0.30.5% , respectively ; see supporting information , figure s7 ) . this result fits well with our experimental observation : the most likely structure for radially merged 2h nws is a mixture between 2h and 3c . the increase in twinned 3c seen in the xrd measurements comes not only from the nws , but also from the parasitic substrate growth , and the 36% of 3c found in the nws in our combinatorial model is not the major contribution to the signal . for a structure with very short segments of 2h and 3c sandwiched , streaking between the xrd peaks is expected , and this is also what we observe in our rsms ; see figure 3b and supporting information figure s4 . using this simple combinatorial approach , we are able to describe the main features of the merging process . the lattice parameter for 2h gaas has been reported previously , both using xrd and electron diffraction on nws and on 2h in gaas powder obtained at high pressure . however , the 2h lattice parameters in the former report were measured from nws with a mixed crystal structure , consisting of 2h segments sandwiched between 3c , and most likely strained by the 3c segments . for our structures , the very few sfs in the pure 2h before any merging process are not likely to influence the overall 2h structure , and the 2h crystal is fully relaxed . using the sample with 60 min radial overgrowth , we measured the 2h and 4h unit cell parameters on the sample with the largest volume of pure 2h . the results are summarized in table 1 . with a larger lattice plane spacing along the c - axis in 2h as compared to the 3c counterpart and a smaller in - plane distance , the c / a ratio is measured to be 1.649 , which is 0.98% higher than the ideal 1.633 for hexagonal close - packed structures with perfect tetrahedral coordination . the value found is in good agreement to theoretical values of 1.6461.651 . similarly , we found a c / a ratio of 3.282 for 4h , which is 0.49% higher than ideal . the measured gaas 4h lattice parameters reported here are to the authors best knowledge the first experimental report on gaas 4h lattice parameters . the higher c / a ratio compared to the ideal is typical for metastable hexagonal structures and gives bonding slightly distorted from an ideal tetrahedron . for 4h , the deviation from the ideal close - packing is smaller than for 2h , suggesting 4h to be the more stable structure , but still with 3c as the most stable one . hence , the cohesive energy for 2h should be lower than for 3c , with 4h in between the two . the difference in cohesive energy gives a higher nucleation barrier for 2h compared to 4h or 3c in gaas , which should result in a higher probability of 4h than 2h during growth . however , the surface energies of the nucleus and its surroundings play a crucial role in determining the structure formed during nw growth . v nws is the assumption that the surface energies of 2h are considerably lower than those of 3c , which overcomes the difference in cohesive energy . assuming surface energies of 2h to be considerably lower than those of 4h as well explains why observations of 4h grown in nws are very few , and mostly only occur in a transition region between 2h and 3c . in this work , we studied the crystal structure of radially overgrown gaas wurtzite ( 2h ) nws , especially the resulting crystal structure as nws progressively grow into each other and merge . we characterized the grown structure by high resolution xrd and tem with diffraction analysis . we observed that the free - standing gaas nws retain the pure 2h structure until they merge with each other , at which point the merged segments transform progressively from pure 2h structure to a mixture between zinc blende ( 3c ) structure with twins and 2h . we also found sections with the 4h crystal structure in the merged nws and determined lattice parameters of both 2h and 4h gaas . the phase transformation continues as long as the merging process proceeds , driven by a dislocation formed between adjacent nws due to lateral mismatch of as in the ( 111)/(000.1 ) plane . using a combinatorial approach , we estimated the probability of the resulting structure after radial merging of 2h nws , which shows that a heavily mixed structure is the most probably outcome , in agreement with our xrd and tem measurements . Output:	iii v nanowires ( nws ) grown with metal organic chemical vapor deposition commonly show a polytypic crystal structure , allowing growth of structures not found in the bulk counterpart . in this paper we studied the radial overgrowth of pure wurtzite ( wz ) gaas nanowires and characterized the samples with high resolution x - ray diffraction ( xrd ) to reveal the crystal structure of the grown material . in particular , we investigated what happens when adjacent wz nws radially merge with each other by analyzing the evolution of xrd peaks for different amounts of radial overgrowth and merging . by preparing cross - sectional lamella samples we also analyzed the local crystal structure of partly merged nws by transmission electron microscopy . once individual nws start to merge , the crystal structure of the merged segments is transformed progressively from initial pure wz to a mixed wz / zb structure . the merging process is then modeled using a simple combinatorial approach , which predicts that merging of two or more wz nws will result in a mixed crystal structure containing wz , zb , and 4h . the existence large and relaxed segments of 4h structure within the merged nws was confirmed by xrd , allowing us to accurately determine the lattice parameters of gaas 4h . we compare the measured wz and 4h unit cells with an ideal tetrahedron and find that both the polytypes are elongated in the c - axis and compressed in the a - axis compared to the geometrically converted cubic zb unit cell .
PubmedSumm2044	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: vasospasm ( vsp ) is one of the major causes for prolonged neurologic deficit in patients with aneurysmal subarachnoid hemorrhage.1 treatment of vsp is therefore one of the main priorities for these patients . balloon angioplasty or stenting are options for vsp of the proximal vessels , whereas distally located vsp is preferably treated pharmacologically.2 a long list of pharmacologic agents has been tested for their effect on vsp . intravenous ( iv ) or oral application of nimodipine combined with triple h therapy ( hypertension , hypervolemia , hemodilution ) is currently recommended as the first - line prevention for vsp.3 in patients not responding to this preventive therapy , short local intra - arterial nimodipine administration ( slina ) is recommended , although it is often only effective during the time of infusion.4 however , intra - arterial ( ia ) nimodipine administration may be prolonged for a few days . few case series have reported about the feasibility , efficacy , and safety of this treatment , and standardized guidelines are lacking.5 6 7 8 we report our experience with continuous local intra - arterial nimodipine administration ( clina ) via a catheter in the internal carotid artery ( ica ) in refractory cerebral vsp . a 40-year - old woman was referred to our hospital due to headache and nausea that started acutely the day of admission . she had no history of hypertension or other diseases but was a former smoker . native computed tomography ( ct ) scan showed a subarachnoid hemorrhage ( sah ) extending to the suprasellar cisterns and both sylvian fissures ( modified fisher grade 4 ) . the hunt & hess grade was 2 , and the glasgow coma scale and the world federation of neurosurgery grade were 15 and 1 , respectively . ct angiography ( cta ) revealed a saccular aneurysm of the anterior communicating artery ( acom ) ( fig . the aneurysm was located at the most distal acom segment , at the origin of the left anterior cerebral artery a2 segment ( fig . 1 ) . ( a ) three - dimensional angiography of left carotid artery showing an anterior communicating artery aneurysm measuring 3 7 mm . the aneurysm was successfully treated by coil embolization , and the patient was asymptomatic after the procedure . according to the department 's standard post - sah treatment protocol , the patient was given continuous iv nimodipine , fluids , and statins . at day 6 , the patient developed hemiparesis predominantly affecting the right arm and severe expressive aphasia corresponding to a national institute of health stroke scale ( nihss ) score of 6 ( facial palsy 1 ; arm 2 ; leg 1 , aphasia 2 ) . cta showed vsp in the intracranial ica , the anterior cerebral artery proximal segment ( a1 ) , and the middle cerebral artery proximal segment ( m1 ) bilaterally , but predominantly on the left side . vsp was present also in the basilar artery and the left posterior cerebral artery . at day 7 , 30-minute ia administration of 4 mg nimodipine in the left intracranial ica and 30 minute ia administration of 4 mg nimodipine in the left intracranial vertebral artery led to a clear improvement of angiographic vsp and neurologic impairment . due to the recurrence of symptoms at day 8 , 30-minute ia administration of 4 mg nimodipine in the left intracranial ica and in the left intracranial vertebral artery were repeated with beneficial but transitory effect . at day 9 , 1 hour ia administration of 8 mg nimodipine followed by 3-minute ia administration of 30 mg papaverine were performed . digital subtraction angiography ( dsa ) showed partial resolution of vsp in the left m1 and a1 but persistent vsp in the left m2 and a2 ( fig . 2 ) . left carotid angiogram at the initiation of continuous local intra - arterial ( ia ) nimodipine infusion day 9 ( three short local ia nimodipine administrations were done on days 7 , 8 , and 9 after coil embolization ) . ( a ) severe vasospasm is seen in the a1 , a2 , and m2 branches . ( b ) partial resolution after 4 mg nimodipine in 30 minutes but still severe vasospasm in a2 . transcranial color - coded sonography ( tccs ) measurements of blood flow velocity in the left distal m1 showed no changes before and after ia treatment ( fig . 3 ) . two hours later , cerebral mri showed multiple watershed infarcts in the left carotid artery territory and in the left parietal lobe ( fig . 4 ) . mean velocity in the left distal m1 segment measured with transcranial color - coded sonography . diffusion - weighted magnetic resonance imaging day 9 showed scattered cortical infarctions in the left hemisphere . the patient was quickly referred to tccs that showed marked worsening of hemodynamics ( mean velocity of 300 cm / s in the left distal m1 , lindegaard index > 7 ) . clina was established by introducing a microcatheter ( progreat 2.7f , terumo interventional system , tokyo , japan ) into the left ica through a guiding catheter . with the microcatheter in place nimodipine dose was set at 2 mg / h . an iv bolus dose of 5000 ie heparin was administered initially , followed by 1500 to 2500 units / h to keep the activated clotting time > 200 s. a few hours later , the patient showed improvement of symptoms with normalized motor function and only a slight residual aphasia ( nihss = 1 ) . at day 10 , clina was stopped , the catheter and the sheath were removed , and the full dosage ( 2 mg / h ) was given iv for 7 days and then orally for another 7 days . in addition to nimodipine , advanced vsp treatment including pressor , fluids , statins , magnesium , and dantrolene was administered . during the hospital stay a lumbar drain was inserted due to transient hydrocephalus , and antibiotics were given due to a urinary tract infection . she had no measurable speech impairment but a slight disturbance of fine motor skills in the right hand , and she complained about gait instability and dysesthesia in the right hemibody ( nihss = 0 ) . at follow - up day 90 , the patient reported reduced stamina , mild headache , and non drug - demanding neuropathic pain in the right arm . motor function was normalized , and she had no apparent speech impairment ( nihss = 0 ) . tccs showed normal blood flow velocities in all intracranial arteries , and mri showed no new ischemic lesions . this is the first case of continuous local intra - arterial nimodipine administration ( clina ) for refractory vsp described in scandinavia . the treatment had a dramatic effect on both angiographic vsp and neurologic impairment with no complications . in line with earlier reports,5 6 7 8 9 10 our case shows that clina is feasible , safe , and may be effective in patients with symptomatic medically refractory vsp following sah . however , there is no consensus when to start the treatment . although other reports recommend starting clina if 30-minute ia nimodipine administration does not show improvement of vsp , the threshold for referring patients to dsa is not always specified . in some earlier reports , patients were referred to dsa either after a decline in the parameters of multimodal neuromonitoring or after progressive loss of consciousness postoperatively.5 7 8 in another report , the patients underwent dsa within the first 2 hours after onset of clinical symptoms and decrease in brain tissue oxygen.6 in our patient , clina was started 2 days after the first focal neurologic impairment . the two slinas performed at day 7 and day 8 showed good angiographic response and partial clinical improvement . the third slina showed partial resolution of angiographic vsp with no clinical and doppler response , and mri showed established cerebral infarction . in an earlier study , 21 of 25 patients treated with clina developed cerebral infarction secondary to vsp,7 and , similarly to our case , the mean onset of clina was 9.19 3.31 days . in another case series of 6 patients , onset of clina was 7.5 2.06 days , and none of the patients had cerebral infarction.6 due to the possibly high safety profile and efficacy of the treatment , it may therefore be appropriate to refer patients promptly to dsa and clina in case of development of focal neurologic impairment6 to prevent ischemic complications . transcranial ultrasound has a high specificity ( 99% ) and positive predictive value ( 97% ) for the detection of vsp in the middle cerebral artery11 and is of established value in the detection and monitoring of vsp after sah ( strong recommendation , class i ii evidence).12 13 in our case , there was a good correlation between ultrasonographic findings , clinical impairment , and dsa . future studies should investigate if tccs may play a major role in the early selection of patients who will benefit from this treatment . there are no standardized guidelines for patient selection , the start and the duration of clina , as well as for the dosage of nimodipine and heparin . clina is an expensive and resource - demanding procedure requiring specialized intensive care with a multidisciplinary team of anesthesiologists , neuroradiologists , neurosurgeons , and the availability of multimodal neurologic monitoring . wall dissection , catheter dislocation , sepsis , air emboli , and embolic infarction due to thrombotic adhesion at catheter or vessel wall are all potential complications of clina . moreover , a too high nimodipine dose may lead to hypotension , whereas inappropriate heparin dosage may lead to either thromboembolic or hemorrhagic complications . none of the earlier studies has reported complications.5 6 7 8 however , there are no conclusive data about the safety of clina because all the current evidence is based on case series that may be affected by publication bias . in conclusion , our case report adds new evidence on the feasibility , safety , and efficacy of clina . further studies are needed to clarify if transcranial ultrasound or other modalities such as mr and/or ct perfusion are able to early select which patients are likely to profit from clina . Output:	vasospasm ( vsp ) is one of the major causes for prolonged neurologic deficit in patients with aneurysmal subarachnoid hemorrhage . few case series have reported about continuous local intra - arterial nimodipine administration ( clina ) in refractory vsp . we report our experience with clina in a patient with refractory cerebral vsp .
PubmedSumm2045	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: about 2 to 3 of every 1000 children are identified with hearing loss at birth each year in the united states , and hearing impairment is , in fact , the most common sensory deficit in the pediatric population [ 14 ] . late - onset hearing loss or acquired hearing loss , in addition to congenital hearing loss , is prevalent as well in young children . for example , hearing loss associated with otitis media with effusion ( ome ) can be seen in 1540% of children under 5 years . since hearing impairment in early childhood can cause significant delays in speech / language developments , early identification and diagnosis of hearing loss become the initial and a critical step for proper treatment and habilitation , regardless of the etiology or the severity of the hearing loss . in clinical audiology , behavioral hearing evaluation is considered the gold standard for evaluating hearing sensitivity in the pediatric population . during a hearing evaluation , audiologists typically use visual reinforcement audiometry ( vra ) , conditioned play audiometry ( cpa ) , or conventional pure - tone audiometry to test children 's hearing . the technique chosen by an audiologist for a specific child is usually dependent upon the child 's age and the child 's developmental skill level . due to developmental and physical limitations , behavioral hearing test such as vra is not possible for any young children under six months of age . therefore , an electrophysiology - based hearing evaluation such as the auditory brainstem responses ( abr ) test becomes the obvious choice for this population . it can be very difficult , if not impossible , for audiologists to conduct behavioral hearing assessments on children with complicated medical conditions and those who are unable to cooperate for the hearing test . in such cases , occasionally , abr test may be requested by physicians to be performed in the operating room in conjunction with other surgical procedures , for which patients are put under general anesthesia . while studies have shown that abr test is a reliable and an objective method to assess children 's hearing sensitivity [ 610 ] , the accuracy of abr results obtained in the operating room has been questioned in previous clinical studies since the abr outcomes can be affected by many factors in the operating room . several studies in the past few years suggested that abr results , obtained in the operating room following otologic procedures such as myringotomy and tube placement , can be inaccurate , overestimating the hearing loss in children who have ome [ 1113 ] . to better understand the benefits and limitations of performing abr tests in the operating room , we conducted a comprehensive review of all abr tests conducted in the operating room at children 's hospital boston from 2007 to 2010 . in doing so , we sought to identify factors that may have some effects on the outcomes of abr test . we also would like to share our clinical experience of performing abr testing in the operating room . a retrospective study , approved by the institutional review board at children 's hospital boston , was conducted on children who had abr test completed in the operating room in a pediatric tertiary care facility from 2007 to 2010 . our database included 452 abr tests completed in the operating room during the study period . a total of 267 pediatric cases , most of them young children , were selected for a thorough review and analysis . all selected cases had sufficient information in their medical records , which include abr results from the operating room , medical and surgical procedure notes , and follow - up evaluation documents . abr testing in the operating room was performed after ear examination under the microscope by a pediatric otolaryngologist while patients were under general anesthesia . other otologic procedures , when deemed necessary , were also performed by attending physicians / surgeons before abr testing . these procedures included myringotomy , suction of middle ear effusion , and tube placement when necessary . the abr was recorded with bio - logic navigator pro evoked potential system ( natus medical inc . , san carlos , ca ) using children 's hospital boston pediatric protocol . specifically , stimuli ( clicks and tonebursts ) , calibrated in db normal hearing level , were presented at 38.1 per second via insert phones . thresholds of abr were determined by the lowest intensity level at which the wave v was present . if the wave v of abr was not identified with the highest intensity stimulus , extra recordings with both rarefaction and condensation clicks were made to investigate the presence / absence of the cochlear microphonics . follow - up audiological evaluation was conducted during return visit when patients recovered from the surgical procedures , usually within 3 months . age - appropriate behavioral test procedures , for example , vra , cpa , or conventional pure - tone audiometry , were typically performed . in some cases , a repeat of abr testing was conducted due to inability to complete behavioral tests or lack of cooperation from the patients . for each patient , frequency - specific hearing thresholds were documented and stored for future analysis . for all patients , clinical diagnosis , the types of hearing loss , and the degree of hearing loss , so forth were summarized . we were able to collect hearing results from abr completed in the operating room and postoperative examination from 121 children with ome , and comparison analysis was conducted for this subgroup of patients . the mean of thresholds at three frequencies , 1000 , 2000 , and 4000 hz , was calculated and defined as the averaged hearing threshold . analysis of variance ( anova ) and nonparametric testing ( mann - whitney u test ) was carried out using the statistical package for social science ( spss v.16.0 , spss inc . , among the 267 patients included in this study , there were 156 boys and 111 girls , with age ranging from 2 months to 18 years . the request for an abr test in the operating room was made by patients ' attending physicians or managing audiologists due to previous unsuccessful attempt of hearing evaluation and patients ' medical condition being not suitable for further testing in the clinic . most of the abr tests were carried out in the operating room while patients were under general anesthesia for other medical procedures as well . there were a variety of specific reasons for clinicians and parents to question patients ' hearing ability , summarized in table 1 . the majority ( 57.3% ) of patients had recurrent otitis media , and most of them needed surgical intervention , such as myringotomy and tube placement . in these cases , there were 36 patients ( 13.5% of all cases ) with at least one type of the neurological disorder , who needed the abr test in the operating room to find out their hearing ability . these neurological disorders included , but were not limited to , seizures , cerebral palsy , hydrocephalus , brain hemorrhage , ischemic encephalopathy , graves disease , chiari malformation , and so forth . there were 29 patients , accounted for 10.9% of all cases , who had chromosomal anomalies ( e.g. , trisomy 21 or trisomy 18 , fragile x , and other chromosomal deletions and/or duplications ) and related medical issues . sixteen patients with autistic spectrum disorders , such as autism and pervasive developmental disorders - not otherwise specified ( pdd - nos ) needed the abr test to find out if their hearing was adequate for communication . fifteen patients with infectious diseases involving their central nerve system , such as meningitis and cytomegalovirus ( cmv ) infection , had abr test in the operating room to see if their auditory function was compromised . eleven children with the diagnosis of global developmental delay ( with not - yet - identified causes ) needed the abr test to address the question whether they were able to hear adequately for verbal communication . moreover , there were four patients with head injury and temporal bone fracture who underwent abr test in the operating room to determine if hearing loss was involved . at last , 3 young children with known hearing loss underwent abr testing in the operating room to confirm the degree of hearing loss before cochlear implantation . based on the abr results , we were able to determine the degree of the hearing loss for each patient . specifically , we defined the averaged hearing threshold better than 25 db as normal hearing ; the averaged hearing threshold between 25 db and 40 db as mild hearing loss ; the averaged hearing threshold between 41 db and 60 db as moderate hearing loss ; the averaged hearing threshold 61 db and 90 db as severe hearing loss ; the averaged hearing threshold higher than 90 db as profound loss . our results found 58 patients ( 21.7% ) had normal hearing . among the other 209 children with hearing loss , 94 patients ( 35.2% ) were found with mild loss , 75 patients ( 28.1% ) with moderate loss , 21 patients ( 7.9% ) with severe loss , and 19 patients ( 7.1% ) with profound loss . in addition , we were able to identify the types of hearing loss in most of our patients . over 60% of the children identified with hearing loss had conductive loss while about 15% of them had sensorineural loss . based on the available results , the type of hearing loss could not be determined for the remaining 26 patients . the above findings are summarized in tables 2 and 3 , respectively . by reviewing patients ' medical records , we identified a number of medical conditions or events that are known to be associated with hearing loss , as summarized in table 4 . over 50% ( 109 patients ) of the patients with hearing loss there were 21 patients ( about 10% ) with syndromic diseases such as charge syndrome and waardenburg syndrome , in which hearing loss is common . congenital cmv and meningitis were diagnosed in 13 patients with hearing loss while metabolic diseases were seen in 9 children with hearing loss . there were 9 patients with hearing loss who had a history of chemotherapy and/or antibiotics treatments , which were known to be ototoxic and can cause significant hearing loss . moreover , 9 children with hearing loss previously underwent extracorporeal membrane oxygenation ( ecmo ) treatment . inner ear structural anomalies in the temporal bone , such as enlarged vestibular aqueduct and malformed cochlea , were found in 9 patients with hearing loss . other medical issues , such as neonatal hypoxia , prematurity , and hyperbilirubinemia , were noted in the remaining 33 patients with documented hearing loss . during our recordings of abr in the operating room first , the background noise level in the operating room , in some cases , was extraordinarily high due to the use of multiple medical devices , which could create elevated abr thresholds at lower frequencies such as 500 and 1000 hz . second , electromagnetic interferences from power sources , medical equipment in the operating room , and patients ' own medical devices ( such as pacemaker and vagal nerve stimulator ) could degrade abr responses , which made the abr waveforms unclear and difficult to read , resulting in overestimation of hearing loss . third , the presence of middle ear fluid could influence the hearing results from the abr testing even if the fluid was removed surgically , more significant in the cases with mucoid than in the cases with serous effusion . finally , the abr could be affected by patients ' physiological and neurological status which might be altered by anesthesia medications . since the majority of our patients had ome , we decided to do further analysis on this group to see if there were any other factors of significance . among the 153 children with suspected ome we divided them into three groups based on the procedures they received in the operating room . group 1 included 47 patients ( 69 ears ) who underwent microscopic examination with no middle ear effusion found ; group 2 included 13 patients ( 18 ears ) who underwent myringotomy only with no tube placement ; group 3 included 61 patients ( 83 ears ) who underwent myringotomy and tube placement . for patients in group 1 , the averaged hearing threshold from abr was 3.5 db ( 3.5 ) higher than the follow - up audiological evaluation . in group 2 , the averaged hearing threshold from abr was 4.0 db ( 3.5 ) higher than the follow - up audiological evaluation . in group 3 , the averaged hearing threshold of abr from the operating room was 9.4 db ( 10.6 ) higher than the follow - up audiological evaluation . comparison analysis showed that the averaged hearing threshold discrepancy between abr from the operating room and the follow - up audiological evaluation was significantly different among the three groups ( p < 0.01 ) . although the averaged hearing threshold discrepancies between the abr from the operating room and the follow - up audiological evaluation seemed to be relatively small in all groups , the results varied greatly among patients . across the three groups , there were 16 ears that had average hearing threshold difference of greater than 20 db , with the majority of these ears being patients in group 3 . in all groups , threshold discrepancy between the abr from the operating room and the follow - up audiological evaluation was notably higher at 1000 hz than that at 2000 and 4000 hz . one common goal in identification of hearing loss in children among hearing care professionals , including otolaryngologists and audiologists , is making an accurate diagnosis as early as possible so that children with hearing loss can receive appropriate treatment and habilitation to avoid delays in speech and language development . clinically , audiologists will choose age - appropriate behavioral evaluation methods to obtain hearing thresholds . abr test may be used in certain cases for difficult - to - test children to estimate their hearing sensitivity although its value remains debatable among professionals when it is used in the operating room . our experience of performing abr testing in the operating room , as shown in our study , is mostly successful , and our findings are tremendously valuable in clinical care of pediatric patients with hearing concerns . not only are we able to define the hearing status for the most difficult - to - test children with a variety of serious medical issues , but we are also able to determine the degree and the type of hearing loss for the majority of the patients . by doing so , we have provided appropriate recommendations for their much needed treatment and habilitation . at the same time , we also gained more knowledge of medical conditions and events that are linked to hearing loss . while we are confident in the justification of using abr in the operating room [ 14 , 15 ] , we remain aware of the recent clinical studies which have revealed significant discrepancies between abr results obtained in the operating room and the results obtained in the follow - up period . these studies have raised a concern regarding the validity of abr outcomes obtained in the operating room [ 1113 ] . therefore , we systematically investigated the abr results from the operating room and follow - up audiologic evaluation in a large group of cases . we found that hearing thresholds obtained from the abr tests in the operating room were significantly elevated in patients who underwent myringotomy plus tube placement . on the other hand , minimal discrepancies , smaller than 5 db , were found in the majority of patients without middle ear effusion and surgical procedures involved . our findings also suggest that the elevated abr thresholds in the operating room are most often temporary and usually resolve within a short time period after their surgery . mechanisms of the temporary shift in hearing may include ( 1 ) the existence of residual fluid in the middle ear space following the surgical procedures ; ( 2 ) the high - intensity noises created by suction , which could cause a noise - induced temporary threshold shift in hearing [ 11 , 16 ] ; ( 3 ) swelling and inflammation of the membranes in the middle ear created by suctioning of fluid and insertion of ear tubes . performing abr testing in the operating room can be , at times , a challenge . through the process of conducting hundreds of abr tests in the operating room , our audiology staff has experienced their success and difficulties in this task and our best practice have evolved . efficiency of troubleshooting adverse abr recording conditions , for example , eliminating electromagnetic inferences and minimizing factors or events that may degrade abr waveform , has improved among our audiologists . abr testing in the operating room , unlike the behavioral hearing tests , is not a direct measure of patients ' hearing sensitivity . accordingly , audiologists have to adjust their routine in explaining the audiologic results , improve their counseling skills , and provide more appropriate recommendations regarding patients ' hearing loss to their parents . at last , better coordination and communication with other medical professionals in the operating room , such as the nurses , the surgeons , and anesthesiologists , is a key for audiologists to achieve the goal of accurate hearing assessment for children with complicated medial issues . based on our study , we have concluded that abr testing can be successfully performed in the operating room when interference factors are properly addressed . in fact , hearing thresholds can be accurately estimated based on the outcomes of abr conducted in the operating room under the right circumstances , for example , in the absence of middle ear effusion or surgical procedure involved . in addition , the degrees and the types of hearing loss can be determined in most of the cases . when myringotomy and tube placement are done prior to the abr testing , follow - up hearing evaluation when the ear is cleared and healthy , either by behavioral tests or by abr testing , is necessary to determine a patient 's true hearing thresholds or to confirm the hearing loss established by the abr results from the operating room . caution should be taken when interpreting the abr results from the operating room , and appropriate parental counseling must be provided . it has to be noted that there are times in which abr test in the operating room is the only option to assess a child 's hearing ability . Output:	objectives . to review our experience of conducting auditory brainstem response ( abr ) test on children in the operating room and discuss the benefits versus limitations of this practice . methods . retrospective review study conducted in a pediatric tertiary care facility . a total of 267 patients identified with usable data , including abr results , medical and surgical notes , and follow - up evaluation . results . hearing status successfully determined in all patients based on the abr results form the operating room . the degrees and the types of hearing loss also documented in most of the cases . in addition , multiple factors that may affect the outcomes of abr in the operating room identified . conclusions . hearing loss in children with complicated medical issues can be accurately evaluated via abr testing in the operating room . efforts should be made to eliminate adverse factors to abr recording , and caution should be taken when interpreting abr results from the operating room .
PubmedSumm2046	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the german pathologist rudolph virchow was probably the first to describe the histologic features of myxofibroma in 1863 , although the lesions of jaws were not particularly mentioned . in 1947 , odontogenic myxoma is an uncommon benign mesenchymal odontogenic tumor arising from the dental papilla , follicle , or the periodontal ligament . the evidence for its odontogenic origin arises from its almost exclusive location in the tooth bearing areas of the jaws , its occasional association with missing or unerupted teeth and the presence of odontogenic epithelium . clinically , odontogenic myxoma is a benign painless , invasive , slowly enlarging mass causing marked asymmetry of the face . it usually occurs in second and third decades of life and causes expansion of bony cortices , displacement and loosening of teeth . radiographically , its appearance ranges from unilocular pericoronal radiolucency with variable trabecular pattern giving rise to soap bubble , tennis racquet , or honey comb appearance . histopathologically , the odontogenic myxoma is characterized by loose , abundant mucoid stroma that contains rounded , spindle shaped or angular cells . various surgical procedures were mentioned which includes curettage , excision ( 0.5 mm from apparent normal bony margin ) , resection ( 1 cm from apparent bony margin ) , resection with disarticulation , excision of tumor with dento - alveolarsegment and preservation of the mandibular lower boarder and maxillectomy . while generally considered a slow - growing neoplasm , odontogenic myxomas may be infiltrative and aggressive with high recurrence rates . due to poor follow - up and lack of reports , a precise analysis of recurrence rates is still missing . in view of its rarity , a 25-year - old male patient presented at st joseph dental college , eluru with painless gradually progressive swelling in the left side of the jaw for 2 years . the intraoral examination of the maxillary and mandibular arch revealed permanent dentition except for the absence of three - dimensional ( 3d ) molars in the maxillary jaw and amalgam filling of tooth 15 , 16 , 26 and 46 . the extra oral and general examinations revealed no other abnormalities . on palpation , there was diffuse , non tender swelling of 2 2 cm involving tooth 36 with the expansion of cortical plates . based on the clinical and radiographic finding , a provisional diagnosis of odontogenic cyst was made . fine needle aspiration was attempted to confirm the provisional diagnosis , but it was inconspicuous and this attempted aspiration precipitated pain and inflammation to the subject . keeping the view of provisional diagnosis and small size of the lesion , surgical excision with curettage was done under local anesthesia and the excised tissue was sent for the histopathological examination . grossly , the excised mass was brownish - white in color , without encapsulation and soft in consistency [ figure 1 ] . microscopic examination revealed haphazardly arranged stellate , spindle - shaped cells in an abundant , loose myxoid stroma that contains only few collagen fibrils , thus confirming it to be odontogenic myxoma . photograph showing the biopsy specimen of case 1 a 26-year female reported to the department of oral and maxillofacial surgery with a chief complaint of swelling on the left side of the face since 1 month . on intraoral examination , the swelling presented as an exophytic growth , soft to firm in consistency and tender . on palpation , the swelling was diffuse , firm , non - fluctuant , hard and tender . displacement of the teeth along with expansion of buccal and lingual cortical plates was evident . preoperative intraoral photograph case 2 the panoramic tomogram was taken which showed well - demarcated multilocular radiolucent lesion with honey comb or tennis racquet appearance [ figure 3 ] . hence , the provisional diagnosis of odontogenic keratocyst , ameloblastoma , central giant cell granuloma and odontogenic myxoma was made . incisional biopsy was performed under local anesthesia and the specimen was sent for the histopathological examination . on examination , the specimen showed a dense and diffuse collection of the fibers and fibroblasts and small inconspicuous strands of odontogenic epithelium . radiograph showing honeycomb or tennis racquet appearance based on the incisional biopsy report , progressive nature of the lesion and cortical expansion , radical resection with hemimandibulectomy was advised . a submandibular approach was followed under general anesthesia to expose this ill - defined mandibular mass [ figure 4 ] . after the incision through skin , dissection was carried out through subcutaneous tissues , platysma , deep fascia and finally , the pterygomassetric sling with periosteum is divided to approach the inferior border of the mandible . subperiosteal reflection of the masseter muscle is done along the lateral surface of the mandible to expose the coronoid process , sigmoid notch and condylar neck . the anterior portion of the resection is determined and dissected using the saw blade for complete osteotomy . the specimen is completely removed and the surgical area is irrigated [ figure 5 ] . reconstruction is done by placing the iliac graft , which can be appreciated in the postoperative radiograph [ figure 6 ] . the resected specimen is sent for the histopathological examination , which discovered it to be odontogenic myxoma [ figure 7 ] . intraoperative procedure opg showing reconstruction with iliac graft h and e stained section showing stellate , spindle - shaped cells in an abundant , loose myxoid stroma a 25-year - old male patient presented at st joseph dental college , eluru with painless gradually progressive swelling in the left side of the jaw for 2 years . the intraoral examination of the maxillary and mandibular arch revealed permanent dentition except for the absence of three - dimensional ( 3d ) molars in the maxillary jaw and amalgam filling of tooth 15 , 16 , 26 and 46 . the extra oral and general examinations revealed no other abnormalities . on palpation , there was diffuse , non tender swelling of 2 2 cm involving tooth 36 with the expansion of cortical plates . based on the clinical and radiographic finding , a provisional diagnosis of odontogenic cyst was made . fine needle aspiration was attempted to confirm the provisional diagnosis , but it was inconspicuous and this attempted aspiration precipitated pain and inflammation to the subject . keeping the view of provisional diagnosis and small size of the lesion , surgical excision with curettage was done under local anesthesia and the excised tissue was sent for the histopathological examination . grossly , the excised mass was brownish - white in color , without encapsulation and soft in consistency [ figure 1 ] . microscopic examination revealed haphazardly arranged stellate , spindle - shaped cells in an abundant , loose myxoid stroma that contains only few collagen fibrils , thus confirming it to be odontogenic myxoma . a 26-year female reported to the department of oral and maxillofacial surgery with a chief complaint of swelling on the left side of the face since 1 month . on intraoral examination , the swelling presented as an exophytic growth , soft to firm in consistency and tender . the mucosa overlying the swelling was pale pink in color [ figure 2 ] . on palpation , the swelling was diffuse , firm , non - fluctuant , hard and tender . displacement of the teeth along with expansion of buccal and lingual cortical plates was evident . preoperative intraoral photograph case 2 the panoramic tomogram was taken which showed well - demarcated multilocular radiolucent lesion with honey comb or tennis racquet appearance [ figure 3 ] . hence , the provisional diagnosis of odontogenic keratocyst , ameloblastoma , central giant cell granuloma and odontogenic myxoma was made . incisional biopsy was performed under local anesthesia and the specimen was sent for the histopathological examination . on examination , the specimen showed a dense and diffuse collection of the fibers and fibroblasts and small inconspicuous strands of odontogenic epithelium . radiograph showing honeycomb or tennis racquet appearance based on the incisional biopsy report , progressive nature of the lesion and cortical expansion , radical resection with hemimandibulectomy was advised . a submandibular approach was followed under general anesthesia to expose this ill - defined mandibular mass [ figure 4 ] . after the incision through skin , dissection was carried out through subcutaneous tissues , platysma , deep fascia and finally , the pterygomassetric sling with periosteum is divided to approach the inferior border of the mandible . subperiosteal reflection of the masseter muscle is done along the lateral surface of the mandible to expose the coronoid process , sigmoid notch and condylar neck . the anterior portion of the resection is determined and dissected using the saw blade for complete osteotomy . the specimen is completely removed and the surgical area is irrigated [ figure 5 ] . reconstruction is done by placing the iliac graft , which can be appreciated in the postoperative radiograph [ figure 6 ] . the resected specimen is sent for the histopathological examination , which discovered it to be odontogenic myxoma [ figure 7 ] . intraoperative procedure opg showing reconstruction with iliac graft h and e stained section showing stellate , spindle - shaped cells in an abundant , loose myxoid stroma odonogenic myxoma is a nonencapsulated benign tumor of the jaw bones , comprising around 36% of all odontogenic tumors . previous studies by white et al . , refer the tip of the incidence with an age range of 1170 years , but the majority of the cases are between 10 and 40 years of age . our two cases surpass the second decade of life , but can be addressed within the range referred in the literature . although its most frequent location is the posterior mandible , other locations such as the incisive sector , upper maxilla and mandibular condyle must be considered . present cases showed lesion in the mandible with extension in the posterior region of the mandible . pain , displacement of teeth and parasthesia are uncommon , thus the lesion can reach considerable size before the patients becomes aware of its presence and seeks treatment . our cases showed a large mass as the main aspect in physical examination , but in case one patient developed pain after aspiration . radiologically , the appearance may vary from a unilocular radiolucency to a multicystic lesion with a well defined or diffused margin . a unilocular appearance may be seen more commonly in children and in the anterior parts of the jaws . in tooth - bearing areas , the tumor is often scalloped between the roots ; differential diagnosis must be established with other clinically similar entities , such as simple cyst , ameloblastoma , intraosseous hemangioma , giant cell granuloma , osseous aneurismatic cyst and metastasis . histologically , odontogenic myxoma shows a spectrum of fibrous connective tissue stroma is present , from myxoid to dense hyalinized and from relatively acellular to cellular . calcification may or may not be present.[1014 ] the variation in the histopathological diagnosis between the initial biopsy as odontogenic fibroma and the final diagnosis as odontogenic myxoma in our case 2 could be attributed either to the biological spectrum of this lesion or non inclusion of myxomatous areas in the incisional biopsy . the tumor is not radiosensitive and treatment is by surgery . despite surgery it has been established as the treatment of choice of these tumors , no unanimous consensus exists concerning the extension of surgery . although it is our opinion that enucleation should be avoided , curettage with scarification of the cavity with acrylic bur may be used for small mandibular lesions particularly in the anterior region and especially where the patient is educated and available for periodic reviews . in the present cases , the surgical management involved an intra and extra oral approaches . prognosis after excision is excellent in this 2-year follow up period in both the cases . odontogenic myxoma is a slow growing benign , locally malignant tumor , notorious for recurrence . due to the unspecific nature of these lesions along with diagnostic and operative dilemmas related to myxomatous tumors , a sound knowledge with proper histopathological diagnosis is required for maxillofacial surgeons to improve skills in their management . Output:	odontogenic tumors represents a broad spectrum of lesions ranging from benign to malignant to dental hamartomas all arising from odontogenic residues , that is , the odontogenic epithelium , ectomesenchyme with variable amounts of dental hard tissues formed in the same sequence as in normal tooth development . we report two cases of myxoma , which were misdiagnosed initially and latter , reported as odontogenic myxoma ; and were treated by conservative surgical excision in one case and radical resection with hemimandibulectomy in the other case .
PubmedSumm2047	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: they prevent the establishment of potentially harmful pathogens and assist in improving the immune system . on the other hand , the microbiota may promote the development of allergic diseases and is a major reservoir for endogenous infections . for bacteria in the nasal habitat , the latter has mainly been demonstrated for the role of nasal staphylococcus aureus carriage in the development of nosocomial infections such as bacteraemia , sternal or orthopaedic infections , , , . recently , noncultural methods have greatly increased knowledge on the communities of microorganisms colonizing different body sites . hence , analyses of 16s rrna sequences in samples from the nares of healthy individuals demonstrated that > 80% of rrna sequences detected belonged to either actinobacteria ( e.g. cornyebacteria ) or firmicutes ( e.g. staphylococci ) . in addition , other bacterial taxa including proteobacteria ( e.g. enterobacteriaceae or pseudomonales ) , bacteriodetes or fusobacteria were also found , , . interestingly , various studies indicated that the composition of the microbiota of different human habitats is influenced by several demographic factors such as gender , age and ethnicity , but also by health - related factors like hospitalization , intake of antibiotics , antipneumococcal vaccination or presence of viral infections as well as by lifestyle - associated factors like frequency of hand washing or smoking , , , , . recognizing the value of culture - independent methods , these techniques have two major limitations . firstly , they can not assess bacterial antibiotic susceptibilities , and secondly , the overload of information on predominating phyla tends to dissolve information on the presence of important human pathogens as a result of the lack of abundance of these species . however , it is important to overcome these limitations , as recent studies have indicated changes in the occurrence of antimicrobial resistant pathogens in the healthy general population . this was demonstrated for extended - spectrum -lactamase ( esbl)- or carbapenemase - producing enterobacteriaceae and methicillin - resistant s. aureus ( mrsa ) , which were increasingly found as colonizers of the nares and the intestinal tract in association with travel activities , contact with or ingestion of contaminated food items or contact with livestock husbandries , , , , . in addition , the emergence of antimicrobial resistant gram - negative bacteria among humans has mainly been documented by studies assessing these pathogens in stool samples or rectal swabs . data on their occurrence and persistence in the nares , which might be important for the probability of transmission to the environment and to other persons , are lacking . therefore , the objective of this prospective cohort study was to assess the occurrence of nasal colonization of important facultative pathogenic bacteria including s. aureus , enterobacteriaceae and nonfermentative bacteria in samples from a large number of nonhospitalized volunteers . in addition , information on associated antimicrobial resistance and lifestyle conditions was obtained and correlated with the nasal bacterial colonization pattern . nonhospitalized adult participants were recruited in ambulatory departments of public health offices ( offering , e.g. , travel vaccination or health checks ) or were employees of these offices ; dental practices ; a family physician 's practice ; and among students at colleges or universities . at the beginning of the study , participants were informed about the study 's objectives and provided written informed consent . ethical approval was obtained by the ethical commission of the westphalian wilhelms - university mnster ( 2006 - 268-f - s ) . samples were taken at three different time points ( june august 2011 , january march 2012 , august october 2012 ) , each separated by at least 4 months . at every time point , participants were asked to provide a nasal swab and to answer a standardized questionnaire . answers of the participants were directly assessed in a protected data - collection application on a portable usb flash drive . from all participants , premoistened nasal swabs ( floqswabs ; copan , murrieta , ca , usa ) were used to sample the anterior nares . after nonselective enrichment in mueller - hinton broth ( 24 hours , 36c ) , swabs were streaked onto columbia blood agar and macconkey agar as well as esbl and s. aureus screening agars ( all oxoid , wesel , germany ) and incubated for 24 hours at 36c . all colonies suspicious for s. aureus , enterobacteriaceae or clinically important nonfermenters ( i.e. acinetobacter baumannii , pseudomonas spp . , all isolates were tested by matrix - assisted laser desorption / ionization time - of - flight mass spectrometry ( maldi - tof ) . antimicrobial susceptibility testing was performed using vitek2 automated systems ( and for some nonfermenter species using agar disc diffusion ) with clinical breakpoints and application of expert rules as recommended by the european committee on antimicrobial susceptibility testing ( eucast ) . all s. aureus isolates were analysed for the presence of the nuc and meca genes . for s. aureus isolates that were cefoxitin resistant , mecc was tested in addition as previously described . the questionnaire assessed the following data for each participant : year of birth , sex , zip code for place of residence , country of birth , employment in healthcare institutions , employment with contact to livestock animals , number of other persons living in the same household , living with companion animals in the same household , being a smoker , having regular ( i.e. at least weekly ) contact with medical personnel , having regular ( i.e. at least weekly ) contact with persons with employment in livestock production , and travelling abroad ( if yes , which countries ; within the past 12 months ) . in addition , participants were asked whether they had diabetes mellitus , allergies , chronic skin diseases ( e.g. psoriasis , atopic dermatitis , acne ) , chronic diseases of the liver or the airways ( e.g. asthma , chronic obstructive pulmonary disease ) , chronic renal insufficiency , chronic inflammatory diseases of the paranasal sinuses or the bowel , chronic osteomyelitis , solid tumors or haemato - oncologic diseases . participants provided information on whether they were immunocompromised ( if yes , why ? ) , whether they had implanted devices ( e.g. heart valves , prostheses ) . moreover , information on hospitalization ( within the past 12 months ) was obtained . participants were asked whether they received antibiotics or antimycotic agents ( if yes , oral / intravenous or topical ) , oral contraceptives or medication against clinical depression or neuroleptic agents . data on density of livestock and human population in different districts as well as geographical data were assessed from the german regional database ( https://www.regionalstatistik.de/ ; codes 116 - 33 - 4 , 173 - 21 - 4 , 449 - 01 - 4 ) . statistical analysis was performed by spss 22 ( ibm , armonk , ny , usa ) . chi - square or fisher 's exact tests were used to calculate differences between categorical variable ; t test was used for parametric variables . p < 0.05 was considered significant . for assessing independent risk factors associated with nasal colonization of different groups of bacteria , we applied multivariable logistic regression by stepwise backward selection of variables and probabilities of 0.05 for entry into and 0.01 for removal from the model ( spss 22 ) . all variables associated with p values of 0.2 were entered into the initial regression model . overall , 1878 persons participated in the study . among these , 1168 ( 62.2% ) participated at all three time points , 283 ( 15.1% ) at two time points and 427 ( 22.7% ) only once ( table 1 ) . the majority of participants were female ( 58.4% ) , and the median ( mean ) age was 45 years ( 45.1 years ) ( range , 797 years ) . 1 shows that all but six of the participants lived in a total of 38 districts in the german federal states of north rhine westphalia and lower saxony . of all 1878 participants , 1203 ( 64.0% ) were colonized with at least one of the bacterial pathogens assessed at least at one time point . the overall prevalence of nasal s. aureus carriage was 41.0% ( n = 768 ) ; 33.4% of the participants were nasally colonized with enterobacteriaceae ( n = 628 ) and 3.7% with nonfermenters ( n = 69 ) ( fig . 2 ) . at the species level , klebsiella oxytoca ( 6.4% ) , escherichia coli ( 6.1% ) , proteus mirabilis ( 6.0% ) , citrobacter koseri ( 3.6% ) , pantoea agglomerans ( 3.1% ) , enterobacter aerogenes ( 2.6% ) , klebsiella pneumoniae ( 2.4% ) , citrobacter freundii ( 2.2% ) , enterobacter cloacae ( 1.4% ) , acinetobacter baumannii ( 1.1% ) , raoultella ornithinolytica ( 1.1% ) and serratia marcescens ( 1.0% ) were detected in more than 1% of the 1878 participants . among those participants carrying enterobacteria , n = 515 were colonized with one enterobacterial species , 96 with two , 16 with three , and one with four different species , respectively . of the 69 participants colonized by nonfermenters , all carried one nonfermentative species only . statistically significant differences in the co - colonization patterns were mainly found for s. aureus carriers , who were less frequently colonized with various enterobacteria and nonfermenters ( fig . table 2 shows the persistence of nasal colonization as indicated by detection of the bacteria at the three different time points of investigation . focusing on those participants who provided three nasal swabs at time points separated by four to six months each , the persistence of colonization , which was firstly detected at the initial time point of the study , was significantly ( p < 0.001 ) different for carriers of s. aureus ( n = 525 ; 63.8% colonized for more than four months and 39.2% for more than eight months ) , enterobacteriaceae ( n = 440 ; 54.7% and 32.1% ) and nonfermenters ( n = 55 ; 5.5% and 0% ) . at the three time points , we obtained a total of 1378 s. aureus isolates ( all nuc positive ) , 1169 enterobacteriaceae isolates ( from 28 species ) and 73 nonfermenter isolates ( from nine species ) . enterobacteriaceae mostly belonged to proteus mirabilis ( n = 211 isolates ; 18.0% of all 1168 enterobacteriaceae isolates ) , e. coli ( n = 180 ; 15.4% ) , k. oxytoca ( n = 175 ; 15.0% ) , c. koseri ( n = 138 ; 11.8% ) , e. aerogenes ( n = 86 ; 7.4% ) , c. freundii ( n = 64 ; 5.5% ) , p. agglomerans ( n = 63 ; 5.4% ) , k. pneumoniae ( n = 61 ; 5.2% ) , e. cloacae , r. ornithinolytica , hafnia alvei , and s. marcescens ( each n = 27 ; 2.3% ) . among all nonfermenter isolates , the following species were detected : a. baumannii ( n = 24 isolates ; 32.9% of all nonfermenter isolates ) , stenotrophomonas maltophilia ( n = 14 ; 19.2% ) , pseudomonas putida ( n = 12 ; 16.4% ) , pseudomonas aeruginosa ( n = 8 ; 11.0% ) , pseudomonas fluorescens ( n = 6 ; 8.2% ) , pseudomonas stutzeri ( n = 5 ; 6.8% ) , pseudomonas spp . ( n = 2 ; 2.7% ) , pseudomonas monteilii ( n = 1 ; 1.4% ) and achromobacter xylosoxidans ( n = 1 ; 1.4% ) . the antibiotic susceptibility test results of every first isolate detected per participant are shown in table 3 ; data are shown for those enterobacterial and nonfermenter species , which were detected in more than ten participants , respectively . resistance against linezolid , vancomycin and tigecyclin thirteen participants ( 0.7% ) were colonized with mrsa , as defined by cefoxitin - resistant isolates harbouring meca . this represents a proportion of 1.7% mrsa on all 768 s. aureus first isolates . in addition , s. aureus isolates from two participants ( 0.1% ) were cefoxitin resistant , but neither meca nor mecc was detectable , and hyperproduction of -lactamases was excluded . for these isolates , the vitek2 esbl screening test was negative for all e. coli and klebsiella spp . overall , participants ' first isolates of enterobacteriaceae ( all species ) were resistant to ampicillin , cefuroxime , cefotaxime , ciprofloxacin , trimethoprim sulfamethoxazole and gentamicin in 67.0 , 11.9 , 1.6 , 0.4 , 2.5 and 2.4% , respectively . supplementary tables s1 , s2 and s3 show the association between the risk factors assessed and nasal colonization ( at any time point ) with s. aureus , enterobacteriaceae and nonfermenters based on univariate analysis and logistic regression . we found that household sizes of three ( odds ratio ( or ) 1.420 , 95% confidence interval ( ci ) 1.1091.818 ) or more than four persons ( or 1.349 , 95% ci 1.0831.682 ) , living in regions with > 100 livestock animals per km ( or 1.251 , 95% ci 1.0011.562 ) , atopic dermatitis ( or 1.706 , 95% ci 1.0252.841 ) and asthma ( or 1.501 , 95% ci 1.0152.219 ) were independently associated with s. aureus carriage , while female gender ( or 0.752 , 95% ci 0.6210.911 ) , smoking ( or 0.726 , 95% ci 0.5880.896 ) , colonization with enterobacteriaceae ( or 0.644 , 95% ci 0.5250.789 ) and unreported household size ( or 0.259 , 95% ci 0.0750.886 ) were significantly less frequent among s. aureus carriers . performing a separate analysis for participants colonized with meca - positive mrsa showed that only occupational contact with livestock animals ( 6/13 vs. 55/1865 ; p < 0.001 ) and taking antidepressant / neuroleptic drugs ( 4/13 vs. 125/1865 ; p 0.009 ) were significantly associated with mrsa in univariate analysis . a regression model initially containing the risk factors associated with p values of < 0.2 revealed that occupational livestock contact ( or 31.036 ; 95% ci 8.480113.585 ) , chronic renal insufficiency ( or 40.611 ; 95% ci 3.735441.627 ) , taking antidepressant / neuroleptic medication ( or 4.974 ; 95% ci 1.14021.701 ) and chronic obstructive pulmonary disease ( or 28.111 ; 95% ci 1.808437.103 ) were independent risk factors for mrsa . nasal colonization with enterobacteriaceae ( supplementary table s2 ) was associated with age of 7180 years ( or 1.505 , 95% ci 1.0021.78 ) , travel to the united states ( or 2.022 , 95% ci 1.1173.660 ) and topical antimycotic therapy ( or 2.933 , 95% ci 1.5855.428 ) . the factors s. aureus carriage ( or 0.671 , 95% ci 0.5470.822 ) , smoking ( or 0.596 , 95% ci 0.4750.748 ) and hemato - oncologic diseases ( or 0.099 , 95% ci 0.0130.760 ) , living in regions with low population densities of < 500 inhabitants per km ( or 0.739 , 95% ci 0.5940.919 ) and with rather low livestock densities of 5099 animals per km ( or 0.779 , 95% ci 0.6280.966 ) were significantly less frequent among participants colonized with enterobacteriaceae . s3 ) was independently associated with living in areas with a livestock density of < 50 animals per km ( or 2.114 , 95% ci 1.2763.504 ) , travel to the united states ( or 3.756 , 95% ci 1.4189.946 ) and oral antibiotic therapy ( or 1.729 , 95% ci 1.0182.937 ) . for both enterobacteriaceae and nonfermenters , the univariate analysis of lifestyle - related risk factors was repeated for colonization , with each of the different bacterial species included in these two groups of bacteria as the independent variable ( supplementary table s4 ) . in this prospective study , we assessed nasal colonization with important human pathogens among a cohort of > 1800 individuals from the general population . because of the design of the study , and because the study was carried out in only two federal states , the participants involved are not fully representative for the german general population . however , our approach of recruiting volunteers resulted in the formation of a sufficiently heterogeneous cohort . comparing the demographic data assessed with reference data for the german population ( federal statistical office of germany ( destatis ) ; https://www.destatis.de/ ; data based on 20112012 census ; date of data retrieval , 11 september 2014 ) , we can assume that basic parameters such as gender ( female ; census / cohort : 52%/58% ) , mean age ( census / cohort : 44 years/45 years ) or migration background ( census : 20% ; german nationality 91.7%/cohort : 12.6% birth country other than germany ) are comparable . also , working in the healthcare sector ( 10.9% of all participants ) , which could significantly influence colonization , was representative for the german general population , where according to destatis data about 12% of the population were employed in the healthcare sector in 2011 . surprisingly , we found that 33.4 and 3.7% of all participants were colonized with enterobacteriaceae and nonfermenters , respectively . data on studies investigating nasal carriage with gram - negative rods are rare because most reports focus on colonization of the intestinal tract . in switzerland , gram - negative rods ( klebsiella , e. coli , e. cloacae , citrobacter diversus ) were isolated from nasal samples obtained from 117 ( 38% ) of 534 healthy men . in a cohort of german and eritrean children , , more than half of the number of the individuals harboured isolates belonging to the enterobacteriaceae family . for k. pneumoniae , some studies even showed that colonization rates in the nares or in the respiratory tract were higher than in stool samples . hence , our data support that a major part of the general population is nasally colonized with enterobacteriaceae . antimicrobial susceptibility testing revealed that among nasal e. coli , k. pneumoniae and k. oxytoca isolates there was hardly any resistance against second- ( 9398% susceptible ) or third - generation cephalosporins ( 99100% susceptible ) . this finding is interesting , as clinical isolates of these species are resistant in higher proportions according to german national surveillance data , where cefuroxime / cefotaxime were susceptible in 88.3/92.4% of e. coli , 81.3/90.8% of k. pneumoniae and 87.5/95.3% of k. oxytoca isolates ( https://ars.rki.de/ ; reference data for ambulatory care , 2013 ) . moreover , large studies recently indicated the emergence of resistance against third - generation cephalosporins ( associated with esbl production ) in e. coli and klebsiella isolates obtained from persons in the general population . in germany , testing of stool samples from healthy individuals and outpatients revealed esbl colonization rates of 4.16.3% , . several investigations demonstrated that rectal colonization with esbl - producing enterobacteriaceae was associated with travel . in the netherlands , the prevalence of esbl genes detected in stool samples of travellers increased from 0 before to 33.6% after travel ( mainly to southeast asia ) . in a scandinavian study , risk factors for esbl carriage were travel to the indian subcontinent ( or 24.8 ) , asia ( or 8.63 ) and northern africa ( or 4.94 ) . coli identified in samples obtained < 72 hours after admission to a german hospital , asian mother tongue ( or 13.4 ) was a significant risk factor . as we detected no esbl production in any of the e. coli , k. pneumoniae and k. oxytoca isolates tested in this study , we assume a divergent antimicrobial resistance pattern between enterobacteriaceae from the nasal and the intestinal habitats with respect to esbl - producing organisms . the reasons for this are unclear , and it is a limitation of this study that it was not possible to analyse faecal and nasal samples in parallel , which would have enabled direct comparability of these two reservoirs . hypothetically , the density and variability of enterobacterial species in the intestinal tract could facilitate exchange of resistance determinants . alternatively , transmission of esbl - producing bacteria could predominantly follow orofaecal transmission pathways , leading to colonization of the intestinal tract rather than droplet or direct smear infection or cross - transmission via the hands enabling nasal colonization . nevertheless , our findings may have an impact on defining active surveillance strategies for prevention of infections with multiresistant enterobacteriaceae . there are only few truly population - based studies on the prevalence of colonization with s. aureus . in the united kingdom , a review of longitudinal studies found that about 20% ( range 1230% ) of individuals persistently carry s. aureus , while about 30% are intermittently colonized ( range 1670% ) , and about 50% ( range 1669% ) are noncarriers . interestingly , we found significantly lower rates of persistent nasal carriage for enterobacteriaceae and for nonfermenters , although 54.5% of participants colonized with enterobacteriaceae were carriers for more than 4 and 32.1% for more than 8 months . however , for drawing more exact conclusions on long - term persistence , genotyping of all isolates would be necessary in order to differentiate between repetitive short - term contamination ( with several bacterial clonal lineages ) and long - term colonization ( with one clonal lineage ) . thirteen participants ( 0.7% ) were colonized with meca - positive mrsa . unlike the united states , where the prevalence of mrsa carriage in the population was estimated to be 1.5% in 2004 , mrsa colonization in the german general population has rarely been assessed systematically . various studies interestingly , we detected two participants who carried cefoxitin - resistant , nuc - positive s. aureus , which did not harbour the resistance determinants meca or mecc . analysis of risk factors confirmed other studies indicating that s. aureus colonization is more frequent among male subjects , , , and some investigators hypothesized that taking hormonal contraceptives could have a preventive effect . although the latter risk factor was not significantly associated with s. aureus in our cohort , factors such as larger household size , atopic dermatitis and asthma were more frequent among s. aureus carriers compared to other participants , which is in line with previous investigations , , , . we found that smokers were less frequently carriers of s. aureus , putatively due to the bactericidal activity of the smoke , which confirms other studies , . in addition , we showed that nasal colonization with enterobacteriaceae also prevented ( or 0.64 ) s. aureus carriage , which was previously described for the competitive effects of corynebacteria and coagulase - negative staphylococci . the geographic area of this study ( as defined by the participants ' places of residences ) comprises both rural and urban parts of the german federal states of north rhine westphalia and lower saxony ( fig . 1 ) . the northwestern areas are represented by a low density of inhabitants and a high density of livestock ( especially pigs ) . persons with occupational livestock contact had a 31 times higher risk for mrsa carriage compared to the rest of the cohort . it has been shown that livestock - associated mrsa plays a major role among persons in this part of germany , where up to 29% of human mrsa carriers in hospitals were colonized with mrsa molecular types belonging to the clonal lineage cc398 as defined by multilocus sequence typing , , , which is predominant in european livestock populations . while the risk of acquiring mrsa cc398 via direct livestock contact has been demonstrated in various studies ( reviewed in ) , recent reports from the netherlands indicated that mrsa cc398 is increasingly acquired independently from livestock exposure by persons in the general population living in pig - dense regions . in this context , the results of this study clearly show that this phenomenon does not affect major parts of the population , as only 0.7% of the participants were colonized with mrsa . taking antidepressant / neuroleptic drugs was another risk factor for mrsa carriage in this study . this raises the question of whether or to what extent pharmaceutical agents contribute to the selection pressure on antibiotic - resistant bacteria . we are not aware of any studies assessing risk factors for nasal colonization with enterobacteriaceae or nonfermenters in the general population . in this study , in addition , factors such as topical antimycotic therapy , age of more than 70 years and oral antibiotic therapy , as well as certain characteristics of the place of residence had an impact of colonization with enterobacteria and nonfermenters . in contrast , our results that s. aureus carriage , smoking and hemato - oncologic diseases were significantly less frequent among participants colonized with enterobacteriaceae could be due to previous antibiotic consumption , the bactericidal effects of smoking and bacterial competition in the nasal habitat . overall , this prospective cohort study provided insight into nasal colonization patterns with s. aureus , enterobacteriaceae and nonfermenters in the german general population as well as in associated antimicrobial resistance profiles . however , mrsa carriage was detected in only 0.7% of the participants . although colonization rates with enterobacteriaceae were unexpectedly high ( overall 33% , with k. oxytoca , e. coli , citrobacter spp . and pantoea spp . being predominant ) , antimicrobial resistance of enterobacteriaceae was infrequent , and colonization with esbl - producing enterobacteriaceae was not detected . Output:	the nares represent an important bacterial reservoir for endogenous infections . this study aimed to assess the prevalence of nasal colonization by different important pathogens , the associated antimicrobial susceptibility and risk factors . we performed a prospective cohort study among 1878 nonhospitalized volunteers recruited from the general population in germany . participants provided nasal swabs at three time points ( each separated by 46 months ) . staphylococcus aureus , enterobacteriaceae and important nonfermenters were cultured and subjected to susceptibility testing . factors potentially influencing bacterial colonization patterns were assessed . the overall prevalence of s. aureus , enterobacteriaceae and nonfermenters was 41.0 , 33.4 and 3.7% , respectively . thirteen participants ( 0.7% ) were colonized with methicillin - resistant s. aureus . enterobacteriaceae were mostly ( > 99% ) susceptible against ciprofloxacin and carbapenems ( 100% ) . extended - spectrum -lactamase producing isolates were not detected among klebsiella oxytoca , klebsiella pneumoniae and escherichia coli . several lifestyle- and health - related factors ( e.g. household size , travel , livestock density of the residential area or occupational livestock contact , atopic dermatitis , antidepressant or anti - infective drugs ) were associated with colonization by different microorganisms . this study unexpectedly demonstrated high nasal colonization rates with enterobacteriaceae in the german general population , but rates of antibiotic resistance were low . methicillin - resistant s. aureus carriage was rare but highly associated with occupational livestock contact .
PubmedSumm2048	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: geolocation and remote sensing technologies are increasingly being applied to the mapping and spatial analysis of infectious diseases , including lymphatic filariasis and malaria . for such maps to reflect the real pattern of infection or disease , lymphatic filariasis and malaria are both currently subject to renewed control programmes , and share anopheline vectors in some parts of the world . filariasis is being targeted for elimination , but reduction in its infection intensity could conceivably , via the removal of infection - induced vector mortality , lead to more efficient transmission of malaria . accurate mapping of the two infections can help monitor their control , including the detection of any unwanted interactions . here , we present spatial analysis of both parasites in the same area of papua new guinea , using a recently developed technique which estimates the scale of variation , taking into account the typically highly skewed distribution of parasite counts . in a rural area of the east sepik province , cross - sectional parasitological surveys were done of the population aged over five years , as part of a trial of diethylcarbamazine ( dec ) plus ivermectin versus dec alone against lymphatic filariasis . although , with regard to malaria , the lower age threshold is a limitation , in the nearby wosera area , peak prevalence was not reached till after five years . wuchereria bancrofti microfilariae were counted microscopically on nuclepore filters , through which 1 ml of night - collected blood had been passed . anopheles mosquitoes , predominantly anopheles punctulatus s.s . , are the vectors of filariasis in the area . villages are divided into subunits , which we call hamlets , distinguished by names in the local languages . these were mapped with a hand - held trimble ensign global positioning system ( gps ) machine . in the 1994 survey , immediately before the first round of treatment , blood samples were obtained from 2,219 people in 149 hamlets . hamlets whose distance to their nearest neighbour was , due to gps inaccuracy , measured as less than 10 m were combined with their nearest neighbour . there were two such instances , so reducing the total number of hamlets to 147 . as described elsewhere , spatial structure in the infection densities was fitted by a negative binomial model . the mean density is fitted as a log - linear function of age , sex , and the hamlet mean . in turn , the hamlet means are given a spatial structure , with closer hamlets being more highly correlated . specifically , hamlet i adds a ' hamlet effect ' ui to the logarithm of the mean density . when exponentiated , these are similar to standardized mortality ratios ( smrs ) although relate to parasite densities rather than death rates , so can be called standardized parasite density ratios . these age- and sex - adjusted hamlet effect uil have a covariance matrix ( 1/)exp(-dil/ ) , where dil is the distance between hamlet i and l. the parameter measures the scale of spatial correlation . more specifically , loge2 is the distance over which the correlation reduces by half , which we call the ' half - distance ' . the process can also be thought of a smoothing process of the raw hamlet means , adjusting for age and sex , and with a large spatial scale ( ) corresponding to a greater degree of smoothing . the model was fitted by programs written in the c and fortran languages , and checked in terms of ability to represent the spatial variability , and robustness to distributional assumptions . at the individual level , the mean pre - treatment microfilaraemia density ( including zeros ) increased with age , till reaching 1591 mf / ml in the 4049 year age group , then flattening out . the mean p. falciparum density was 18 asexuals per 200 white cells in those aged 510 years , decreasing to below 2 for ages over 30 years . at the hamlet level , the median of the crude hamlet - specific pre - treatment mean microfilarial densities was 552 mf / ml , range 04625 . for p. falciparum , the median was 2.2 asexual parasites per 200 white cells , range 0113 . the figure shows the spatial variation in the pre - treatment density of both parasites in this area , estimated from the model , taking into account age and sex effects , and spatial correlation . w. bancrofti had an estimated half - distance of 1.7 km ( = 2.41 , = 1.79 ) . p. falciparum showed virtually no spatial correlation , the half - distance being just 14 m ( = 0.0208 , = 1.13 ) , around the resolution limit of the gps . spatial variation of a ) w. bancrofti and b ) p. falciparum in parts of urim and urat census districts , east sepik province . the scale is in kilometres with an arbitrary origin near the centre of the study area ( 142.7e , 3.6s ) . each hexagon represents one hamlet , with the shading showing the quartile of the standardized parasite density ratio . for w. bancrofti , the 0 , 25 , 50 , 75 and 100th percentiles are 0.27 , 0.56 , 1.02 , 1.71 and 4.3 respectively . for p. falciparum the curve plotted against the vertical axis shows the rate at which correlation in mean parasite density decays with distance . the solid lines in the body of the map show unpaved roads , and the dashed line indicates the division of villages between the two census districts . one year after the first round of mass treatment with anti - filarial drugs ( coverage 88% ) , all 14 randomized clusters showed a reduction in williams mean w. bancrofti density , ranging from 47 to 96% , mean 80% . all 14 clusters showed an increase in williams mean p. falciparum , the increases ranging from 2 to 255% , mean 91% . however , although dec+ivermectin was more effective against w. bancrofti , no tendency was seen for those clusters to have larger increases in p. falciparum : in 3 of the 7 pairs the dec+ivermectin cluster had the larger increase , with 4 having the opposite tendency . to further investigate the possibility that this increase was due to the treatment , the results of the spatial models were put into a hamlet - level analysis : an ' ecological ' analysis in epidemiological terms . the difference in the post- and pre - treatment hamlet effects was calculated for both filariasis and malaria , and a linear relation tested using the pearson correlation coefficient . there was negligible correlation between the changes in the densities of the two infections ( r = 0.026 , p = 0.75 ) . the world health organization methodology of ragfil ( rapid assessment of the geographical distribution of bancroftian filariasis ) is based on a 50 50 km grid . gyapong et al . have compared this to a finer 25 km grid in ghana , with both leading to operationally similar conclusions . when extended to three other countries of west africa , the method showed filariasis to have an unexpectedly wide geographical distribution . the current results suggest that , at the opposite end of the spatial scale , it may be possible for foci to persist within the interstices of a 50 50 km grid , or even a 25 25 km one ; a conclusion similar to that reached by srividya et al . in india . however , the overall importance of any such effects will also depend on the relative magnitudes of variation at different scales , which we are unable to measure beyond the extent of our current study area . the need for accurate geographical monitoring may increase if the current elimination campaign reduces filariasis over a wide scale and proceeds from an ' attack ' to a ' consolidation ' phase ( to borrow malaria eradication terminology ) . for example , thompson et al . , found the risk of malaria varying by a factor of 6 over 500 m . however , the scale of variation found in the current study was still smaller than expected , with even the closest hamlets showing little correlation . the much smaller scale of variation of malaria is probably related to its rapid variation over time . by comparison , the risk of acquiring w. bancrofti infection per mosquito bite seems to be much smaller , but infections can last much longer . a single cross - sectional survey therefore reflects a longer - term aggregate of filariasis exposure history . our estimation of the hamlet effects can be seen as a smoothing method , in which the degree of smoothing is determined by the data . in this example , the west - east trend in filariasis density was not easily discernible in the raw data , while the low spatial correlation of malaria meant that the smoothed map is similar to the crude one . any pattern presumably reflects spatial variation in factors which affect the parasite , vector or host , but which are not included in the model . in the current study , such factors are likely to include those related to vector density , such as distance to breeding sites , but this information is not available for the whole study area , and the lack of pattern in malaria infection suggests that other factors must also be involved . we view the technique as potentially useful in identifying such factors , as well as identifying ' hot spots ' of infection . our method takes account of the extreme skewness shown by most parasite distributions , which can cause problems even at the exploratory stage of analysis . for example , we found that using the common log(x+1 ) transformation to plot the hamlet ' geometric means ' induced a spurious spatial correlation in malaria infection . excess mosquito mortality caused by w. bancrofti would make it feasible for filariasis control to enhance malaria transmission . an ' ecological ' analysis did not show such an effect in this study , but monitoring should continue in those areas where anopheles are vectors of both infections . for the elimination of filariasis , mapping at small as well as large scales will be necessary , including urban areas , especially as efforts proceed beyond the ' attack ' phase . dec diethylcarbamazine gps global positioning system ragfil rapid assessment of the geographical distribution of bancroftian filariasis smr standardized mortality ratio the work originated in a drug trial of which james kazura and michael alpers were the co - principal investigators , and which was led in the field by phil hyun and zachary dimber . neal alexander worked on the drug trial and , for this paper , made the gps readings and wrote the first draft . moses bockarie also contributed to the trial , including ensuring that the blood slides were read . the spatial model was initially developed by neal alexander under the supervision of bryan grenfell . the model was finalized by neal alexander , rana moyeed and julian stander , and fitted by programs written by rana moyeed and julian stander . the drug trial was supported financially by the world health organization ( grant number tdr 910466 ) and the government of papua new guinea . julian stander was partially supported by the european union tmr network erb - fmrx - ct96 - 0095 . bryan grenfell was supported by the wellcome trust and the biotechnology and biological sciences research council . Output:	the spatial variation of wuchereria bancrofti and plasmodium falciparum infection densities was measured in a rural area of papua new guinea where they share anopheline vectors . the spatial correlation of w. bancrofti was found to reduce by half over an estimated distance of 1.7 km , much smaller than the 50 km grid used by the world health organization rapid mapping method . for p. falciparum , negligible spatial correlation was found . after mass treatment with anti - filarial drugs , there was negligible correlation between the changes in the densities of the two parasites .
PubmedSumm2049	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: there has been a lot of debate not only about how dcsg sorting occurs but also about exactly where in the cell this triage takes place . all cells have the capacity to rapidly secrete proteins after their transit through the constitutive secretory pathway . a great deal of evidence supports the view that in the appropriate cell type , dcsg sorting signals can redirect proteins from the constitutive secretory pathway to dcsgs , thus confirming that it is not a default secretory pathway . some groups have proposed that dcsg sorting occurs through the action of a sorting receptor present in the tgn that latches onto granule - destined proteins at sites where nascent granules will bud ( chung et al . , 1989 ; cool et al . , 1997 this has been referred to as the sorting by entry model . on the other hand , convincing evidence has been presented that in cells that generate dcsgs , all of the contents of the tgn are initially encapsulated into the nascent granules ( arvan and castle , 1998 ) . model proposes that those proteins destined to be secreted constitutively are progressively extruded in low - density vesicles as the granule matures , ultimately leaving only the correct cargo protein in the mature dcsg . the first truth is that , regardless of the site at which sorting occurs , a mechanism has to exist that establishes and then maintains the segregation of dcsg cargo proteins from those that are constitutively secreted . thus , it is a reasonable postulate that some mechanism exists to anchor the appropriate cargo proteins to the dcsg as it forms or matures . a second truth is that the sorting of proteins to dcsgs is a prerequisite for certain posttranslational processing steps in hormone and protease activation . for example , the conversion of proinsulin to active insulin , the conversion of proopiomelanocortin ( pomc ) to its many peptides , including acth , and the proteolytic activation of prorenin to renin all occur only after the precursors are encapsulated in the nascent secretory granules ( orci et al . , 1986 ; taugner et al . , 1987 this makes sense for the organism because it ensures that the secretion of the active hormones or proteases is under appropriate physiological control . however , for granule - restricted activation to occur , it is necessary that both the protein precursors and the appropriate processing enzymes end up in the same dcsg . in the case of proinsulin , this means that the proprotein convertases pc1/3 and pc2 , as well as carboxypeptidase e ( cpe ) , all of which are required for generation of active insulin , have to be cotargeted with proinsulin in the budding granules . thus , a second postulate is that a mechanism exists to ensure efficient cotargeting of protein precursors and their processing enzymes in the same organelle . dcsgs also share , by definition , the distinguishing trait of a core that appears dark or dense in electron micrographs . however , in spite of this common appearance , there may be important functional and mechanistic differences in dcsgs . for example , the gonadotropes of the pituitary store luteinizing hormone and follicle - stimulating hormone in separate dcsgs , and their release is independently controlled ( for review see dannies , 2001 ) . likewise , there are two types of dcsg in chromaffin cells containing either epinephrine or norepinephrine , and these are morphologically distinct ( hendy et al . , 2006 ) . the signals for targeting proteins to dcsgs also show tissue - specific variations ; removal of 90 amino acids from the c terminus of the granin chromogranin a ( cga ) prevents its sorting to dcsgs in pituitary gh4 cells , but has no effect on dcsg sorting in sympathoadrenal pc12 cells ( cowley et al . likewise , pomc is efficiently stored in dcsgs when transfected into cultured pituitary cells , but not in sympathetic neurons ( marx et al . , 1999 ) . thus , a third truth is that not all dcsgs are alike , and it is a reasonable postulate that dcsgs can be assembled , even within the same cell , through more than one mechanism . could some of these truths explain the difficulties in reaching consensus on the protein signals necessary for dcsg targeting ? there has been no shortage in the variety of dcsg sorting mechanisms proposed in the last 20 yr ; these include protein domains that interact with or that traverse membranes and that may or may not interact with additional proteins on the cytoplasmic side of the dcsg , proteins proposed to be a master switch for granule formation , universal granule cargo receptors , protein domains that mediate aggregation in the late tgn , certain paired basic protease cleavage sites or helices in secretory proteins , disulfide - constrained loops , acidifying proton pumps , and other mechanisms . as a result , investigators have become progressively entrenched in defending their favorite mechanisms and commonly use the descriptors controversial and difficult to repeat to describe the work of others in their publications . nevertheless , it is possible to accommodate most of these findings in a model that subdivides targeting function into three components ( fig . 1 ) : membrane associated ( or traversing ) tethers , tether - associated cargo , and aggregation . many dcsg cargo proteins also aggregate to form the dense core ( blue ) , and these aggregates may contain more than one protein . the yellow boxes indicate the various protein domains or mechanisms that have been implicated in dcsg sorting . note that some proteins ( such as insulin ) may have multiple dcsg sorting mechanisms . peptidyl--amidating monooxygenase ( pam ) , phogrin , and muclin are all type 1 membrane - spanning proteins that are targeted to dcsgs ( bell - parikh et al . , 2001 ; wasmeier et al . , 2002 , the granule sorting domain is located in the cytoplasmic tail of the protein , and although the exact nature of the signal is still debated , it appears that this domain can bind the clathrin adaptor proteins ap-1 and -2 in vitro ( wasmeier et al . such interactions might provide a means of communication between the granule cargo proteins and the membrane domains or cytoplasmic proteins that will define the budding dcsgs . the membrane - binding domains of the granule - resident protein cpe ( cpe ; dhanvantari et al . , 2002 ) and the prohormone convertases pc1/3 ( jutras et al . , 2000 ; arnaoutova et al . , 2003 ) and pc2 ( assadi et al . , 2004 ) are also key for their targeting to dscgs , and there is agreement that the granule sorting is mediated by short helical domains . an helical domain has also been shown to be important for targeting prosomatostatin ( mouchantaf et al . , 2001 ) , cga ( taupenot et al . , 2002 ) , and vgf ( garcia et al . , 2005 ) to dcsgs . recent results suggest that helices with the ability to direct granule sorting in secretory proteins share the characteristic of charge segregation from a hydrophobic patch , which is consistent with a shallow membrane interaction ( dikeakos et al . , 2007 ) . this first group of dcsg proteins could therefore be said to be tethered to the membranes of the tgn or the maturing granule . a second group of granule sorting domains may act by binding cargo proteins to these granule - tethered proteins . for example , cpe has been proposed to interact with several granule cargo proteins including proenkephalin , proinsulin , pomc ( cool and loh , 1998 ) , brain - derived neurotropic factor ( probdnf ; lou et al . , 2005 ) , and 2005 ) to promote their retention in secretory granules , even though some of these are not enzymatic substrates of cpe . no common mechanism for interaction of these cargo proteins with cpe has yet emerged , although undefined residues within an n - terminal disulfide - constrained hydrophobic loop in pomc ( loh et al . , 2002 ) and acidic residues in probdnf ( lou et al . , 2005 ) have been reported , and both seem to be important for sorting the respective proteins to dcsgs ( cool et al . , 1995 ; lou et al . , 2005 ) . paired basic amino acids have also been reported to direct dcsg sorting in some proteins , including proneurotensin ( feliciangeli et al . , 2001 ) , prorenin ( brechler et al . , 1996 ) , prothyrotropin - releasing hormone ( mulcahy et al . , 2005 ) , and progastrin ( bundgaard et al . , 2004 ) , and to increase the sorting efficiency of proinsulin ( kuliawat et al . , 2000 ) . in the analyses performed to date , it appears that these paired basic amino acids must constitute a cleavage site for one of the granule - resident prohormone convertases ( pc1/3 or pc2 ) to function as a granule sorting domain because changing the cleavage site to one recognized by furin ( another member of the family that cleaves its substrates in the early secretory pathway ) causes the proteins to be secreted through the default constitutive pathway ( brechler et al . , 1996 ) . these results raise the possibility that certain dcsg - targeted proteases can act as sorting chaperones for their substrates , in addition to being processing proteases . muclin has also been suggested to act as a granule cargo receptor in pancreatic cells through its binding of sulfate groups on o - linked glycosylated proteins ( boulatnikov and de lisle , 2004 ) . atrial natriuretic factor ( anf ) has also been shown to be tightly bound to the membranes of atrial myocyte secretory granules through its interaction with pam ( o'donnell et al . , 2003 ) , although it is not a substrate of pam . thus , a variety of interactions with tethers may serve to target proteins to secretory granules . notably , if this mechanism is correct , it would , in some cases , provide a means to ensure that processing enzymes and their substrates end up in the same dcsgs . a third category of granule - targeting mechanisms involves formation of high molecular weight protein complexes or aggregates . indeed , many granule - targeted cargo proteins have the ability to multimerize or aggregate , leading , in most cases , to the formation of electron - dense cores . a direct correlation between the ability to aggregate in vitro and to be sorted to secretory granules in transfected cells has been reported for rat pro - anf ( canaff et al . , 1996 ) and cga ( jain et al . , 2002 ) . because granins are acidic proteins that cluster in the slightly acidic environment present in dcsgs ( for review see dannies , 2001 ) it has been suggested that aggregation may serve to prevent their extrusion from the maturing granule . ( 2005 ) showed that treatment of pc12 cells with bafilomycin a1 , which is a specific inhibitor of vacuolar h - atpase , resulted in a decrease in regulated secretion of cga with a concomitant decrease in visible dcsgs , suggesting that regulated secretion of cga and dense core formation are linked in dcsgs . kim et al . ( 2001 ) also showed that silencing cga expression in pc12 cells results in a loss of visible dcsgs , leading the authors to the striking conclusion that cga is not only a component of the dense core , but that it is also a master regulator of dcsg biogenesis . ( 2004 ) suggests that the role of cga in this process may not be that simple ; they found no correlation between dscg content and cga expression in isolated clonal lines of pc12 cells , suggesting that other proteins could be contributing to dcsg appearance . in fact , expression of several other dcsg cargo proteins , including provasopressin , prooxytocin , pomc , secretogranin ii , and chromogranin b , is sufficient to induce aggregate - containing cytoplasmic vesicles , even in cells with no regulated secretory pathway ( beuret et al . , 2004 ) , although these probably do not display all of the functional characteristics of dcsgs ( meldolesi et al . , 2004 ) . ( 2004 ) recently reported that a polypyrimidine - binding protein ( ptb ) , which is up - regulated under conditions of high insulin demand , stabilizes messenger rnas of many of these same dcsg cargo proteins in insulin - producing cells and leads to increased granule formation . cga has also been reported to induce the expression of pn-1 , which is a serine protease inhibitor that slows the turnover of several dcsg cargo proteins ( kim and loh , 2006 ) that could provide an additional mechanism for increasing dcsg aggregate formation . because cga binds to another granin partner , sgiii ( hosaka et al . , 2004 ) , which , in turn , can associate with cholesterol ( hosaka et al . , 2004 ) and cpe ( hosaka et al . , 2005 ) , aggregation may synergize with protein protein and protein membrane interactions to improve the retention of cargo proteins in the maturing granule and their regulated secretion . in spite of the compelling arguments presented for these various dcsg sorting mechanisms , their translation to the whole animal has been anything but simple . one example of this difficulty is the proposed role of cga as a master regulator of granule formation . although down - regulation of cga expression was reported to result in the loss of detectable dcsgs in cultured pc12 cells ( kim et al . , 2001 ) , ( mahapatra et al . , 2005 ) or no discernable effect ( hendy et al . , 2006 ) on dcsg formation in the cga - rich adrenal chromaffin cells in two independent studies . in spite of the differences in the effects on dcsg morphology , both groups report a similar and dramatic effect on catecholamine secretion in the cga - deficient mice , proving that cga deficiency is not entirely without consequence . how can these apparent differences in the requirement for cga be explained ? one obvious possibility is that in vivo , other dcsg cargo proteins can complement the function of cga in the formation of the dense core , but can not compensate for its absence in catecholamine storage and secretion . in support of this possibility , the group that saw no effect of cga inactivation on dcsg formation reported an up - regulation of cgb and sgii in the adrenal glands of the engineered mice ( hendy et al . , 2006 ) . thus , although cga may affect dcsg formation in some cultured cells , this particular function can obviously be replaced in vivo . nevertheless , although experiments to date have not identified a master regulator of dcsg formation , the concept may not be entirely wrong in specific cell types ; anf inactivation in mice leads to a complete loss of visible dcsgs in the cardiac atrium ( john et al . , 1995 ) , and inactivation of the renin gene leads to a complete disappearance of dcsgs in the juxtaglomerular cells of the kidney ( clark et al . , it 's important to note , however , that regulated secretion can occur in the absence of a dense core as it does in many neurosecretory vesicles . in the case of the anf and renin - deficient mice , it will be intriguing to determine if the remaining cargo proteins are still packaged in such vesicles in the absence of the aggregating partner . a similar conundrum exists with cpe as a sorting receptor for a variety of dcsg cargo proteins . ( 1997 ) originally proposed cpe as the regulated secretory pathway sorting receptor because they observed endocrine disorders in the cpe fat / fat mouse that harbors a mutation in the cpe gene . proinsulin and pomc are among the several proteins that were shown to bind to cpe and that were proposed to enter dcsgs by this association ( cool and loh , 1998 ) . however , recent results demonstrate that both proinsulin and pomc are correctly targeted to dcsgs in cpe fat / fat mice ( irminger et al . , 1997 ; hosaka et al . , although there may be many reasons why it has been hard to derive a consensus for the mechanisms and components of the dcsg sorting machinery , the most intuitive is that we are the victims of our own scientific reductionism , i.e. , that in our search for a simple canonical sorting mechanism we have developed a grossly oversimplified view of the way in which proteins enter dcsgs . nearly 100% of the proinsulin produced in pancreatic enters dcsgs ( rhodes and halban , 1987 ) , whereas only about 25% of the prorenin in the secretory pathway of kidney juxtaglomerular cells is sorted to dcsgs ( pratt et al . , 1987 ) . proinsulin contains numerous potential dcsg sorting domains , such as a binding domain for cpe ( cool and loh , 1998 ) , two paired basic amino acid protease cleavage sites ( steiner et al . , 1996 ) , and the ability to hexamerize and subsequently aggregate ( quinn et al . , 1991 ) , whereas prorenin only contains a single dcsg sorting domain : a paired basic amino acid protease cleavage site ( brechler et al . , 1996 ) . in the case of prorenin , changing even a single one of these basic amino acids completely eliminates dcsg targeting in tissue culture cells ( brechler et al . , 1996 ) . in contrast , neither the mutation of the protease cleavage sites ( halban and irminger , 2003 ) nor the hexamerization domain ( quinn et al . , 1991 ) of proinsulin appears to affect its dcsg sorting . combined with the finding that proinsulin is still efficiently sorted to dcsg in cpe - deficient mice ( irminger et al . , 1997 ) however , another possible explanation is that , with its many dcsg sorting signals , proinsulin might be able to compensate for the loss of any single sorting domain . there is , in fact , some evidence to support the view that dcsg sorting signals can synergize ; duplicating the disulfide - constrained loop dcsg sorting signal normally found at the n terminus of cgb results in a greater sorting efficiency to dcsg than the native protein ( glombik et al . , 1999 ) . furthermore , combination of helical and paired basic amino acid sorting domains on either the same protein or on two proteins capable of dimerizing led to a dramatic increase in dcsg sorting over proteins containing either individual domain ( lacombe et al . , 2005 ) . thus , diverse sorting signals may be able to functionally complement each other even through protein ( 2005 ) also reported that pituitaries of the cpe fat / fat mouse contain elevated levels of both sgiii and cga that might compensate for the loss of cpe in targeting pomc to dcsgs . all of these cases are consistent with the existence of multiple sorting mechanisms , each of which can contribute to the overall efficiency of protein sorting or retention in dcsgs . cell types and the nature and/or the number of the sorting domains contained in the cargo protein would ultimately determine the extent to which each mechanism is active . multimerization and aggregation could add synergy between mechanisms used by other dcsg cargo proteins in the aggregate . with such a model , it 's also easy to imagine how changing conditions within the cell could alter dcsg sorting efficiency of a protein , which is a potential control point that has important implications for hormone secretion but has received little attention to date . work to date has largely supported the three previously outlined postulates : ( a ) proteins exist that could explain the anchoring of dcsg cargo to the granule membrane , ( b ) protein complexes between processing enzymes and their substrates have been proposed that could explain how these end up in the same granules , and ( c ) the diversity of sorting mechanisms helps to understand how there could be distinct dcsgs even within single cells and how there could be such a lack of agreement on the mechanism of dcsg sorting . unfortunately , this lack of simple consensus has limited progress , primarily because of attempts to determine the role of single targeting motifs in given proteins in various cell culture models that may or may not be entirely appropriate . although this approach has certainly not been without worth , a better understanding of the complexity of this important cellular event may help to design experiments that will help to significantly advance this field of research . how does the dcsg protein cargo identify the membrane patches that will make up the mature granule ? how do these complexes communicate with the cytoplasmic accessory proteins that are necessary for the formation of the budding dcsgs ? in fact , although it has been difficult to explain the entry of proteins into dcsgs , we can expect that describing the assembly of proteins on the surface of the dcsgs that are necessary for their cytoplasmic transport , docking at the membrane , and exocytosis will be an equally daunting challenge . a recent report on the components of the functionally related synaptic vesicle identified > 400 associated proteins ( takamori et al . , 2006 ) . what is abundantly clear is that in the characterization of this unique organelle , there is still a lot to sort out . Output:	the intracellular sorting of peptide hormone precursors to the dense core secretory granules ( dcsgs ) is essential for their bioactivation . despite the fundamental importance of this cellular process , the nature of the sorting signals for entry of proteins into dcsgs remains a source of vigorous debate . this review highlights recent discoveries that are consistent with a model in which several protein domains , acting in a cell - specific fashion and at different steps in the sorting process , act in concert to regulate the entry of proteins into dcsgs .
PubmedSumm2050	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: four months before , the patient had developed cardiogenic shock while attempting stent insertion ( endeavor 3.0~18 ) into the obstructed left main coronary artery ( 80% ) , which necessitated urgent institution of extracorporeal membrane oxygenation ( ecmo ) and intra - aortic balloon pump . after coming off extracorporeal life support systems , she underwent stenting ( taxus 2.75~38 ) of the obstructive left anterior descending coronary artery ( lad ) , which resulted in second attack of cardiogenic shock and ecmo support , which was successfully weaned off . she had been discharged home after these events , and had been followed up in the outpatient department . on arrival at er , her consciousness was clear , and chest x - ray showed pulmonary congestion . on echocardiography , severe mitral regurgitation due to restricted valve motion was noted , and left ventricular ejection fraction was 28% . she developed the third attack of cardiogenic shock during emergency coronary angiogram , and was put on ecmo again ( fig . 1 ) . coronary angiogram on ecmo support showed restenosis of the previous lad stent ( 80% ) and complete obstruction of the left circumflex and right coronary arteries . after ecmo circuit was converted to conventional cardiopulmonary bypass using the same femoral artery - vein cannulae , a great saphenous vein graft was anastomosed to the lad and an additional saphenous vein graft was placed from the lad graft to the posterior descending coronary artery in an inverted - y fashion . operative findings of the mitral valve consisted of severe fibrotic change of the anterior leaflet , dilatation of the mitral valve annulus , and central non - coaptation . the patient was able to come off bypass without any event . cardiopulmonary bypass time and aortic cross - clamping time were 168 minutes and 85 minutes , respectively . in provision for the later postoperative use , we decided to leave the femoral arterial and venous cannulae in situ , which were removed on postoperative day 1 . she was extubated on postoperative day 4 , but developed superficial wound infection with dehiscence which was repair on postoperative day 36 . she was readmitted in 2 months after discharge due to general weakness , fever , and septicemia . echocardiography showed 73 mm sized vegetation at the prosthetic mitral valve , and bacteriologic examination identified methicillin - resistant staphylococcus aureus as a causative microorganism . surgical intervention was recommended , but the patient refused to receive further treatment and died of sepsis . he had a smoking history of 10 pack - years , and had been followed up in the out - patient clinic for systemic hypertension , myocardial infarction and moderate mitral regurgitation . four months before , he had undergone coronary artery stenting ( cypher 3.0~33 ) to relieve complete obstruction of lad , which resulted in the deterioration of mitral regurgitation . echocardiography at this admission revealed severe mitral regurgitation due to restricted valve motion , and left ventricular ejection fraction was 30% . upon coronary angiography x ray and moderate left ventricular dysfunction , we elected to put the patient on ecmo temporarily to ameliorte cardiopulmonary function . after 2 days of ecmo support , the patient was brought to the operating room and cardiopulmonary bypass was instituted using the same femoral arterial and venous cannulae . an additional venous cannula was inserted into the superior caval vein after sternotomy due to insufficient venous drainage from the single femoral venous cannula . operative findings of the mitral valve were annular dilatation and restrictive motion of the posterior leaflet . ring annuloplasty using commercialized mitral ring ( carpentier - edwards physio ring 28 mm , edwards ) was performed . the patient came off cardiopulmonary bypass without any event , and , differing from the 1st case , the femoral arterial and venous cannulae were removed after the completion of the procedures . intra - operative trans - esophageal echocardiography showed mild residual mitral regurgitation . cardiopulmonary bypass time and aortic cross - clamping time were 103 minutes and 53 minutes , respectively . the patient was extubated on postoperative day 1 , and echocardiography on postoperative day 19 showed mild to moderate residual mitral regurgitation . he was discharged home on postoperative day 29 , and has been followed up for 14 months in a good clinical condition . four months before , the patient had developed cardiogenic shock while attempting stent insertion ( endeavor 3.0~18 ) into the obstructed left main coronary artery ( 80% ) , which necessitated urgent institution of extracorporeal membrane oxygenation ( ecmo ) and intra - aortic balloon pump . after coming off extracorporeal life support systems , she underwent stenting ( taxus 2.75~38 ) of the obstructive left anterior descending coronary artery ( lad ) , which resulted in second attack of cardiogenic shock and ecmo support , which was successfully weaned off . she had been discharged home after these events , and had been followed up in the outpatient department . on arrival at er , her consciousness was clear , and chest x - ray showed pulmonary congestion . on echocardiography , severe mitral regurgitation due to restricted valve motion was noted , and left ventricular ejection fraction was 28% . she developed the third attack of cardiogenic shock during emergency coronary angiogram , and was put on ecmo again ( fig . 1 ) . coronary angiogram on ecmo support showed restenosis of the previous lad stent ( 80% ) and complete obstruction of the left circumflex and right coronary arteries . after ecmo circuit was converted to conventional cardiopulmonary bypass using the same femoral artery - vein cannulae , a great saphenous vein graft was anastomosed to the lad and an additional saphenous vein graft was placed from the lad graft to the posterior descending coronary artery in an inverted - y fashion . operative findings of the mitral valve consisted of severe fibrotic change of the anterior leaflet , dilatation of the mitral valve annulus , and central non - coaptation . the patient was able to come off bypass without any event . cardiopulmonary bypass time and aortic cross - clamping time were 168 minutes and 85 minutes , respectively . in provision for the later postoperative use , we decided to leave the femoral arterial and venous cannulae in situ , which were removed on postoperative day 1 . she was extubated on postoperative day 4 , but developed superficial wound infection with dehiscence which was repair on postoperative day 36 . she was readmitted in 2 months after discharge due to general weakness , fever , and septicemia . echocardiography showed 73 mm sized vegetation at the prosthetic mitral valve , and bacteriologic examination identified methicillin - resistant staphylococcus aureus as a causative microorganism . surgical intervention was recommended , but the patient refused to receive further treatment and died of sepsis . he had a smoking history of 10 pack - years , and had been followed up in the out - patient clinic for systemic hypertension , myocardial infarction and moderate mitral regurgitation . four months before , he had undergone coronary artery stenting ( cypher 3.0~33 ) to relieve complete obstruction of lad , which resulted in the deterioration of mitral regurgitation . echocardiography at this admission revealed severe mitral regurgitation due to restricted valve motion , and left ventricular ejection fraction was 30% . upon coronary angiography x ray and moderate left ventricular dysfunction , we elected to put the patient on ecmo temporarily to ameliorte cardiopulmonary function . after 2 days of ecmo support , the patient was brought to the operating room and cardiopulmonary bypass was instituted using the same femoral arterial and venous cannulae . an additional venous cannula was inserted into the superior caval vein after sternotomy due to insufficient venous drainage from the single femoral venous cannula . operative findings of the mitral valve were annular dilatation and restrictive motion of the posterior leaflet . ring annuloplasty using commercialized mitral ring ( carpentier - edwards physio ring 28 mm , edwards ) was performed . the patient came off cardiopulmonary bypass without any event , and , differing from the 1st case , the femoral arterial and venous cannulae were removed after the completion of the procedures . intra - operative trans - esophageal echocardiography showed mild residual mitral regurgitation . cardiopulmonary bypass time and aortic cross - clamping time were 103 minutes and 53 minutes , respectively . the patient was extubated on postoperative day 1 , and echocardiography on postoperative day 19 showed mild to moderate residual mitral regurgitation . he was discharged home on postoperative day 29 , and has been followed up for 14 months in a good clinical condition . coronary artery disease is frequently associated with mitral regurgitation . as benefits are substantiated by the meta - analysis of the reports pertaining to the outcome of surgical intervention on ischemic mitral regurgitation , concomitant mitral valve surgery upon coronary artery bypass grafting is recommended by the major academic bodies in north america ( i.e. american college of cardiology , american heart association ) and in europe ( i.e. european society of cardiology ) . elimination of the left ventricular volume overloading by mitral valve surgery is believed to be beneficial for the recovery of left ventricular function after coronary artery revascularization . with respect to the types of mitral valve surgery , either mitral valve replacement or mitral annuloplasty could be performed according to the disposition of valve pathology and patient characteristics . for instance , valve replacement is recommended in case of papillary muscle rupture or leaflet motion restriction caused by papillary muscle displacement . there are a number of reports asserting the superiority of the mitral valve repair over replacement , but selection from the two surgical options should be undertaken based on the exact valve regurgitation mechanism of individual patients . in this report , the first patient underwent valve replacement due to the severe fibrotic change of the anterior leaflet , while ring annuloplasty alone was deemed appropriate to alleviate severe mitral regurgitation in the second case . as for the selection of the life support system , intra - aortic balloon pump ( iabp ) has been widely used as a rescue measure for circulatory collapse . iabp is believed to improve left ventricular function by reducing left ventricular afterload and augmenting coronary perfusion while maintaining pulsatility of the systemic circulation . however , barlett et al . claimed that ecmo is more beneficial for cardiogenic shock , given that patients in this setting may need both circulatory and pulmonary support . in between these two assertions , proposed that iabp is indicated for patients with coronary artery disease while ecmo is preferred in patients with valvular heart diseases . in our first case , ecmo was used because coronary arteries were severely obstructed and iabp alone might have not been effective . in the second case , indications for ecmo or other percutaneous circulatory support systems are diverse , such as cardiac arrest , cardiogenic shock , high - risk cardiovascular intervention , bridge to emergency cardiac operation or cardiac transplantation , and pulmonary thromboembolism [ 6 - 10 ] . conducted a meta - analysis of 85 publications between 1966 and 2005 regarding percutaneous cardiopulmonary bypass , and concluded that this measure is mostly effective for rescuing critically ill patients , providing the median survival of 47% in 1,495 patients enrolled in this analysis . in the literature , however , survival rate after instituting mechanical circulatory support widely varies between 15% and 60% , which may well be attributed to small number of patients in most of the reports , diversity of the underlying diseases , and differences in experience and quality of care from one institution to another . it has been reported that selection of candidates , timing of life support system installation and technical aspects mainly determine the survival . mechanical circulatory support may be mandatory upon coronary angiography or catheter intervention in high - risk patients with cardiogenic shock or uncontrolled arrhythmia . reported that mechanical circulatory support was necessary in 275 ( 15.4% ) among 1,785 patients who underwent cardiovascular intervention for acute myocardial infarction . with respect to the institution of life support system as a bridge to a definitive cardiac operation , there are reports of successful cardiac operation after an urgent installation of mechanical circulatory support for cardiogenic shock which developed during coronary angiography . grambow et al . reported 20% survival in patients with circulator assist device instituted for circulatory collapse in the catheterization laboratory . for the percutaneous circulatory support , we are using capiox emergency bypass system ( ebs ; terumo inc , tokyo , japan ) , which consists of centrifugal pump , membrane oxygenator ( polypropylene hollow fiber ) and heparin - coated circuit . this system is characterized by lower heparin administration , longer durability of the oxygenator , fast and easy auto - priming , and small - sized equipment with excellent adaptability to a small space . thus , we have an institutional strategy to institute ebs system as soon as possible in case of cardiogenic shock after coronary artery angiography or intervention , and this might have lead to rescue the two patients with cardiogenic shock in the catheterization laboratory . Output:	indication for extracorporeal membrane oxygenation ( ecmo ) has been extended as the experience of ecmo in various clinical settings accumulates and the outcome after ecmo installation improves . we report two cases of successful mitral valve surgery for severe ischemic mitral regurgitation in patients on ecmo support for cardiogenic shock which developed upon coronary angiography .
PubmedSumm2051	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the morbidity of breast cancer is gradually increasing , and a growing number of young people are now at risk for this disease . in china , breast cancer continues to be the leading cause of death among women . with the improvement in diagnosis and development of drugs and radiotherapy technology . therefore , early detection of the tumor and accurate analysis of molecular biology characteristics are still of great importance . microrna ( mirna ) is a non - coding protein and single - stranded small rna . it mainly inhibits translation or induces the degradation of some related mrnas through target mrna recognition , leading to changes in objective mrna at the protein expression level . recent studies showed that some mirnas have functions similar to that of oncogenes or anti - oncogenes , which may play an important role in the invasion and development of breast cancer . mir-125a , mir-206 , and mir-145 are similar to anti - oncogenes , and they are related to the occurrence of tumors . thus , mir-206 plays a key role in adjustment during breast cancer proliferation . according to the research of scott and gk , mir-125a can affect the protein levels of her-2 and her-3 , thereby inhibiting the metastatic and invasive abilities of breast cancer cells . moreover , lee showed that mir-145 expression increases after mir-145 precursors transfect breast cancer cell lines , but can dose - dependently inhibit cell growth . mir-21 and mir-10b are similar to oncogenes , and related to the invasion and metastasis of tumors . in vitro and in vivo experiments showed that eliminating mir-21 gene fragments can inhibit the proliferation and invasion of mcf-7 and mda - mb-231 cells . in the 4t1 rat breast tumor metastasis model , mir-10b can effectively inhibit the metastasis of malignant breast cancer cells with high specificity . the possible mechanism for this effect may be the inhibition of the entry of 4ti tumor cells into blood vessels . however , the specific roles of mir-21 and mir-10b in the occurrence and development of breast cancer remain unclear . the expression of mir-21 , mir-10b , mir-125a , mir-206 , and mir-145 in breast cancer tissues and para - carcinoma tissues was examined . the expression of estrogen receptor ( er ) and progesterone receptor ( pr ) in breast cancer was analyzed by immunohistochemistry , whereas that of her-2 and topoisomerase 2a ( top 2a ) was analyzed by fluorescence in situ hybridization ( fish ) . the relationship between the expression levels of mirnas and breast cancer clinical pathology was analyzed . thus , this research aimed to investigate the function of mirnas in the invasion and development of breast cancer . cancer tissues were obtained from these patients , who were pathologically confirmed to have invasive ductal carcinoma from january 1 2013 to december 1 2013 in the tumor hospital of the second affiliated hospital of xjtu . all patients were female , with an age range of 4262 years ( mean age , 51 years ) . these patients underwent modified radical mastectomy or reserved radical mastectomy , and had complete clinical pathology data . the diameter of tumors ranged from 1.6 cm to 4.5 cm ( mean diameter , 2.2 cm ) . one sample was cancer tissues , and the other sample was para - carcinoma normal gland tissues , which were more than 3 cm away from cancer tissues . the samples were immediately flushed with 0.9% rna- and enzyme - free nacl , rapidly immersed in rna enzyme inhibitor , and incubated at 4c overnight . the protective liquid was removed from the samples , which were stored in a refrigerator at 80c . the patients were not treated before the operation , and neither neoadjuvant chemotherapy nor endocrine therapy was conducted . based on histological classification by the who ( 2003 ) standards , m - mlv reverse transcriptase , rna enzyme inhibitor , oligo dt , dntps , kod - plus , and sybr mastermix were obtained from takara ( japan ) , and primers were from shanghai shenggong company ( china ) . breast cancer her-2 , neu ( 17q12 ) , top 2a ( 17q21 ) , and csp 17 multicolor detection kits were purchased from guangzhou biping medical technology company . total rna ( 500 ng ) was used for rt by mirna stem - loop rt primers . the reaction was conducted in a 10-l system at the following reaction conditions : 16c for 30 min , 42c for 15 min , and 85c for 5 s. real - time qpcr was performed with a 20-l reaction system . the mirna reaction detection system included 1 l of rt products , 1 sybr green i mastermix , 0.5 ixmol / l mirna - specific f - primer and 0.5 ixmol / l universal r - primer . the conditions of immediate quantitative pcr were as follows : 95c for 10 min , followed by 40 cycles of 95c for 15 s and 60c for 1 min . the circulation times in which real - time qrt - pcr reached the set - up threshold in every reaction tube ( ct data ) were recorded . according to some related studies , the 2 method was used to express the gene expression of tumor tissues relative to that of the paired normal tissues . moreover , ct=(ct mirna - ct u6 ) tumor ( ct mirna - ct u6 ) was normal . samples were fixed in formalin , embedded in paraffin , and sliced into 4-m - thick sections . the results show positive expression of ep and er at 10% , and the cancer cell nucleus was brown . samples were fixed in formalin , embedded in paraffin , and sliced into 4-m - thick sections . sections were initially observed under the 40 objective with a suitable filter , and counted under the 100 objective . the signals of gene - specific probe ( gsp ) her-2 ( red ) , gsp top 2a ( green ) , and centromeric probe ( csp ) 17 ( cyan ) were counted in 60 nuclei at the clear tumor areas . the sum of signals of gsp her-2 , gsp top 2a , and csp 17 were recorded , and the data of gsp her-2/csp 17 and gsp top 2a / csp 17 were calculated . the signals of her and top 2a were analyzed according to the following criteria : a ) the signal of gsp her-2 and top 2a ( red , green ) combined to bunches ; b ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) 2.2 and 2 ; c ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) < 1.8 and 0.8 ; and d ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) between 1.82.2 and 0.82.0 . if a or if d , new signal data should be counted in 20 other nuclei , or another researcher should recalculate the data . researchers should indicate the consequence if the recalculated data were at the critical value . under situations with weak signal or high background , signal judgment may be disturbed , so the experiment was repeated using another tissue slice . differences in expression were analyzed by spss 13.0 . given the limited number of samples in this study , the data demonstrate an abnormal distribution . all mirna expression data are shown as the median and interquartile range ( iqr ) . the nonparametric wilcoxon test was used to rank matched - pair data , and the mann - whitney u test was used to compare data between the 2 groups . cancer tissues were obtained from these patients , who were pathologically confirmed to have invasive ductal carcinoma from january 1 2013 to december 1 2013 in the tumor hospital of the second affiliated hospital of xjtu . all patients were female , with an age range of 4262 years ( mean age , 51 years ) . these patients underwent modified radical mastectomy or reserved radical mastectomy , and had complete clinical pathology data . the diameter of tumors ranged from 1.6 cm to 4.5 cm ( mean diameter , 2.2 cm ) . one sample was cancer tissues , and the other sample was para - carcinoma normal gland tissues , which were more than 3 cm away from cancer tissues . the samples were immediately flushed with 0.9% rna- and enzyme - free nacl , rapidly immersed in rna enzyme inhibitor , and incubated at 4c overnight . the protective liquid was removed from the samples , which were stored in a refrigerator at 80c . the patients were not treated before the operation , and neither neoadjuvant chemotherapy nor endocrine therapy was conducted . based on histological classification by the who ( 2003 ) standards , m - mlv reverse transcriptase , rna enzyme inhibitor , oligo dt , dntps , kod - plus , and sybr mastermix were obtained from takara ( japan ) , and primers were from shanghai shenggong company ( china ) . breast cancer her-2 , neu ( 17q12 ) , top 2a ( 17q21 ) , and csp 17 multicolor detection kits were purchased from guangzhou biping medical technology company . total rna ( 500 ng ) was used for rt by mirna stem - loop rt primers . the reaction was conducted in a 10-l system at the following reaction conditions : 16c for 30 min , 42c for 15 min , and 85c for 5 s. real - time qpcr was performed with a 20-l reaction system . the mirna reaction detection system included 1 l of rt products , 1 sybr green i mastermix , 0.5 ixmol / l mirna - specific f - primer and 0.5 ixmol / l universal r - primer . the conditions of immediate quantitative pcr were as follows : 95c for 10 min , followed by 40 cycles of 95c for 15 s and 60c for 1 min . the circulation times in which real - time qrt - pcr reached the set - up threshold in every reaction tube ( ct data ) were recorded . according to some related studies , the 2 method was used to express the gene expression of tumor tissues relative to that of the paired normal tissues . moreover , ct=(ct mirna - ct u6 ) tumor ( ct mirna - ct u6 ) was normal . samples were fixed in formalin , embedded in paraffin , and sliced into 4-m - thick sections . the results show positive expression of ep and er at 10% , and the cancer cell nucleus was brown . samples were fixed in formalin , embedded in paraffin , and sliced into 4-m - thick sections . sections were initially observed under the 40 objective with a suitable filter , and counted under the 100 objective . the signals of gene - specific probe ( gsp ) her-2 ( red ) , gsp top 2a ( green ) , and centromeric probe ( csp ) 17 ( cyan ) were counted in 60 nuclei at the clear tumor areas . the sum of signals of gsp her-2 , gsp top 2a , and csp 17 were recorded , and the data of gsp her-2/csp 17 and gsp top 2a / csp 17 were calculated . the signals of her and top 2a were analyzed according to the following criteria : a ) the signal of gsp her-2 and top 2a ( red , green ) combined to bunches ; b ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) 2.2 and 2 ; c ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) < 1.8 and 0.8 ; and d ) gsp her-2 and top 2a / csp 17 ( red , green / cyan ) between 1.82.2 and 0.82.0 . if a or d , new signal data should be counted in 20 other nuclei , or another researcher should recalculate the data . researchers should indicate the consequence if the recalculated data were at the critical value . under situations with weak signal or high background , signal judgment may be disturbed , so the experiment was repeated using another tissue slice . differences in expression were analyzed by spss 13.0 . given the limited number of samples in this study , the data demonstrate an abnormal distribution . all mirna expression data are shown as the median and interquartile range ( iqr ) . the nonparametric wilcoxon test was used to rank matched - pair data , and the mann - whitney u test was used to compare data between the 2 groups . among the 35 samples , the relative quantity ( 2 ) of mir-125a was below 1 ( 80% ) in 28 cases and above 1 ( 20% ) in 7 cases . the relative quantity ( 2 ) of mir-206 was below 1 ( 86% ) in 30 cases of breast cancer and above 1 ( 14% ) in 5 cases . the relative quantity ( 2 ) of mir-145 was below 1 ( 60% ) in 21 cases of breast cancer and above 1 ( 40% ) in 14 cases . the data ranged from 0.13 to 7.73 and the iqr was 0.511.72 . the median was 0.78 , which is 0.78 times that of normal tissue . the relative quantity ( 2 ) of mir-21 was above 1 ( 94% ) in 33 cases of breast cancer and below 1 ( 6% ) in 2 cases . the relative quantity ( 2 ) of mir-10b was above 1 ( 63% ) in 22 cases of breast cancer and below 1 ( 37% ) in 13 cases . the difference was statistically significant ( p<0.05 ) . among the 35 patients , 16 cases ( 45.7% ) exhibited her-2 gene amplification , which was demonstrated as a red cluster or multiple red points , whereas 18 cases ( 51.4% ) exhibited top 2a gene amplification , which was demonstrated as a green cluster or multiple green points ( table 1 ) . negative expression of her-2 and top 2a was observed in the 17 chromosome ( figure 1 ) . the expression levels of mir-125a , mir-206 , and mir-145 in breast cancer tissues were all lower than those in para - carcinoma tissues . low expression of mir-125a in the metastatic lymph node group and high negative expression of her-2 and top 2a in patients were unrelated to the tumor size , patient age , menstruation , negative er , and pr expression ( p<0.10 ) . mir-206 was highly expressed in patients with small tumors , negative er , negative pr , and negative her-2 ( p<0.10 ) . thus , mir-206 was irrelevant to patient age , menstruation , metastatic lymph node , and top 2a expression ( table 2 ) . mir-145 was negatively correlated with the tumor size and lymph node metastasis ( p<0.05 ) , and had no correlation with patient age , menstruation , and pr expression . however , mir-145 was correlated with negative er , negative her-2 , and negative top 2a ( p<0.05 ) ( table 3 ) . the expression of mir-21 and mir-10b in breast cancer tissues was obviously higher than that in para - carcinoma tissues . mir-21 exhibited high expression in older , postmenopausal patients with various symptoms , such as lymph node metastasis and positive her-2 ( p<0.10 ) , but it was not related to the tumor size and expression levels of top 2a , er , and pr . the expression of mir-10b was related to the tumor size , lymph node metastasis , and positive top-2a ( p<0.05 ) , but it was not associated with age , menstruation , er , pr , and her-2 ( table 3 ) . among the 35 samples , the relative quantity ( 2 ) of mir-125a was below 1 ( 80% ) in 28 cases and above 1 ( 20% ) in 7 cases . the relative quantity ( 2 ) of mir-206 was below 1 ( 86% ) in 30 cases of breast cancer and above 1 ( 14% ) in 5 cases . the relative quantity ( 2 ) of mir-145 was below 1 ( 60% ) in 21 cases of breast cancer and above 1 ( 40% ) in 14 cases . the data ranged from 0.13 to 7.73 and the iqr was 0.511.72 . the median was 0.78 , which is 0.78 times that of normal tissue . the relative quantity ( 2 ) of mir-21 was above 1 ( 94% ) in 33 cases of breast cancer and below 1 ( 6% ) in 2 cases . the relative quantity ( 2 ) of mir-10b was above 1 ( 63% ) in 22 cases of breast cancer and below 1 ( 37% ) in 13 cases . among the 35 patients , 16 cases ( 45.7% ) exhibited her-2 gene amplification , which was demonstrated as a red cluster or multiple red points , whereas 18 cases ( 51.4% ) exhibited top 2a gene amplification , which was demonstrated as a green cluster or multiple green points ( table 1 ) . negative expression of her-2 and top 2a was observed in the 17 chromosome ( figure 1 ) . the expression levels of mir-125a , mir-206 , and mir-145 in breast cancer tissues were all lower than those in para - carcinoma tissues . low expression of mir-125a in the metastatic lymph node group and high negative expression of her-2 and top 2a in patients were unrelated to the tumor size , patient age , menstruation , negative er , and pr expression ( p<0.10 ) . mir-206 was highly expressed in patients with small tumors , negative er , negative pr , and negative her-2 ( p<0.10 ) . thus , mir-206 was irrelevant to patient age , menstruation , metastatic lymph node , and top 2a expression ( table 2 ) . mir-145 was negatively correlated with the tumor size and lymph node metastasis ( p<0.05 ) , and had no correlation with patient age , menstruation , and pr expression . however , mir-145 was correlated with negative er , negative her-2 , and negative top 2a ( p<0.05 ) ( table 3 ) . the expression of mir-21 and mir-10b in breast cancer tissues was obviously higher than that in para - carcinoma tissues . mir-21 exhibited high expression in older , postmenopausal patients with various symptoms , such as lymph node metastasis and positive her-2 ( p<0.10 ) , but it was not related to the tumor size and expression levels of top 2a , er , and pr . the expression of mir-10b was related to the tumor size , lymph node metastasis , and positive top-2a ( p<0.05 ) , but it was not associated with age , menstruation , er , pr , and her-2 ( table 3 ) . recent studies have already proved that the expression of mirnas changes is closely related to various types of human cancer . however , the functions of mirnas are complicated because they can effectively regulate and control hundreds of genes , including oncogenes and anti - oncogenes . studies indicated that mir-125a ( has - mir-125a ) is located at chromosome 19q13.41 , including 2 mirnas that are in the same precursor ( mir-125a.5p and mir-125a.3p ) , and has the same origin as lin-4 in nematodes . mir-125a can be detected as a phylogenetic mutation , which is associated with the occurrence of breast cancer . mir-125a can also inhibit tumors by directly lowering the mrna and protein levels of some genes , such as erbb2 and rna binding protein hur . mir-206 is located at human chromosome 6p12.2 , and is considered to be a type of mirna related to skeletal muscles and the myocardium . it can boost muscle differentiation by lowering the p180 subunit of dna polymerase and muscle transcription factors ( e.g. , is13 and myor ) . mir-206 can also inhibit the proliferation of breast cancer cells , mainly by lowering the mrna and protein expression levels of er and er co - regulatory proteins via the egf / mapk pathway [ lowering steroid acceptor nuclear co - activators ( e.g. , src-1 and src-3 ) and transcription factor gata-3 ] . mir-206 can also act on the protein cx43 to inhibit the invasion and metastasis of cancer cells . mir-206 can result in mutations at the rs6920648 locus , and is associated with the existence of breast cancer . the expression of mir-145 decreases in many types of tumor tissues , including breast cancer , colon cancer , and lung cancer . this phenomenon might be related to several factors , such as the location of mir-145 , ability of mir-145 to easily cause deficiency in tumor cells , and genetic changes in the receptor s surface . similarly , mir-145 can inhibit hyperplasia in tumor cells by affecting target genes , such as c - myc , irs-1 , oct4 , sox2 , and klf4 . mir-21 is an mirna located in the fra17b fragile area of chromosome 17q23.2 , and has an independent transcription unit . moreover , mir-21 controls the expression of many anti - oncogenes , including tpm1 , programmed cell death 4 , maspin , and bcl-2 , to prompt the metastasis and hyperplasia of breast cancer cells . mir-10b is located between the hoxd4 and hoxd8 genes of 2p31.1 , and its structure is 5-uacccuguagaaccgaauuugug-3. mir-10b is related to many types of tumors . adjusting the twist - mir-10b - hoxd 10-rhoc pathway is the main mechanism by which mir-10b inhibits the metastasis of breast cancer cells . mir-10b also facilitates the metastasis of breast cancer cells by regulating the expression of the target genes upar and syndecan-1 . early diagnosis is essential to prevent and cure tumors . in this study , the relative transcript levels of mir-21 and mir-10b in patients tumor tissues were higher than those in para - carcinoma normal tissues . by contrast , the relative transcript levels of mir-145 , mir-125a , and mir-206 in patients tumor tissues were apparently lower than those in para - carcinoma normal tissues . according to the literature [ 1214 ] , the relative transcript levels of mir-145 , mir-125a , and mir-206 in serum are also visibly abnormal compared with those in serum from healthy females . therefore , these mirnas can be used as markers of breast cancer for early diagnosis . our results show that mir-145 and breast cancer tumor size , as well as mir-145 and lymph node metastasis , was negatively correlated in 35 cases of breast cancer . although mir-125a and mir-206 were expressed at low levels in the large mass group and lymph node metastasis group , statistical analysis showed no significant correlation between the expression of mir-125a and mir-206 and breast tumor size , as well as between the expression of mir-125a and mir-206 and lymph node metastasis . we are currently further expanding the number of samples for analysis to determine the intrinsic correlations . preliminary results show that mir-145 , mir-125a , and mir-206 were similar to tumor suppressor genes , and may be involved in the suppression of breast cancer invasion and metastasis ; these findings are consistent with those reported in the literature . as indicated in the literature , mir-10b expression may indicate the invasion and development of breast cancer . combined with the findings of lee , the results show that mir-21 may be correlated with old patients with lymph node metastasis . hormone receptor - positive breast cancer , her-2-positive breast cancer , and triple - negative breast cancer ( tnbc ) ] based on er , pr , and her-2 expression . her-2-positive breast cancer is highly malignant , and can reappear and transfer easily . herceptin - targeted treatment has made significant progress for this type of cancer . lee showed that mir-21 is related to the metastasis of breast cancer lymph nodes and positive her-2 expression . moreover , mir-21 is an independent prognostic factor of breast cancer . combined with cong s research , the results showed that mir-21 is over - expressed in breast cancer cells resistant to trastuzumab , which causes the loss of pten expression . a previous study showed that mir-125a is negatively associated with her-2 , indicating that the her-2 gene may be under the regulation and control of mir-125a ; therefore , mir-125a can distinctly inhibit the expression of oncogenes her-2 and her-3 . mir-125a expression is up - regulated when the hdac inhibitor entinostat is used to treat her-2-positive breast cancer cells , and has been shown to be an important mechanism of the functions of entinostat . thus , mir-21 and mir-125a are significant to developing new anti - her-2 drugs and overcoming herceptin resistance in drugs . tnbc is characterized by high malignancy , high possibility of reappearance and metastasis , and poor treatment methods . in this study , mir-145 and mir-206 were found to be related to negative er and negative her-2 ( p<0.05 ) . mir-145 was highly expressed in negative pr ( p>0.05 ) , whereas mir-206 was negatively correlated with negative pr ( p<0.10 ) . showed that the metastasis of cancer cells could be inhibited by leading mir-145 and mir-206 into metastatic breast cancer cells . mir-145 and mir-206 are expected to be the new target spots in tnbc treatment . besides inhibiting tnbc development , mir-206 can also inhibit positive er lines , and is considered as the switch of breast cancer when it is transferred to the base type from lumen - a . top 2a codes dna topoisomerase ii , and is one of the components of the nuclear matrix . top 2a can adjust the copy , transcription , recombination , and repair of dna , which is not only a hyperplasia index of tumor cells , but also a treatment target of anthracycline chemotherapy drugs . in breast cancer tissues , tumors with high expression of top 2a this situation can be improved after using anthracycline alone or anthracycline combination chemotherapy . in this research , mir-21 was found to be positively related to top 2a , whereas mir-125a and mir-145 were negatively associated with top 2a . these findings show that mir-145 and mir-125a might inhibit the top 2a gene , and were connected with anthracycline drug resistance . therefore , mir-21 , mir-125a , and mir-145 may contribute to the chemotherapy effect of epirubicin , and are important in guiding the development of chemotherapy for breast cancer . therefore , mir-125a , mir-206 , mir-145 , mir-21 , and mir-10b are highly expected to be novel tumor markers for the diagnosis and prognosis of breast cancer . mir-21 and mir-125a could contribute to the treatment of her-2-positive breast cancer cells , whereas mir-145 and mir-206 could accelerate the development of treatment methods for tnbc . however , the detailed mechanism and whether these mirnas can be used as treatment targets in clinical applications should be verified in future investigations . Output:	backgroundthis study aimed to investigate the expression and significance of 5 types of mirnas in breast cancer to provide a theoretical and practical foundation for using these mirnas in the diagnosis and treatment of breast cancer , thereby improving medical services.material/methodsstem-loop real - time rt - pcr was used to detect the expression levels of mir-145 , mir-21 , mir-10b , mir-125a , and mir-206 in 35 cases of breast cancer and adjacent normal breast tissues , and to analyze the relationship of mirnas expression with clinicopathological features of breast cancer . the expression levels of estrogen receptor ( er ) and progesterone receptor ( pr ) were examined by immunohistochemistry . fluorescence in situ hybridization was used for the detection of her-2 and top 2a.resultsthe expression levels of mir-145 , mir-125a , and mir-206 in breast cancer were lower than those in adjacent normal tissues . mir-145 was negatively correlated with tumor size , lymph node metastasis , er , her-2 , and top 2a ( p<0.05 ) , regardless of age , menstruation , and pr . mir-125a was correlated with negative node status , negative her-2 status ( p<0.05 ) , whereas tumor size , age , menstruation , er , and pr were independent factors . mir-206 expression was correlated with negative er status , negative pr status , and negative her-2 status ( p<0.05 ) , regardless of age , menstruation , lymph node metastasis , and top 2a . mir-21 and mir-10b expression in breast cancer tissues was significantly higher than that in adjacent tissues ( p<0.05 ) . mir-21 in post - menstrual patients with lymph node metastasis was highly expressed ( p<0.05 ) , and had no correlations with tumor size , er , pr , and top 2a expression . mir-10b expression was positively correlated with breast cancer tumor size , lymph node metastasis , and top 2a status ( p<0.05 ) , but had no correlations with age , menstruation , er , pr , and her-2.conclusionsmir-145 , mir-21 , mir-10b , mir-125a , and mir-206 may play important roles in breast cancer development and invasion .
PubmedSumm2052	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: trypanosoma cruzi is an intracellular protozoan parasite causing chagas disease , which affects millions of people in latin america . during the acute inflammatory phase of the t. cruzi infection , high - level expression of inducible nitric oxide synthase ( inos ) , proinflammatory cytokines , and arachidonic acid- ( aa- ) derived lipids such as leukotrienes , lipoxins ( lxs ) , h ( p ) etes , prostaglandins , and thromboxane is prevalent [ 3 , 4 ] . in the early t. cruzi infection , nitric oxide ( no ) and arachidonic acid metabolites could be attributed to resistance , but later on to tissue damage . prostaglandins ( pgs ) are oxygenated lipid mediators formed from the 6 essential fatty acid , arachidonic acid ( aa ) . the committed step in pg biosynthesis is the conversion of aa to pg h2 ( pgh2 ) , catalyzed by either pg endoperoxide h synthase-1 or -2 , commonly known as cyclooxygenase-1 ( cox-1 ) and cyclooxygenase-2 ( cox-2 ) , respectively [ 5 , 6 ] . both cox-1 and cox-2 are nonselectively inhibited by nonsteroidal anti - inflammatory drugs ( nsaids ) such as aspirin and ibuprofen , whereas cox-2 activity is selectively blocked by cox-2 inhibitors called coxibs ( e.g. , celecoxib ) [ 7 , 8 ] . the relevance of these enzymes and the bioactive lipids that they produce are not well understood in parasitic disease , although the role of eicosanoids in the pathogenesis of chagas disease is becoming more defined . pharmacological antagonists of cox-1 ( aspirin , asa ) , cox-2 ( celecoxib ) , or both ( indomethacin ) have been found to increase mortality and parasitemia ( parasite load in peripheral blood and cardiac tissue ) regardless of which mouse or t. cruzi strains were used [ 913 ] . conversely , others have found that inhibition of pg synthesis / release reduces parasitemia and extends survival of mice infected with t. cruzi [ 1417 ] . this was often associated with a decrease in the levels of circulating inflammatory cytokines ( such as tnf- , ifn- , and il-10 ) . more recently , treatment with asa during chronic infection was found to be beneficial with no increase in mortality and substantial improvement in cardiac function . additionally , the protective effect of asa could be mediated by the synthesis of 15-epi - lipoxin a4 ( 15-epi - lxa4 ) . given the increasing interest in the role of eicosanoids in t. cruzi infection , we decided to investigate the effect of prostaglandin synthesis inhibition with asa on inflammatory response and macrophage invasion by t. cruzi . six- to eight - week - old balb / c female and male mice were supplied by the multi - institutional center for biological investigation , state university of campinas , brazil . mice were maintained under standard conditions in the animal house of the department of pathological sciences , center for biological sciences , state university of londrina . commercial rodent diet ( nuvilab - cr1 , quimtia - nuvital , colombo , brazil ) and sterilized water were available ad libitum . all animal procedures were performed in accordance with the guidelines of the brazilian code for the use of laboratory animals . the protocols were approved by the internal scientific commission and the ethics in animal experimentation committee of londrina state university ( approval number : ceea 5492.2012.22 ) . t. cruzi y was maintained by weekly intraperitoneal inoculation of swiss mice with 2 10 trypomastigotes . to conduct our experiments , blood from previously infected mice was obtained by cardiac puncture without anticoagulant . the blood was centrifuged at 1,500 g for 1 min and allowed to stand at 37c for 60 min . the supernatant serum containing most of the t. cruzi was centrifuged at 1,200 g for 15 min . the sediment was resuspended in 1 ml of rpmi 1640 medium ( gibco , gran island , ny ) containing 10% inactivated fetal bovine serum ( fbs ) , 100 units of penicillin , and 100 g streptomycin ( gibco , gran island , ny ) . trypomastigotes were derived from the supernatant of t. cruzi - infected llc - mk2 culture cells ( atcc ccl-7 ; american type culture collection , rockville , md ) grown in rpmi 1640 medium containing 10% inactivated fetal bovine serum ( fbs ) , 40 g ml gentamicin , 100 units of penicillin , and 100 g streptomycin ( gibco , gran island , y ) . free parasites were removed after 24 h and cultures were maintained in 10% fbs - rpmi 1640 . mice were inoculated intraperitoneally with 2 ml of 5% thioglycollate and , 4 days later , the elicited cells from the peritoneal exudates were harvested in cold pbs . mouse peritoneum was washed with 5 ml ice - cold , serum - free rpmi . peritoneal cells from 36 mice were pooled and left to adhere in complete medium ( rpmi , 2 mm glutamine , 1 mm sodium pyruvate , 40 g ml gentamicin , and 10 mm hepes ) for 24 h in 24-well plates at 2 10 cells / well . the macrophages were plated onto 13 mm round glass coverslips and washed in warm phosphate - buffered saline ( pbs ) before the interaction assays . one set of plates was used to quantify il-1 and the other set for no detection . before the experiments , peritoneal macrophages previously washed were incubated for 30 min at 37c in a 5% co2 atmosphere in the presence of different concentrations of asa ( 2.5 mm , 1.25 mm , and 0.625 mm ) to test its effect on internalization of the parasite into the host cell . after incubation , the medium containing asa was removed , and macrophages were allowed to interact with trypomastigote forms added in a ratio of 5 parasites per cell . the interaction was allowed to proceed for 2 h , at 37c in a 5% co2 atmosphere . the cells were then washed three times , fixed with bouin 's fixative , stained with giemsa ( merck ) stain , and observed with a light microscope at 1000x magnification . other treatments included incubation with aminoguanidine ( 1 mm ) or l - name ( 1.0 mm ) for 60 min at 37c with or without asa . the internalization index was calculated by multiplying the percentage of infected cells by the mean number of parasites per infected cell . experiments were performed in triplicate , and six independent experiments were completed . all experiments included untreated , infected peritoneal macrophages as controls . the quantification was carried out using light microscopy where a total of 500 cells were randomly counted . the viability of the cells obtained from the cultures before and after incubation experiments was determined using mtt ( sigma - aldrich ) assay , showing the mitochondrial activity of living cells . the culture medium was aspirated , and mtt ( 0.5 mg ml ) was added to the cells prior to incubation at 37c for 4 h. the supernatant was aspirated and dimethyl sulfoxide ( sigma - aldrich ) was added to the wells . insoluble crystals were dissolved by mixing and the plates were read using a biorad multiplate reader ( hercules , ca ) , at a test wavelength of 570 nm and a reference wavelength of 630 nm . production of nitric oxide ( no ) was determined by measuring the level of accumulated nitrite , a metabolite of no in the culture supernatant using griess reagent ( sigma - aldrich ) . after 24 h of treatment with asa ( 0.625 mm ) , the culture supernatants were collected and mixed with an equal volume of griess reagent in 96-well culture plates and incubated at room temperature for 10 min . the absorbance was measured at 540 nm and nitrite concentrations were calculated by reference to a standard curve generated by known concentrations of sodium nitrite . immunocytochemistry for inos was performed on coverslip - adherent cells using the labeled streptavidin biotin method with a lsab kit ( dako japan , kyoto , japan ) without microwave accentuation . the coverslips were incubated with 10% triton x-100 solution for 1 h , washed 3 times in pbs , and treated for 40 min at room temperature with 10% bsa . the coverslips were then incubated overnight at 4c with the primary antibody ( anti - inos rabbit monoclonal antibody diluted 1 : 200 , bd biosciences , catalog number 610599 ) , followed by secondary antibody treatment for 2 h at room temperature . horseradish peroxidase activity was visualized by treatment with h2o2 and 3,3-diaminobenzidine ( dab ) for 5 min . at the last step , the sections were weakly counterstained with harry 's hematoxylin ( merck ) . for each case , negative controls were prepared by omitting the primary antibody . intensity and localization of immune reaction against primary antibody used were examined on all coverslips using a photomicroscope ( olympus bx41 , olympus optical co. , ltd . , after conversion of the images into grey scale ( adobe photoshop ) inos - positive pixels and total pixels became thresholds and were processed by image j program . positive immunostained area was calculated as the proportion of positive pixels to total pixels ( % ) . ( 0.625 mm ) or untreated , either infected or not infected with t. cruzi , were incubated for 24 h. levels of il-1 in 100 l medium were measured by commercial elisa kits ( ready - set - go ! the statistical analysis was conducted using one way anova with bonferroni 's multiple comparison test . statistical analysis was performed using the graphpad prism 5.0 computer software application ( graphpad software , san diego , ca , usa ) . to determine whether cox - derived mediators are involved in t. cruzi entry into host cells , cells were treated with increasing amounts of asa for 30 min and after treatment , the medium containing asa was removed before macrophage invasion assay in order to guarantee that asa affected only the host cell and not the parasites . after 2 h of incubation with parasites , which provides sufficient time for them to enter into macrophages , the free parasites were removed . aspirin irreversibly inhibits cox-1 by acetylation of a single serine residue on the enzyme and this inactivation persists , that is , 24 hours . in some cases , the medium with increasing asa concentrations was added every 24 hours until the end of the t. cruzi infection period ( 7 days , figure 4(d ) ) . figure 1(a ) shows that asa markedly inhibited the internalization of trypomastigote by macrophages at all concentrations tested ( p < 0.0001 ) . thus , pge2 synthesis inhibition using asa improves macrophage response against t. cruzi infection . the cytotoxicity of asa in macrophages was evaluated by mtt assay ( figure 1(a ) , insert ) . asa did not induce cell death , as the concentrations of asa used in all experiments reported were too low to cause cytotoxicity . light microscopy observations confirm t. cruzi inhibition invasion of asa treated cells ( figure 1(c ) ) . we examined the combined effect of celecoxib ( cox-2 selective inhibitor ) and asa on the entry of t. cruzi into phagocytic cells . peritoneal macrophages were incubated separately , with or without drugs ( asa and celecoxib ) either alone or in combination . figure 1(b ) shows that the drugs in combination significantly ( p < 0.01 ) restored the invasive capacity of trypomastigotes . none of the drugs ( asa , celecoxib , or their combination ) showed cytotoxicity against uninfected peritoneal macrophages ( data not shown ) . these results indicate that the enzyme activities of cox-1 and cox-2 favor invasion , but simultaneous inhibition of both activities also favors parasite invasion , probably by allowing a new panorama of eicosanoids production . four days postinfection , macrophages began releasing trypomastigotes into the supernatant ( figure 4(d ) ) . trypomastigotes release to culture supernatants from t. cruzi - infected macrophages was diminished by asa ( 0.625 mm ) . this result is in agreement with previous reports , showing that aspirin and other cox inhibitors decrease t. cruzi infection in vitro [ 1416 , 23 ] . lipoxins ( lxs ) are endogenous lipid mediators with potent anti - inflammatory and proresolving actions [ 24 , 25 ] . native lxs and their stable analogues exert their biological effects by binding to a g - protein - coupled receptor , denoted as alx [ 26 , 27 ] . the effect of asa has been associated , in part , to a switch to the aa pathway linked to the acetylation of the cox-2 isoenzyme . this reaction enables cox-2 to synthesize lxs as 15-epi - lxs ( 15-epi - lxa4 ) . to evaluate the effect of asa - triggered lxs on the entry of t. cruzi into phagocytic cells , peritoneal macrophages were incubated separately either with or without drugs ( asa or boc-2 , a specific antagonist of the 15-epi - lxs ) either alone or in combination . the inhibitory effect of asa on the entry of t. cruzi into macrophages was prevented by boc-2 demonstrating that the effect of asa on the entry of t. cruzi into macrophages could be mediated by the synthesis of 15-epi - lxa4 . furthermore , boc-2 did not have any effect on the entry of t. cruzi into macrophages when used alone ( figure 2(a ) ) . figure 2(b ) ( insert ) shows that treatment of macrophages with high concentrations of pge2 causes inhibition of t. cruzi entry into macrophages . when pge2 ( 3.52 ng ml ) was added alone or in combination with asa , the effect of asa was inhibited , indicating that pge2 is involved in the internalization of t. cruzi trypomastigotes into peritoneal macrophages . the internalization of t. cruzi into macrophages stimulated the release of il-1 while asa increased il-1 production by infected macrophages ( figure 3(a ) ) . the effect of asa on no production was evaluated by detection of nitrite in t. cruzi - infected macrophage supernatants using griess reaction ( figure 3(b ) ) . no production in macrophages was stimulated by t. cruzi and was also increased by prior treatment of macrophages with asa . in addition , we observed that asa treatment stimulated inos expression in t. cruzi - infected macrophages ( figure 4(a ) ) . to confirm that the effect of asa on macrophage activity depends on no levels , we assessed the entry of trypomastigotes into macrophages incubated with aminoguanidine ( ag , inos inhibitor ) or with l - name ( c - nos inhibitor ) . we found that both inhibitors reversed the effects of asa ( figures 4(b ) and 4(c ) ) . previous studies have shown that the release of eicosanoids during infection with t. cruzi regulates host responses and controls disease progression [ 3 , 10 , 11 , 13 , 14 , 2831 ] . pgs , together with no and tnf- , participate in a complex circuit that controls lymphoproliferative and cytokine responses in t. cruzi infection . however , the involvement of cox - mediated pg production in the entry of t. cruzi into macrophages is largely unexplored . the data shown herein demonstrates that the treatment of macrophages with asa significantly inhibits internalization of t. cruzi trypomastigotes and strongly supports the idea that the cox pathway plays a fundamental role in the process of parasite invasion . in fact , pge2 production increases significantly in t. cruzi - infected macrophages as compared to uninfected macrophages and synergistically enhances the activity of nifurtimox and benznidazole on infected raw 264.7 cells . this is demonstrated by the observation that celecoxib reversed the aspirin - induced inhibition of entry of t. cruzi into macrophages . the problem of nsaids coadministration is actively discussed in literature in the context of uncertainty of the resulting therapeutic and side effects arising from such combinations . the mechanism of such a suppression of aspirin inhibitory effect on cox-1 by other nsaids has been difficult to satisfactorily explain . this could be due to celecoxib binding strongly to a monomer of cox-1 without affecting aa oxygenation . in fact , a related study has shown that celecoxib prevented asa inhibition in a dog model of thrombosis . however , the application of our approach to investigate the combined effect of aspirin and celecoxib on cox-1 during coadministration confirmed the ability of celecoxib to suppress the aspirin - mediated inhibition of cox-1 in vitro conditions . in addition , the effects of asa on t. cruzi infection have been associated in part , to a switch to the aa pathway linked to the acetylation of the cox-2 isoenzyme . such modification promotes the synthesis of the 15-r - hete intermediate , which can be transformed by 5-lipooxygenase to 15-epi - lxa4 , a lipid involved in the resolution of inflammation . accordingly , we assessed the effect of boc-2 ( a specific antagonist of the 15-epi - lxs ) on low doses of asa , indicating that 15-epi - lxa4 is probably involved in the inhibitory effect exerted by asa on the internalization of t. cruzi by macrophages . interestingly , when macrophages were treated with pge2 concentration as high as 3.52 ng ml , we observed a reduction in the entry of parasites , but when we used low concentration of pge2 ( 0.35 ng ml ) , the entry of t. cruzi was similar to that observed in untreated macrophages . in addition , we found reversal of asa effect , even when pge2 concentration as low as 3.52 ng ml was used , indicating that pge2 has an important role in asa effect . inhibition of cox activity may increase no levels , thus restoring the antiparasitic activity of macrophages . our results are in agreement with this hypothesis . to confirm that the effect of asa on macrophage activity depends upon restoring no levels , we assessed the invasive capacity of t. cruzi when cells were incubated with aminoguanidine ( ag ) , an inos inhibitor . in addition , we showed that inos expression in macrophages was increased with asa treatment , suggesting that inos - dependent no production is responsible for asa effects . we did find reversal of asa effect with l - name ( 1.0 mm ) , indicating the role of cnos in asa activity . this can be explained as high concentrations of l - name may interfere in the selectivity for cnos and can inhibit other isoforms of nos , such as nnos and inos . additionally , polyamines seem to be crucial for trypomastigote internalization process in , at least , some cellular types and infection progression . in t. cruzi - infected macrophages , cox is related to the increase of ornithine decarboxylase ( odc ) activity , which might increase the polyamine content in macrophages . since t. cruzi uses these polyamines to synthesize trypanothione ( an enzyme that participates in the hydroperoxide detoxification of t. cruzi ) , the inhibition of cox by asa probably resulted in a reduction in polyamine levels caused by inhibition of odc , indirectly contributing to decrease trypanothione synthesis in t. cruzi , as suggested by lpez - muoz and collaborators . finally , in t. cruzi - infected macrophages , cox inhibition by asa was related to the increase of il-1 , which also might explain the increase of antiparasitic activity of macrophages treated with asa . in fact , il-1 is critical for the restriction of leishmania amazonensis infection and it recently was demonstrated that macrophages treated with il-1 released fewer trypomastigotes than untreated macrophages and il-1 triggered no release by t. cruzi - infected macrophages in a dose dependent manner . in conclusion , this is the first report , to our knowledge , showing the in vitro effect of aspirin on t. cruzi entry into peritoneal macrophages and the influence of cox pathway on innate inflammatory response to t. cruzi infection , adding a new perspective to immune interventions against chagas disease . Output:	the intracellular protozoan parasite trypanosoma cruzi causes chagas disease , a serious disorder that affects millions of people in latin america . cell invasion by t. cruzi and its intracellular replication are essential to the parasite 's life cycle and for the development of chagas disease . here , we present evidence suggesting the involvement of the host 's cyclooxygenase ( cox ) enzyme during t. cruzi invasion . pharmacological antagonist for cox-1 , aspirin ( asa ) , caused marked inhibition of t. cruzi infection when peritoneal macrophages were pretreated with asa for 30 min at 37c before inoculation . this inhibition was associated with increased production of il-1 and nitric oxide ( no ) by macrophages . the treatment of macrophages with either nos inhibitors or prostaglandin e2 ( pge2 ) restored the invasive action of t. cruzi in macrophages previously treated with asa . lipoxin alx - receptor antagonist boc2 reversed the inhibitory effect of asa on trypomastigote invasion . our results indicate that pge2 , no , and lipoxins are involved in the regulation of anti - t . cruzi activity by macrophages , providing a better understanding of the role of prostaglandins in innate inflammatory response to t. cruzi infection as well as adding a new perspective to specific immune interventions .
PubmedSumm2053	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: to describe a patient with intrachoroidal cavitation in the normal eye that caused self - limiting recurrent macular detachment and retinoschisis . an 80-year - old female patient with intrachoroidal cavitation in the normal eye presented with macular detachment and retinoschisis after cataract surgery . one year after cataract surgery , the patient presented with macular detachment and retinoschisis in the same eye again . this case suggests that similar cases of cystoid macular edema after cataract surgery can occur , and that intrachoroidal cavitation is observed not only in eyes with pathologic myopia but also in normal eyes with peripapillary atrophy ; intrachoroidal cavitation can cause macular detachment and retinoschisis . macular detachment and retinoschisis are often associated with x - linked retinoschisis , pit - macular syndrome and coloboma [ 13 ] . however , recent studies have reported other causes of macular detachment and retinoschisis [ 47 ] . intrachoroidal cavitations are often observed in pathologic myopia , but not in normal eyes , and can result in retinoschisis and macular serous detachment [ 5 , 8 ] . other studies have shown peripapillary retinoschisis in glaucomatous eyes [ 9 , 10 ] , which resolved without treatment in some cases . here , we present a patient with intrachoroidal cavitation in the normal eye , with peripapillary atrophy ( ppa ) , that caused self - limiting recurrent macular detachment and retinoschisis . an 80-year - old woman presented with visual loss in her right eye to the sensho - kai eye institute . her best - corrected visual acuity measured 20/100 od and 20/16 os . in the right eye , a grade 2 nuclear cataract based on the emery scale was observed with an axial length of 23.80 mm and a refractive error of 1.0 diopters . the intraocular pressure measured 15 mm hg od and 14 mm hg os . no significant changes were observed in other parts of her eyes except small temporal conus . macular serous detachment and cysts were observed using rtvue-100 fourier - domain optical coherence tomography ( rtvue - oct ; optovue inc . , although rtvue - oct showed prominent serous detachment and cysts , fluorescent angiography showed slight pooling in the cystic space and no robust leakage comparable to configuration obtained by rtvue - oct ( fig . the eye was treated as cystoid macular edema with intravitreal bevacizumab ( 1.25 mg ) 2 months after surgery . the patient presented with decreased vision and central scotoma in her right eye to kyoto medical center . rs-3000 spectral - domain oct examination ( sd - oct ; nidek co. ltd . , tokyo , japan ) showed macular detachment and retinoschisis extending to ppa ( fig . intrachoroidal cavitations connecting to the cystic space were observed adjacent to the disc ( fig . the connection between the intrachoroidal cavitation and cystic space appeared as breakage of elschnig 's collagenous limiting tissue , which connects the choroid to the optic disc . since the patient refused further examinations , such as fluorescent angiography , sd - oct was used to monitor the condition monthly . serous retinal detachment and retinoschisis gradually absorbed , and was almost fully absorbed 4 months after onset ( fig . the left eye also showed intrachoroidal cavitation ; however , no incidence of retinoschisis or retinal detachment has been noted ( fig . no coloboma or optic disc pit was observed on ophthalmoscopy , but some ppa was found in the patient 's eyes . in her right eye , a yellow - orange lesion was also found that was reported as peripapillary detachment in pathologic myopia ( fig . 2 g ) . the hyporeflective space observed with sd - oct suggested the presence of fluid was not in large blood vessels but in the space of the intrachoroidal cavitation because no corresponding large vessels were visible in the ppa- area . the first incidence was resolved within 9 months with intravitreal bevacizumab , whereas the second incidence was resolved without treatment . this is similar to irvine - gass syndrome that is characterized by cystoid macular edema following cataract surgery . however , fluorescent angiography did not show the robust leakage associated with serous detachment and retinoschisis , suggesting another source of fluid like pit - macular syndrome could exist . retinoschisis appeared to extend to the disc , unlike that observed in common cystoid macular edema . recent studies have reported that peripapillary retinoschisis can be seen in glaucomatous eyes [ 9 , 10 ] and resolved without treatment . in our case , however , macular detachment and retinoschisis occurred immediately after cataract surgery , and may have therefore caused the fluctuation of intraocular pressure . the period of self - resolution of the second incidence was even shorter than that of the first . we have encouraged the patient to follow up periodically to assess any changes . although previous studies on peripapillary detachment in pathologic myopia showed small pit formation connecting the cystic space and vitreous , no such pit formation was observed in our case on thorough observation using sd - oct . the subretinal fluid in other cases may have been derived from the vitreous ; however , the fluid in our case may have been derived from intrachoroidal cavitation or spinal fluid . since there was no connection between vitreous and macular detachment , such a condition may facilitate self - resolution . previous studies have reported intrachoroidal cavitation or peripapillary detachment in pathologic myopia but not in normal eyes [ 47 ] . here , we reported a case of intrachoroidal cavitation in an eye with normal length and having ppa , resulting in self - limited recurrent macular detachment and retinoschisis . our case suggests that similar cases of cystoid macular edemas after cataract surgery exist , and that intrachoroidal cavitation is observed not only in pathologic myopia but also in normal eyes with ppa ; intrachoroidal cavitation can cause macular detachment and retinoschisis . in cases of cystoid edema following cataract surgery , we should carefully observe not only the macular area but also the peripapillary area . Output:	purposeto describe a patient with intrachoroidal cavitation in the normal eye that caused self - limiting recurrent macular detachment and retinoschisis.case reportan 80-year - old female patient with intrachoroidal cavitation in the normal eye presented with macular detachment and retinoschisis after cataract surgery . these were treated with intravitreal bevacizumab and then absorbed within 9 months . one year after cataract surgery , the patient presented with macular detachment and retinoschisis in the same eye again . these were absorbed within 4 months without treatment.conclusionthis case suggests that similar cases of cystoid macular edema after cataract surgery can occur , and that intrachoroidal cavitation is observed not only in eyes with pathologic myopia but also in normal eyes with peripapillary atrophy ; intrachoroidal cavitation can cause macular detachment and retinoschisis .
PubmedSumm2054	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: traditionally , the neisseria species has two potential pathogenic strains ; neisseria meningitidis and neisseria gonorrhoeae . the remainder of neisseria species , including neisseria mucosa , are considered normal flora colonizing the respiratory tract , living in symbiosis . one analysis showed that the majority of people are colonized by one to two of these neisseria species . specifically , neisseria mucosa is increasingly found to cause a multitude of diseases , in particular peritonitis . part of the difficulty in associating disease with other species of neisseria , such as neisseria mucosa , is the fact that they do not contain the typical virulence factors . the established pathogens neisseria meningitidis and neisseria gonorrhoeae derive much of their virulence from attaching to mucosal surfaces . however , the lack of these virulence factors does not necessarily make the other neisseria species benign . the advent of chronic ambulatory peritoneal dialysis ( capd ) has permitted significant patient autonomy with the concomitant risk of bacterial peritonitis . the typical culprits are gram - positive species , such as staphylococcus species and streptococcus species . we performed a pubmed literature search using the terms neisseria mucosa , peritonitis , and chronic ambulatory peritoneal dialysis . this yielded very few published cases of capd patients afflicted by neisseria mucosa peritonitis . in this clinical vignette , we add another case of capd associated with neisseria mucosa peritonitis . the patient was a 28 year - old hispanic man with a history of end stage renal disease due to hypertensive disease since the age of 10 years , complicated by malignant hypertension . his surgical history was remarkable for a remotely failed renal transplant requiring an allograft nephrectomy . he had been on hemodialysis for about a decade since , and transitioned to peritoneal dialysis just six months prior to presenting to our facility with sepsis due to new - onset peritonitis . in the outpatient clinic , the patient had been diagnosed with external cheek cellulitis that responded to a short course of empiric cephalexin . he overcame this only to experience three days of abdominal pain near his dialysis catheter . the pain was constant and severe pain , worsened by draining and filling during dialysis sessions . he denied any fevers , chills , nausea , vomiting , diarrhea , oropharyngeal and genitourinary complaints or decrease in recent ultrafiltration . exam was remarkable for a blood pressure of 167/99 mmhg , heart rate of 107 beats per minute and abdominal tenderness to palpation diffusely with the catheter exit site clear dry and intact without exudate or erythema . he quickly underwent peritoneal fluid examination , revealing an absolute neutrophil count of 14,458 cells / mm , confirming the diagnosis of bacterial peritonitis . his gram stain revealed gram - negative diplococci and subsequent chocolate agar plated cultures grew neisseria species , not gonorrhoeae or meningitidis . further biochemical speciation ( via rapid nh system by remel , thermo fisher scientific inc . , waltham , usa ) showed neisseria mucosa to be the etiology of the patient s bacterial peritonitis . given this is typically nonpathogenic , further susceptibility was not performed . the patient was placed on empiric intraperitoneal vancomycin 1 gram and cefepime 1 gram therapy and later transitioned to oral ciprofloxacin 500 mg daily for two weeks . during the hospitalization , the likely source of neisseria mucosa contamination of the dialysis port was the oral cellulitis . the transmission from the oral site to catheter site is supported by the well - known colonization of the oropharyngeal tract . capd seems to be associated with an increased risk for neisseria mucosa , particularly in cases of peritonitis . in general , this case adds to the current literature highlighting the importance of considering all types of disease caused by traditionally - benign neisseria mucosa species . although , nongonococcal and nonmeningococcal neisseria strains are usually benign and infections are rare , they may cause a variety of disease . this case highlights the danger of neisseria mucosa peritonitis in patients undergoing capd , and is the fourth reported case . the rarity of case reports and lack of the established pathogenic factors of nongonococcal and nonmeningococcal neisseria may limit recognition of these infections . this makes the association with clinical disease complex , and further investigation regarding their pathogenicity is warranted . in previously described case reports of nongonococcal and nonmeningococcal neisseria , our patient was also successfully treated with ciprofloxacin . given the increasing frequency of infections by nongonococcal and nonmeningococcal neisseria , guidelines regarding appropriate antimicrobial therapy should be established . further case reports should also establish the safety of preserving the peritoneal dialysis catheters , as in our patient s case . Output:	peritonitis is a leading complication of chronic ambulatory peritoneal dialysis . however , very rarely does neisseria mucosa cause peritonitis . we describe an unusual case of n. mucosa peritonitis in a chronic ambulatory peritoneal dialysis patient . a 28-year - old hispanic male presents with diffuse abdominal pain exacerbated during draining of the peritoneal fluid . peritoneal fluid examination was remarkable for leukocytosis and gramnegative diplococci . bacterial cultures were positive for n. mucosa growth . the patient was treated with ciprofloxacin with preservation of the dialysis catheter . this case highlights the rarity and importance of neisseria mucosa causing peritonitis in chronic ambulatory peritoneal dialysis patients. there seems to be a unique association between n. mucosa peritonitis and chronic ambulatory peritoneal dialysis patients. the patient was successfully managed with ciprofloxacin along with salvaging of the dialysis catheter .
PubmedSumm2055	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: good quality water is gradually getting beyond the reach of average households in developing countries . this is attributed to low income earnings vis - - vis the ever increasing cost of other sources of energy for water disinfectant such as kerosene and firewood . the use of chloride solution concentrate has been reported to be an effective means of cleaning vessels for water disinfection and prevention of waterborne diseases [ 2 , 3 ] . it was therefore suggested that efforts should be made to protect water after treatment up until the point of use . to achieve this , who suggested the use of residual chlorine of between 0.2 and 0.5 this is far from reality in developing nations due to the collapse of public water supply systems . most rural and semiurban settlements do not have the privilege of being connected to national electricity grid either . where they are connected , the erratic supply makes it imperative for cheaper and more reliable source of energy that could be used especially for water pasteurization to be sourced . solar energy is a free , inexhaustible , and environment - friendly resource [ 6 , 7 ] . it is estimated that one billion people worldwide do not have access to treated drinking water . harvesting of solar energy and its use may be a way out of this crisis , not only because of its availability in nigeria , but mainly due to its relative safety as well , compared to the use of fossil fuels . solar water heater can be used to pasteurize water thereby destroying harmful food - water microbes including bacteria and viruses when heated to temperatures of about 65c [ 10 , 11 ] . countries that could be considered as having potential for solar cooking must satisfy certain criteria , some of which include high insolation , fuel wood shortages , low per capital income , and high population density . these conditions are satisfied by nigeria with a ranking of five ( 5 ) in the list of countries with the highest potential benefit from solar cooking . contamination of surface and ground waters by pathogens and chemicals tends to make domestic roof - harvested rainwater an alternative . in the study environment , most households are in the habit of harvesting and storing rainwater in jars and cisterns to supplement other sources of potable water during the raining season . however , a small increase in temperature can improve the solar disinfection effectiveness for certain microbial strains . the populace is constantly faced with the problem of high cost of cooking fuels as well as diminishing wood supply . there is also the problem of overdependence on wood , leading to deforestation and its attendant problems of erosion and global warming . other alternatives such as the use of kerosene , gas , and electricity are unattractive because they are costly , unavailable , or unreliable . hence , this study investigated the use of locally available materials to develop solar heating system that can be used to improve the quality of domestic roof - harvested rainwater in nigeria and other developing countries . the study was conducted at the university of ibadan , ibadan , nigeria ( latitude 726n ; longitude 354e ) . solar water heating system was designed to handle continuous flow of water with a mean design temperature of 65c . the solar water heater developed and used in this work has five important components , namely , the outer box , the inner box , double - walled glass cover , reflector lid , and absorber plate . due to its local availability in some part of southwestern nigeria and low thermal conductivity , this was introduced to fill the space between the inner and outer boxes such that there is 100 mm thickness of insulating materials all around the inner box and between the inner and outer box . the insulating material was closely packed together at about 95.58 kg m density which is comparable to the average density of 112 kg m recommended by baryeh . the space housing the insulating materials was then sealed up with four pieces of plywood noggins . the solar water heating system was evaluated in an open field belonging to the nigerian micrometeorological experiments ( nimex ) research group , department of physics , university of ibadan , nigeria , for eleven days . a 15 m mast was set up to measure the profiles of the mean wind speed at 0.7 , 1.2 , 2.2 , 3.3 , 5.2 , 7.2 , 10.2 , and 14.8 m ( the mean wind direction is inclusive only at the 14.8 m height ) and air temperature ( wet and dry bulb ) at 0.9 , 4.9 , and 10.0 m. the same mast also supported radiation sensors for both global and net radiation at 1.5 m. the slow measurements were controlled by the use of two campbell cr10x data loggers which sampled the data every 1 second and subsequently stored them as 1-minute averaged value . a list of all the meteorological equipment used in this study is contained in table 1 . the arrangement was to monitor the rise in temperature of the absorber plate under no - load condition . hence , the maximum temperature attainable at a given solar radiation on a particular day can be determined . similarly , a pyranometer was used from the same station to monitor solar radiation . from standard procedure , thermal efficiency of a solar water heater or cooker is determined from water heating test using the relationship ( 1)=mwcw+mcuccutiat , where = thermal efficiency ( % ) , mw = mass of water ( kg ) , mcu = mass of the copper pipe ( kg ) , cw = specific heat capacity of water ( 4200 j kg k ) , ccu = specific heat capacity of copper pipe ( 400 j kg k ) , i = total solar radiation ( w m ) , a = area of absorber plate ( m ) , t = difference in temperature ( c ) , and t = difference in time ( seconds ) . a typical box - type heater requires adjustment every 15 to 30 min or when shadow appears on the absorber plate . the solar water heater was designed such that the orientation of the reflector lid ( when operated ) is facing the sunset ( west ) . this was found to be representative of local conditions , since the users will not likely have the time to stay with the system and be turning it every 15 to 30 min . the limitation of this fixed condition is that the system will only operate effectively between the hours of 11:30 a.m. and 4:30 p.m. on clear / sunny days . readings were monitored and recorded at 10 min interval , similar to the method used by mahavar et al . . most of the studies reported in the literature on solar disinfection used water with laboratory grown organisms subjected to simulated solar irradiation [ 17 , 22 , 23 ] . bacteria inactivation rate was reported to be slower for naturally occurring organisms compared with laboratory grown organisms . in this study , naturally contaminated drhrw in ibadan , nigeria , was harvested and released into the system at 2.61 10 m s ( 93.96 lh ) in a continuous flow process . water temperatures at the inlet , within the heating chamber , and at the outlet to the heating chamber and solar radiation were monitored at 10 min interval . samples were also collected into mccartney bottles at both the inlet to and the outlet from the heating chamber at 10 min interval for microbiological analysis . the total viable count was carried out by means of the standard plate count technique using plate count agar . dilutions of water samples in buffered peptone water were inoculated by putting 1 ml into each 10 ml molten standard plate count agar in mccartney bottles . after thorough mixing , these were poured into sterile petri dishes and incubated for 48 hours at 22c . petri dishes from dilutions counting 50 discrete colonies were counted and the results expressed as the number of bacteria colonies per millilitre . the isolates were further identified using their macroscopic , cultural , physiological , and biochemical characteristics . presumptive coliform test for the detection of coliform was done after the methods stated in . growth characteristics in methylene blue as well as reactions to indole , methyl red , voges - proskauer , and citrate utilization tests were used as confirmation of the presence of escherichia coli . the chemicals used in the study were of analytical grade and the preparation was done according to test guidelines ( [ 2426 ] , api 20e and api 20ne biomerieux , france ) . the schematic drawing for the design and fabrication of a flat plate collector , box solar water heater is presented in figure 2 . the design of the system is made such that the temperature is used up as soon as it is being built up by the incoming water that is constantly flowing through it ( which is at a lower temperature , relative to the temperature within the heating chamber ) . the procedure for the design and fabrication of the solar water heater is presented in figure 2 . design conditions are as follows : transparent surface area = 0.84 m 0.56 m = 0.47 m;absorber plate = 0.3 m 0.6 m black coated ( front and back ) ; aluminium sheet 1.4 mm gauge was used as the absorber plate;absorber pipe = 4.34 m length and 0.0064 m diameter , black coated , was used as the absorber / conveyance pipe.aluminium plate and copper pipe were used because of their high thermal conductivity , low weight per unit area , availability , affordability , workability , and good resistance to corrosion . corrosion of copper is most often associated with soft , acidic waters with ph below 6.5 . it is also a micronutrient needed by the body with a dietary value of 2 mg / kg . domestic roof - harvested rainwater used in this study has mean values of ph ranging from 6.8 to 7.6 , irrespective of roof types and those that were collected directly without contact with roof materials . also , a contact time of 7 min and a temperature of less than 80c are not sufficient to leach copper quantity that will cause toxic effect . transparent surface area = 0.84 m 0.56 m = 0.47 m ; absorber plate = 0.3 m 0.6 m black coated ( front and back ) ; aluminium sheet 1.4 mm gauge was used as the absorber plate ; absorber pipe = 4.34 m length and 0.0064 m diameter , black coated , was used as the absorber / conveyance pipe . aremu suggested that the base area should be smaller than the surface area and that the solar water heater should be shallow enough so as to avoid side shading effect . for maximum concentration at the base of the solar water heater dimensions of the inner box are as follows : surface area = 0.84 m 0.56 m;base area = 0.6 m 0.3 m;sides = 0.56 m 0.30 m 0.17 m and 0.84 m 0.60 m 0.17 m ( in pairs).the inner box was lagged 0.10 m on all sides . surface area = 0.84 m 0.56 m ; base area = 0.6 m 0.3 m ; sides = 0.56 m 0.30 m 0.17 m and 0.84 m 0.60 m 0.17 m ( in pairs ) . ( 2)dimension of the outer box=1.07 m0.76 m0.20 m. a reflector lid of dimension equal to the area of the lid was attached to one side of the lid , with a mirror of 0.91 m 0.61 m , so it may act as a booster to maximize solar radiation transfer into the box . for a flat plate collector , applying the first law of thermodynamics ( conservation of energy ) , ( 3)output energy qa = input energy quthe losses at equilibriumql . since the average energy from the sun is constant on a very clear day for a given locality , the energy transferred can be optimized by minimizing the heat loss ( ql ) component . for a double cover arrangement , stout reported that the energy input into the collector when the sun is at the zenith and its radiation is at right angle is 1 kwm given a cloudless sky and clear air . approximately 12% of the energy reaching each cover is reflected for each of the glass covers in the double cover arrangement . hence , for a solar water heater of effective surface area of 0.47 m , ( 4)qu=1000 wm20.880.880.47 m2=363.97 w. if cleaned and set at the correct angle , the reflector lid ( booster ) can reflect about 25% of the available insolation into the system . hence , ( 5)qu=363.97 + 25% of 363.97=454.96 w. total heat losses from the system are the sum total of heat losses from the bottom ( qb ) , sides ( qs ) , and covers ( qc ) given as ( 6)ql = qb+qs+qc , where ql = heat loss ( overall , w ) , qb = heat loss from the bottom ( w ) , qs = heat loss from the sides ( w ) , qc = heat loss from the cover ( w ) , ( 7)qb = ubat , where ( 8)ub = kl , k = thermal conductivity of lagging material = 0.0295 w m c , l = thickness of lagging material = 0.10 m , a = base area 0.60 m 0.30 m = 0.18 m , ( 9)qs = ubast , as = area of the sides ( m ) , ( 10)qc = utactqc.from duffie and beckman , ( 11)ut = n344/tptptan+f0.31 + 1hw1 n344/tptptan+f0.31 + 1hw1+(tp+ta)(tp2+ta2 ) p+0.0425n1p1 + 2n+f1ghhhp+0.0425n1p1 + 2n+f1gn1 , where hw = wind heat transfer coefficient = 5.7 + 3.8 v , n = number of glass covers ( 2 ) , g = emittance of glass ( 0.88 ) , p = emittance of plate ( 0.95 ) , tp = plate temperature.total heat loss was calculated to be 77.04 w. ql = heat loss ( overall , w ) , qb = heat loss from the bottom ( w ) , qs = heat loss from the sides ( w ) , qc = heat loss from the cover ( w ) , ( 7)qb = ubat , k = thermal conductivity of lagging material = 0.0295 w m c , l = thickness of lagging material = 0.10 m , a = base area 0.60 m 0.30 m = 0.18 m , ( 9)qs = ubast , as = area of the sides ( m ) , ( 10)qc = utactqc . hw = wind heat transfer coefficient = 5.7 + 3.8 v , n = number of glass covers ( 2 ) , g = emittance of glass ( 0.88 ) , p = emittance of plate ( 0.95 ) , tp = plate temperature . hence , output energy ( 12)qa=454.96 w77.04 w=377.92 w. the user can reckon on a total of 377.92 w , taking the losses into consideration . 1.16 w - hours is needed to heat up 1 litre of water by 1c . to heat up from ambient temperature of 30c to 65c , that is , temperature rise of 35c , 1.16 wh 35 = 40.60 wh . hence , 40.60 wh heat is required which will take 40.60 wh/377.92 w , that is , 0.11 hr ( 6.45 minutes ) . diameter ( d1 ) = 0.0127 m = d3 , d2 = 0.0064 m , length ( l1 ) = 3.8 m = l3 , and l2 = 4.34 m. neglecting form losses , ( 13)discharge q = q1=q2=q3 . the discharge through the system was then calculated to be 93.96 lh . to ensure the detention period of about 7 minutes within the solar panel , a serpentine or sinusoidal shape was assumed for the copper pipe used . on a clear day , the system can work effectively for about 4 hours on average , treating about 375.84 l of water . this amount should be enough to meet the minimum volume of 7.5 l per capital per day recommended by who for a family of six , for one week . the effect of temperature on microbial load of treated water with solar water heater is presented in table 2 while the percentage reductions in microbial load due to solar disinfection are presented in figure 3 . it attained a maximum operating temperature of 75c . at this temperature , a reduction of 89.6% in total viable count and 94.4% in total coliform count as low as 49c , the heating system was still able to achieve 41.7 , 33.3 , 20.0 , and 33.3% reduction in microbial load , respectively , for total viable count , total coliform count , e. coli , and s. aureus . had reported that temperature of about 60 to 70c can prevent permanent colonization of legionella spp . mcguigan et al . also observed that children ( 5 to 16 years of age ) who stored their drinking water in 1.5-litre plastic bottles that were placed in direct sunlight for continuous periods of not less than six hours in kenya experienced a 9% reduction in incidences of severe diarrhea over three months ' duration of the trial , compared with the control group . meera and ahammed , heaselgrave et al . , and mcguigan et al . reported the effectiveness of solar disinfection ( sodis ) in disinfecting water contaminated with total coliforms : cyst of giardia muris and oocysts of cryptosporidium parvum poliovirus and acanthamoeba polyphaga , respectively , at temperatures of 40 to 55c . the averagecost of fabricating a unit of the solar water heater at the time of this research was 32,161.76 ( $ 201.01 ) at the rate of $ 1 = 160 . this amount is equivalent to the cost of purchasing 402.02 l of bottled water in ibadan , nigeria , where the experiment was conducted . a bottle of 1.5 l of water is being sold for about 120 ( $ 0.75 ) in the area at the time of the experimentation . it would take the developed solar heating system 2 clear / sunny days to treat 402.02 l. hence , the cost of constructing this system would be recovered in only 2 clear / sunny days . an appropriate low cost solar heating system was developed and evaluated as a way of reducing microbiological contamination of domestic roof - harvested rainwater . the solar water heater recorded a maximum operating temperature of 75c with 89.6 and 94.4% reduction in total viable count and total coliform count , respectively , while e. coli and s. aureus were completely eradicated at this temperature . the drhrw in ibadan , nigeria , contains some contaminants and is therefore not safe for potable uses without treatment . the solar heater developed proved to be effective in improving the quality of drhrw . it is however not recommended for use in disinfecting water with ph less than 6.5 . Output:	a box - type solar heater was designed , constructed , and used to determine the effect of solar heating on quality of domestic roof - harvested rainwater ( drhrw ) . during testing , naturally contaminated drhrw was harvested in ibadan , nigeria , and released into the system at 93.96 lh1 ( 2.61 105 m3 s1 ) in a continuous flow process . water temperatures at inlet , within the heating chamber , and at outlet from the heating chamber and solar radiation were monitored at 10 min interval . samples were collected at both inlet to and outlet from the heating chamber at 10 min interval for microbiological analysis . the highest plate stagnation temperature , under no - load condition , was 100c . the solar water heater attained a maximum operational temperature of 75c with 89.6 and 94.4% reduction in total viable count and total coliform count , respectively , while escherichia coli and staphylococcus aureus were completely eradicated at this temperature . the solar heater developed proved to be effective in enhancing potability of drhrw in ibadan , nigeria . this may be an appropriate household water treatment technology for developing countries , hence , a way of resolving problem of low quality water for potable uses .
PubmedSumm2056	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: cells continuously encounter a wide variety of intrinsic and extrinsic stresses that damage the integrity of dna ( lindahl and barnes , 2000 ) . mitochondrial respiratory chain generates reactive oxygen species ( ros ) , which are the most prevalent intrinsic source of dna damage . extrinsic sources of dna damage are ionizing radiation ( ir ) , ultraviolet ( uv ) , and genotoxic chemical agents , including chemotherapeutic drugs ( e.g. , camptothecin , doxorubicin , and etoposide ) . both of the sources attack dna and produce dna lesions or breaks ( dipple , 1995 ) . if not properly repaired , these damages are capable of blocking dna replication and transcription , leading to mutations or genome aberrations , which consequently give rise to various human diseases , such as cancer . to maintain genome stability against the stresses , cells operate dna damage response ( ddr ) to detect dna lesions , signal their presence , and promote their repair ( harrison and haber , 2006 ; harper and elledge , 2007 ; rouse and jackson , 2002 ) . if ddr alone can not completely handle dna damage , cells employ an additional defense mechanism , called dna damage tolerance ( ddt ) that allows dna replication with the bypass of dna lesions , which eventually is repaired at a later stage ( sale et al . , 2012 ) . both ddr and ddt are tightly controlled by post - translational protein modifications ( ptms ) , including phosphorylation , acetylation , methylation , poly ( adp - ribosyl)ation , and modification by ubiquitin and ubiquitin - like proteins ( ubls ) . when cells are seriously damaged by genotoxic stresses and can not be repaired by ddr and ddt , they are then committed to kill themselves through a suicide process , called apoptosis . ptms are also critically involved in the control of apoptotic process for selective and permanent elimination of damaged cells . ubls are small - size polypeptides whose three - dimensional structures are strikingly similar to that of ubiquitin , although the similarity in their amino acid sequences to ubiquitin significantly varies ( kerscher et al . , 2006 ) . so far more than ten ubls have been identified and they include atg8 , fat10 , hub1 , isg15 , nedd8 , sumo(14 ) , ufm1 , and urm1 ( jentsch and pyrowolakis , 2000 ) . most ubls are conjugated to specific target proteins by a three - enzyme cascade system ( e1 , e2 , and e3 ) that resembles ubiquitination . protein modifications by ubiquitin and ubls are reversible processes that are catalyzed by isopeptidases , called deubiquitinating enzymes ( dubs ) and ubl - specific proteases ( ulps ) , respectively . these reversible protein modification processes play important roles in the regulation of key cellular processes , including cell proliferation , cell differentiation , and apoptosis . furthermore , deregulation of these modification systems results in a wide variety of human diseases , including cancer , neurodegenerative disorders , and immune diseases . of the ubiquitin family , the roles of ubiquitin , sumo , and nedd8 in the control of ddr and ddt as well as of apoptosis have been extensively studied and reviewed in detail ( brown and jackson , 2015 ; dantuma and van attikum , 2016 ; jackson and bartek , 2009 ; roos et al . , 2016 ) . however , relatively little is known about the function of isg15 under dna damage conditions . in this review , we overview the recent progress made in exploring the functional significance of isg15 and its reversible modification of target proteins in the regulation of cellular responses to genotoxic stress and thus in their implication in human diseases , particularly cancer . the product of interferon ( ifn)-stimulated gene 15 ( isg15 ) is the first reported ubl ( haas et al . , 1987 ) . this 15-kda protein consists of two ubiquitin - like domains corresponding to the n- and c - terminal regions , which share 29% and 31% amino acid sequence identities with ubiquitin , respectively ( haas et al . , 1987 ; isg15 is conjugated to target proteins by an enzymatic cascade : isg15-activating e1 enzyme ( ube1l ) , isg15-conjugating e2 enzyme ( ubch8 ) , and isg15 e3 ligases ( e.g. , herc5 and efp ) ( kim et al . , 2004 ; 2006 ; yuan and krug , 2001 ; zhao et al . , 2004 ; zou and zhang , 2006 ) . this conjugation process can be reversed by the major deisgylating enzyme ubp43 , also called usp18 ( malakhov et al . , 2002 ) . isg15 is not present in drosophila , caenorhabditis , and saccharomyces , indicating that its functions are restricted to higher animals . isg15 is robustly induced by type - i ifns , lipopolysaccharide , and viral infection ( farrell et al . , 1979 ; kim et al . , 2002 ; loeb and haas , 1992 ; malakhova et al . , 2002 ) one of the major components of the innate immune response is the activation of type i ifn signaling pathways ( platanias , 2005 ; taniguchi and takaoka , 2001 ) . proteomic analysis has revealed that regulatory proteins involved in antiviral ifn signaling pathways , such as rig - i , irf3 , mda-5 , stat1 , jak1 , and filamin b , serve as targets for isg15 modification ( arimoto et al . , 2008 ; jeon et al . , 2009 ; kim et al . , 2008 ; zhao et al . , 2005 moreover , isg15 and its conjugation to target proteins have been implicated in impairment of viral replication in vivo ( lu et al . , 2006 ; malakhova and zhang , 2008 ; okumura et al . , 2008 ) . conversely , certain viruses induce their own proteins that can block the expression of isg15 ( e.g. , ns3/4a ) or generation of isg15-conjugated proteins ( e.g. , ns1b ) or promote deconjugation of isg15 from its target proteins ( i.e. , deisgylating enzymes , including otu , plpro , and viral e3 protein ) , thus abrogating the antiviral immune responses ( frias - staheli et al . , 2007 ; lindner et al . , 2005 ; loo et al . , 2006 ; ratia et al . , 2008 ; yuan and krug , 2001 ; zhao et al . , 2016 ) . in addition , mice lacking isg15 or ube1l exhibit increased susceptibility to infection by certain viruses , including influenza b virus , herpes simplex virus , and/or sindbis virus ( giannakopoulos et al . , 2009 ; lai et al . , 2009 ; lenschow et al . , 2007 ) , further indicating the role of isg15 and its conjugation of target proteins in antiviral responses . uv was first shown to increase the level of isg15 transcripts by about four fold in human skin fibroblasts ( gentile et al . , 2003 ) . camptothecin , an inhibitor of topoisomerase i , also increases the level of isg15 mrna and this increase requires protein synthesis and a functional p53 protein ( liu et al . , 2004 ) . significantly , camptothecin - mediated induction of isg15 is independent of ifn signaling pathway and isg15 conjugates formed by the drug treatment are different from those generated by type i ifns , indicating that different signals induce isgylation of different target proteins ( park et al . , 2016 ) . moreover , the isg15 gene has a putative p53-responsive element ( p53re ) for its induction in addition to ifn - sensitive response element ( isre ) ( reich et al . , 1987 ) . collectively , these findings suggest that isg15 and its conjugated proteins participate in the control of cellular responses to dna damage . the p53 tumor suppressor coordinates cellular responses to dna damage as well as to other stresses , such as abnormal oncogene activation , telomere erosion , and hypoxia ( green and kroemer , 2009 ; riley et al . , 2008 ) . under normal conditions , the level of p53 protein is kept low by several e3 ligases - mediated ubiquitination . among them , mdm2 is the major ubiquitin e3 ligase that leads to degradation of p53 by proteasome . interestingly , the expression of mdm2 is induced by p53 , thus forming a negative feedback loop for down - regulation of p53 ( ashcroft and vousden , 1999 ; oliner et al . , 1992 , however , the interaction of p53 with mdm2 and other negative regulators is disrupted by phosphorylation and acetylation , leading to stabilization and activation of p53 . the activated p53 then binds to p53res for transcriptional activation of its target genes ( e.g. , bax , cdkn1 , and puma ) that mediate cell cycle arrest and/or apoptosis , depending on the degree of stresses ( el - deiry et al . , 1994 ; miyashita and reed , 1995 ; nakano and vousden , 2001 ) . recently , we have shown that p53re is present not only in the isg15 gene but also in the promoter regions of the genes encoding ube1l ( e1 ) , ubch8 ( e2 ) , and efp ( e3 ) , all of which are henceforth referred to as the isg15-conjugating system ( park et al . , 2016 ) . accordingly , treatment with dna - damaging agents , such as uv , camptothecin , and doxorubicin , markedly induces both the mrna and protein levels of ube1l , ubch8 , and efp in p53 cells , but not in p53 cells , and this induction can be abrogated by caffeine , an inhibitor of atm / atr kinases ( sarkaria et al . , 1999 ) , which phosphorylate chk1 and p53 for the expression of p53 . in addition , dna damage - mediated induction of the isg15-conjugating system is independent of type i ifns , indicating that p53 alone can positively regulate the expression of isg15 and its conjugation system . dna - damaging agents are capable of inducing isgylation of p53 as well as overexpression of the isg15-conjugating system ( park et al . , 2016 ) . lys291 and lys292 serve as the major isg15-acceptor sites in p53 . of two known isg15 e3 enzymes , efp , but not herc5 , acts as a p53-specific ligase . herc5 lacks p53re , consistently with the finding that the ligase is not induced under dna - damaging conditions . intriguingly , isgylation of p53 promotes its transcriptional activity and in turn in the expression of its downstream target genes , including cdkn1 , mdm2 , bax , and isg15 , as well as of its own gene . this increase of the p53 activity is mediated by the ability of isg15-conjugated p53 to promote its phosphorylation and acetylation and thereby to increase its affinity toward p53re . knockdown of isg15 or any of the isg15-conjugating system or lys - to - arg mutations of the isg15 acceptor sites in p53 strongly attenuates dna damage - induced p53 activity and in turn its tumor suppressive function ( park et al . , 2016 ) . thus , cells appear to operate a novel feedback circuit between p53 and the isg15-conjugating system for positive control of p53 tumor suppressive function under genotoxic stress conditions . different isotypes can be generated from their genes due to the presence of different promoters ( levine et al . , 2011 ) . for example , the p63 gene generates two types of transcripts : one for p63 having an n - terminal transactivation domain ( ta ) and the other for p63 lacking ta domain ( n ) . in addition , both ta and n transcripts are differentially spliced at their 3 ends to generate the p63 proteins with unique c - termini , termed , , , , and ( melino , 2011 ) . similar to p53 , tap63 isotypes can activate transcription from p53-responsive genes , which induce cell cycle arrest and apoptosis , thus also functioning as tumor suppressors ( flores et al . , np63 has the transactivation inhibitory domain ( ti ) but lacks the ta domain and therefore can dominant - negatively suppress transcriptional activation of the p53 family member by binding to their ta domains ( guo et al . , 2009 ; 1998 ) , contributing to its anti - apoptotic , mitogenic , and tumorigenic functions . np63 is the most abundant p63 isotype in many proliferating epithelial cells , such as mcf10a ( carroll et al . , significantly , its expression is frequently amplified in human epithelial cancers , such as squamous cell carcinomas , advanced cervical carcinomas , and human breast carcinomas , supporting its role in tumorigenesis ( hibi et al . , 2000 ; leong et al . , 2007 ) . dna - damaging agents , such as camptothecin and doxo - rubicin , induce isgylation of np63 in mcf10a and various epithelial cancer cell lines , including hnscc013 , hcc1937 , and fadu ( jeon et al . , 2012 ) . lys139 and lys324 serve as the isgylation sites in np63. upon exposure to the dna - damaging agents , caspase-2 is activated , although with an unknown mechanism(s ) , and cleaves off the ti domain from isgylated np63 , but not from its unmodified form , suggesting that isg15 molecules conjugated tonp63 act as molecular scaffolds for recruiting activated caspase-2 . asp452 , asp469 , and asp489 are the cleavage sites in np63. the cleaved ti domain is exported to the cytoplasm from the nucleus , thus losing its ability to bind the ta domain and inhibit the transcriptional activity of ta domain - containing p53 family members in the nucleus . under the same stress conditions , tap63 , is also isgylated and cleaved by caspase-2 and its ti domain is released to the cytoplasm , thus yielding a transcriptionally active form of tap63. furthermore , isgylation of np63 abrogates its ability to induce cell growth and tumor formation ( jeon et al . , 2012 ) . knockdown of isg15 , lys - to - arg mutations of isgylation sites , or asp - to - ala mutations of cleavage sites by caspase-2 strongly potentiate the ability of np63 to promote anchorage - independent cell growth and tumor development in vivo . these findings indicate that isg15 and its conjugation to np63 play critical roles in suppression of tumor - igenesis particularly in epithelial cancer cells under genotoxic stress conditions . as both camptothecin and doxorubicin are well - known anticancer drugs , these findings also provide a molecular basis for chemotherapeutic drugs against np63-mediated cancers . notably , cisplatin , unlike camptothecin and doxorubicin , is unable to induce the isg15-congugating system and np63 isgylation , although it also acts as a dna - damaging agent as well as an anticancer drug . however , cisplatin is capable of inducing cabl - mediated phosphorylation of tap73 , which causes the dissociation of tap73 from np63 and in turn the promotion of its transcriptional activity to induce apoptosis ( leong et al . , 2007 ) . thus , cisplatin , like camptothecin and doxorubicin , impairs the dominant - negative function ofnp63 toward ta domain - containing p53 family members , although it does not exhibit any effect on isgylation and caspase-2-mediated cleavage of np63 , unlike camptothecin and doxorubicin . the sliding clamp proliferating cell nuclear antigen ( pcna ) serves as a processivity factor as well as a platform for recruiting necessary components for dna replication . moreover , pcna is critically involved in dna lesion bypass by acting as a scaffold that recruits essential components for ddt ( moldovan et al . , 2007 ) , indicating that pcna plays an additional key role in the maintenance of genome stability and cell survival under dna damage conditions . when replicating cells encounter dna damage , pcna undergoes numerous ptms , such as ubiquitination and sumoylation ( bergink and jentsch , 2009 ; jackson and durocher , 2013 ; mailand et al . , 2013 ; ulrich and walden , 2010 ) . uv induces mono - ubiquitination of a highly conserved lys164 residue in pcna by the ubiquitin e3 ligase rad6-rad18 complex ( hoege et al . , 2002 ) . this pcna ubiquitination triggers the replacement of replicative dna polymerases , such as pol , by damage - tolerant y family of dna polymerases , including pol , for translesion dna synthesis ( tls ) ( bienko et al . , 2005 ; kannouche and lehmann , 2004 ; kannouche et al . , 2004 ; lehmann et al . tls polymerases bypass dna lesion and therefore dna replication can proceed without the need of removal of the damage and the risk of fork collapse ( sale , 2012 ; sale et al . , 2012 ) . however , tls polymerases lack proofreading activity and frequently incorporate incorrect nucleotides , therefore are potentially mutagenic ( loeb and monnat , 2008 ; matsuda et al . , 2000 ; sale , 2012 ; sale et al . , 2012 ) . thus , error - prone tls polymerases have to be released from pcna after dna lesion bypass for preventing excessive mutagenesis . upon dna damage by uv , pcna consisting of three identical subunits is first mono - ubiquitinated ( at lys164 ) for recruitment of pol and thus for initiation of tls , as previously documented in detail ( bienko et al . , 2005 ; kannouche and lehmann , 2004 ; kannouche et al . , 2004 ; lehmann et al . , 2007 ; stelter and ulrich , 2003 ) . after dna lesion bypass , the isg15 e3 ligase efp , but not herc5 , is tethered to mono - ubiquitinated pcna , and generates mono - isgylated pcna ( at lys168 ) , leading to formation of pcna conjugated with both ubiquitin and isg15 in different subunits ( park et al . , 2014 ) . isgylated pcna then recruits pcna - interacting peptide ( pip ) motif - containing usp10 for deubiquitination of pcna , which causes the release of pol from pcna for termination of tls . efp conjugates an additional isg15 to pcna at lys164 , thus forming pcna ligated with two isg15 molecules in the same subunit , likely for preventing mono - ubiquitination at lys164 and re - recruitment of pol. finally , ubp43 is induced at a later period and cleaves off isg15 molecules from pcna for reloading of replicative dna polymerases and resuming of dna replication . for this sequential modification of pcna ( i.e. , mono - ubiquitination , isgylation , deubiquitination , isgylation , and deisgyation ) , expression of the proteins responsible for the processes ( i.e. , rad6-rad18 , isg15 and efp , usp10 , and ubp43 ) is induced in order after uv irradiation , although it remains mysterious how the timely expression of each component of pcna modifications is regulated . moreover , knockdown of any of isg15 , efp , and usp10 as well as lys - to - arg mutations of the isgylation sites in pcna lead to a dramatic increase in uv - mediated mutation frequency with a decrease in cell survival ( park et al . , 2014 ) . thus , pcna isgylation appears to play a crucial role in termination of error - prone tls after dna lesion bypass for escaping from excessive mutagenesis and thereby maintaining genome stability . further , its conjugation to target proteins plays essential roles in innate immune responses , such as anti - viral and anti - microbial functions . recent studies indicate that isg15 and its conjugating system , including ube1l ( e1 ) , ubch8 ( e2 ) , and efp ( e3 ) , are also induced under dna damage conditions and this induction is positively regulated by p53 . moreover , p53-mediated increase in the levels of isg15 and its conjugating system leads to isgylation ofnp63 and pcna as well as itself . isgylation of p53 forms a positive feedback loop for its transcriptional activation , resulting in the expression of p53-downstream target genes for cell cycle arrest and apoptosis under dna damage conditions . on the other hand , dna damage - induced isgylation of np63 negatively regulates its anti - apoptotic and mitogenic functions by caspase-2-mediated destabilization . in addition , reversible pcna isgylation relays a signaling pathway to turn off error - prone tls after dna lesion bypass for suppressing uv - induced mutagenesis as well as for resuming normal dna replication . thus , it appears clear that isg15 and its conjugation to target proteins play a crucial function in the control of cellular responses to genotoxic stresses and in turn in suppression of dna damage - mediated tumor - igenesis . Output:	error - free replication and repair of dna are pivotal to organisms for faithful transmission of their genetic information . cells orchestrate complex signaling networks that sense and resolve dna damage . post - translational protein modifications by ubiquitin and ubiquitin - like proteins , including sumo and nedd8 , are critically involved in dna damage response ( ddr ) and dna damage tolerance ( ddt ) . the expression of interferon - stimulated gene 15 ( isg15 ) , the first identified ubiquitin - like protein , has recently been shown to be induced under various dna damage conditions , such as exposure to uv , camptothecin , and doxorubicin . here we overview the recent findings on the role of isg15 and its conjugation to target proteins ( e.g. , p53 , np63 , and pcna ) in the control of cellular responses to genotoxic stress , such as the inhibition of cell growth and tumorigenesis .
PubmedSumm2057	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: duodenal atresia and stenosis are rare causes of intestinal obstruction in the newborn , with a prevalence of 1 in 5,000 to 10,000 live births , with duodenal stenosis occurring less frequently than duodenal atresia.1 however , these intestinal tract anomalies occur significantly more frequently in patients with down syndrome . approximately 2.5% of infants with down syndrome are ultimately diagnosed with either duodenal atresia or stenosis.2 conversely , 24% of patients with duodenal atresia or stenosis carry a diagnosis of down syndrome.3 duodenal atresia invariably presents within the first few hours to days of life with persistent vomiting after all feeds . in general , the atresia occurs at the level of , or just distal to , the ampulla of vater.4 as a consequence , the emesis is frequently bilious and quickly brought to the attention of the medical provider . on the other hand , recurrent episodes of vomiting and failure to thrive are the most common presenting symptoms of duodenal stenosis.5 we report this case because of its unusual and delayed presentation despite the presence of high - grade stenosis . moreover , few endoscopic images of duodenal stenosis have been published in the literature , suggesting that this finding is uncommon , but providers need to be prepared to encounter this complication . a 5-month - old male infant with down syndrome presented to the emergency department with coffee - ground emesis . two weeks before admission , the patient had a single episode of emesis with dark streaks concerning for blood . he looked well on examination by his pediatrician and was started on lansoprazole with the plan to follow - up at our pediatric gastroenterology clinic . however , coffee - ground emesis recurred , and the infant was referred to the emergency department for evaluation . growth had been appropriate for a child with down syndrome . at the time of the emergency room evaluation , abdominal radiography showed a normal bowel gas pattern . on the day of admission , the patient underwent esophagogastroduodenoscopy using a gif - xp 160 endoscope ( olympus , tokyo , japan ) . there was mild erythema in the distal esophagus , and a dark brown substance was observed in the stomach , consistent with blood exposed to gastric secretions . 1a ) , with a tubular structure containing normal - appearing villous mucosa in the center ( fig . 1b ) . no lumen could be identified in this structure . an upper gastrointestinal series ( fig . 2 ) demonstrated a markedly dilated duodenal bulb and narrowing of the second portion of the duodenum ( arrows ) , confirming the suspicion of duodenal stenosis . intestinal malrotation was also identified at the time of surgery ; and therefore , the patient underwent ladd 's procedure . duodenal stenosis frequently presents with recurrent vomiting and failure to thrive . owing to its chronic and variable presentation it has been reported that the presentation and diagnosis of duodenal stenosis may be delayed to as late as adolescence.6 therefore , pediatric providers should maintain a high index of suspicion for duodenal stenosis , particularly in patients with down syndrome . our patient presented at the age of 5 months , outside of the immediate neonatal period , and with the atypical presenting sign of hematemesis . however , other studies have reported that hematemesis is the primary presenting symptom of duodenal stenosis and atresia in children.7,8 the etiology of hematemesis in these patients is not clear but may be related to esophageal or gastric irritation in the setting of gastric stasis , recurrent reflux , and vomiting . hematemesis has been better described in pyloric stenosis and is secondary to reflux esophagitis in these patients.9 the mechanism of duodenal stenosis may be similar . our patient showed erythema in the distal esophagus and had no evidence of duodenitis ; therefore , esophagitis was considered to be the most likely cause of bleeding . a typical finding of duodenal atresia on abdominal radiography in infants is the " double bubble " sign , which is marked gaseous distention of the proximal duodenum and stomach . duodenal stenosis is classically diagnosed by an upper gastrointestinal series , which demonstrates a dilated stomach and duodenal bulb and a narrowing in the duodenum at the location of stenosis.4 an upper gastrointestinal series was not used as the initial diagnostic modality in our patient because he reported hematemesis without a history of chronic vomiting as the primary presenting sign . diagnosis by endoscopy is unusual , and few endoscopic images of congenital duodenal stenosis have been published in the literature so far . once medically stable , the patients can undergo operative repair with duodenoduodenostomy performed with either side - to - side or diamond - shaped anastomosis . our patient underwent a diamond - shaped duodenoduodenostomy , which allows earlier postoperative feeding and a shorter hospital stay and is therefore the preferred procedure.10 in conclusion , patients with down syndrome are at a higher risk of gastrointestinal anomalies . even high - grade duodenal stenosis can present late and atypically in children , as observed in our patient with hematemesis at 5 months of age . therefore , pediatric gastroenterologists should maintain a high index of suspicion for congenital duodenal stenosis in older infants and children and should be familiar with the endoscopic appearance of this lesion . Output:	duodenal stenosis and duodenal atresia are well - known gastrointestinal anomalies in patients with down syndrome . although duodenal atresia presents early and classically with vomiting in the immediate neonatal period , the presentation of duodenal stenosis can be significantly more subtle and the diagnosis delayed . here , we describe the case of a 5-month - old male infant with down syndrome and delayed presentation of high - grade duodenal stenosis diagnosed endoscopically . pediatric gastroenterologists should include duodenal stenosis in the differential diagnosis of older infants and children with vomiting and should be familiar with the endoscopic appearance of this lesion .
PubmedSumm2058	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: as the population ages , interest in maintaining and improving older adults psychological well - being as an important indicator of quality of life , has received substantial scholarly attention in gerontological research . in light of this consideration , several scientific investigations have been conducted to find the biopsychosocial factors affecting psychological well - being in later life.1,2 in recent years , the role of attitudes toward one s own aging as a psychosocial determinant of older adults health has been studied.3 attitudes toward one s own process of aging , as a modifiable factor,4 refers to expectations about the personal experience of aging . in other words , the personal experience of aging refers to the way that individuals perceive their own aging.5 there is a growing body of literature that indicates that attitudes toward aging significantly affect older adults psychological well - being.6,7 for example , levy et al4 conducted a longitudinal study from 1977 to 1995 to investigate the impact of self - perceptions of aging on functional health over time . the authors found that more positive attitudes toward aging were significantly associated with better functional health , after controlling for baseline measures of functional health and several sociodemographic factors . the ohio longitudinal study of aging and retirement ( olsar)8 also showed that beliefs about aging predict cause - specific mortality . the olsar found that a positive attitude toward aging was significantly related to lower mortality rates among patients with respiratory problems . similarly , the results of a cross - sectional study conducted on a sample of 405 older adults at six elderly welfare centers in a metropolitan city in south korea showed that attitudes about aging were significant factors for enhancing life satisfaction.9 conversely , the findings from a random sample of 316 community - dwelling older adults in taiwan showed that negative attitudes toward aging contributed to depressive symptoms.10 overall , the existing research shows that self - perceptions of aging are independently correlated with physical and psychological health,11 life - satisfaction,12 and depression.13 although existing evidence shows physical and psychological benefits of positive attitudes toward one s own aging among older adults , it is worth mentioning that the role of attitudes toward aging on psychological well - being should also be investigated from the dyadic perspective because husbands and wives are interdependent and affect each other . kenny et al14 stated that people involved in dyadic relationships may influence each other s thoughts , emotions , and behaviors . to the best of our knowledge , no studies have examined the extent to which attitudes toward aging among older adults and their spouses may exhibit reciprocal benefits . therefore , the main aim of the present study is to examine the dyadic effects of attitudes toward aging on the psychological well - being of older couples in malaysia . in order to examine the potential dyadic effects of attitudes toward aging on the psychological well - being of older couples a family is viewed as an interdependent system , whereby each member reciprocally influences and is influenced by other family members thoughts , feelings , and actions . in other words , family members emotions , cognition , and behaviors affect other family members emotions , cognition , and behaviors.15 therefore , each family member has a psychological and emotional influence on other family members . according to the proposed model , the actor effect is explained in terms of how one s attitude toward the aging of older adults and their spouses influences one s own well - being . the partner effect is explained in terms of how older adults attitudes toward aging influence the well - being of their spouses , and vice versa . in addition , nonindependence is explained in terms of how older adults well - being is similar to spouses well - being , and how older adults attitudes toward aging are similar to their spouses attitudes toward aging . as illustrated in figure 1 , the arrows a and b demonstrate actor effects ; wherein the husband s attitude toward aging influences his well - being , and the wife s attitude toward aging influences her well - being . c and d , where the husband s attitude toward aging influences the well - being of his wife , and the wife s attitude toward aging influences her husband s well - being . the arrow e proposes an association between older adults and their spouses in terms of their attitudes toward aging . finally , arrow f suggests an association between the husband s well - being and the wife s well - being . according to the proposed model , which is in line with a previous study which mentioned that attitudes toward aging are significantly associated with the health and well - being of older adults,3,16,17 it is hypothesized that older adults attitudes toward aging significantly contribute to their own psychological well - being . in addition , in accordance with previous studies showing interspousal associations between well - being and health status,1820 this model will test the interdependence effects of attitudes toward aging and psychological well - being between spouses . the most important goal of this model is to test the partner effects of attitudes toward aging on the psychological well - being of older adults and their partners . in order to examine the potential dyadic effects of attitudes toward aging on the psychological well - being of older couples , we developed a theoretical model called the spousal attitude a family is viewed as an interdependent system , whereby each member reciprocally influences and is influenced by other family members thoughts , feelings , and actions . in other words , family members emotions , cognition , and behaviors affect other family members emotions , cognition , and behaviors.15 therefore , each family member has a psychological and emotional influence on other family members . according to the proposed model , the actor effect is explained in terms of how one s attitude toward the aging of older adults and their spouses influences one s own well - being . the partner effect is explained in terms of how older adults attitudes toward aging influence the well - being of their spouses , and vice versa . in addition , nonindependence is explained in terms of how older adults well - being is similar to spouses well - being , and how older adults attitudes toward aging are similar to their spouses attitudes toward aging . as illustrated in figure 1 , the arrows a and b demonstrate actor effects ; wherein the husband s attitude toward aging influences his well - being , and the wife s attitude toward aging influences her well - being . c and d , where the husband s attitude toward aging influences the well - being of his wife , and the wife s attitude toward aging influences her husband s well - being . the arrow e proposes an association between older adults and their spouses in terms of their attitudes toward aging . finally , arrow f suggests an association between the husband s well - being and the wife s well - being . according to the proposed model , which is in line with a previous study which mentioned that attitudes toward aging are significantly associated with the health and well - being of older adults,3,16,17 it is hypothesized that older adults attitudes toward aging significantly contribute to their own psychological well - being . in addition , in accordance with previous studies showing interspousal associations between well - being and health status,1820 this model will test the interdependence effects of attitudes toward aging and psychological well - being between spouses . the most important goal of this model is to test the partner effects of attitudes toward aging on the psychological well - being of older adults and their partners . the study population , consisting of 300 couples aged 50 years and older , was drawn from a community - based survey entitled poverty among elderly women : case study of amanah ikhtiar conducted in 2011 . the survey utilized a multistage stratified sampling technique to select couples from peninsular malaysia ( west malaysia ) . the research team went through each question and asked the enumerators to practice recording possible answers . completed questionnaires were checked by members of the research team to ensure complete responses to the questions . in certain cases , attitudes toward aging were measured using a four - item scale developed and validated by the authors . the items included : 1 ) older adults are respected for their knowledge and wisdom ; 2 ) i will have more time with family and friends when i grow old ; 3 ) i hope to be able to do what i want when i grow old ; and 4 ) old age is time for resting . each item was rated on a three - point likert scale , with responses ranging from 13 . the internal consistency for attitude toward aging ( using cronbach s alpha ) was 0.80 for women and 0.82 for older men . the findings from the exploratory factor analysis ( efa ) showed construct validity for both groups . the results among women revealed that the four items had factor loadings of more than 0.6 , and that collectively , all four items explained 62.6% of the variance in attitudes toward aging ( kaiser meyer olkin = 0.78 ; eigenvalue = 2.5 ) . among older men , the results from the efa also revealed one factor with an eigenvalue of 2.6 ( kaiser meyer olkin = 0.77 ) ; 64.9% of the total factor variance was explained by four items . the world health organization five well - being index ( who-5 ) was used to measure psychological well - being . i woke up feeling fresh and rested ; and my daily life has been filled with things that interest me . the theoretical raw score ranges from 0 to 25 , and is transformed into a scale with scores ranging from 0 to 100 ; higher scores suggest higher levels of psychological well - being.21,22 the scale has received good reliability and validity among older malaysians.23,24 in the present study , the who-5 showed strong internal consistency , with a cronbach s alpha of 0.89 for the female respondents , and 0.90 for the males . the efa established the unidimensionality of the scale , where the first factor with an eigenvalue of 3.46 explained 69% of the total variance for females . unidimensionality of the scale was also established for males , where the first factor with an eigenvalue of 3.5 accounted for 70.3% of the total factor variance . first , exploratory and confirmatory factor analysis and internal consistency reliability were applied to evaluate the psychometric properties of the scales used in the study . next , descriptive statistics ( for example , mean , standard deviation , and frequency ) were performed to provide a profile of the general characteristics of the sample . finally , the actor partner interdependence model ( apim ) using the statistical package for the social sciences ( ibm corporation , armonk , ny , usa)/amos version 20 ( europress software , cheshire , uk ) was used to perform dyadic analysis . the apim as a new statistical modeling technique , was proposed by a number of authors,14,2527 to explore how individuals independent variables simultaneously and independently contribute to their own dependent variables and to their partners dependent variables . this statistical approach is commonly used to study pairs of individuals nested within family systems , such as couples , siblings , and in parent child relationships , allowing researchers to better understand intra- and interpersonal effects simultaneously.14,26 the model allows researchers to study the impact of a individual factor on his or her own outcome variable ( actor effect ) , as well as on the outcome variable of the partner ( partner effect).28 the apim can be categorized into four types of sub - models , including the actor - only pattern , the partner - only pattern , the couple pattern , and the social comparison pattern . the actor - only pattern indicates that individuals outcomes are influenced only by their own characteristics , and that the partner does not have any effect on the outcome . the partner - only pattern suggests that individual s factors do not affect their own outcomes . in other words , the next pattern , which was adopted for the current study , is the couple pattern . this pattern suggests that the individual is as affected by his / her own characteristics as well as by his / her spouse s characteristics . another pattern that is slightly similar to the couple pattern is the social - comparison pattern . the most important difference between the couple pattern and the social - comparison pattern is that the actor effect in the social - comparison pattern is a positive predictor of a person s own outcomes , but his or her partner s effect is a negative predictor of the spouse s outcomes.29 in the present study ( couple pattern ) , the actor effect is the outcome obtained from the husbands/wives attitudes toward aging on their own psychological well - being . the partner effect is that a husbands / wife s attitudes toward aging affects the spouse s psychological well - being . the following indices were used to assess the adequacy of the model fit : the goodness - of - fit index ( gfi ) ; the root mean square error of approximation ( rmsea ) ; the comparative fit index ( cfi ) ; the standardized root square mean residual ; and the tucker according to hu and bentler,30 values of 0.06 and below for the rmsea and the standard - ized root square mean residual , values greater than or equal to 0.90 for the gfi , and values greater than or equal to 0.95 for the cfi and the tucker lastly , interdependence within spousal dyads on attitudes toward aging and psychological well - being were tested using the intraclass correlation coefficient ( icc).31 attitudes toward aging were measured using a four - item scale developed and validated by the authors . the items included : 1 ) older adults are respected for their knowledge and wisdom ; 2 ) i will have more time with family and friends when i grow old ; 3 ) i hope to be able to do what i want when i grow old ; and 4 ) old age is time for resting . each item was rated on a three - point likert scale , with responses ranging from 13 . the internal consistency for attitude toward aging ( using cronbach s alpha ) was 0.80 for women and 0.82 for older men . the findings from the exploratory factor analysis ( efa ) showed construct validity for both groups . the results among women revealed that the four items had factor loadings of more than 0.6 , and that collectively , all four items explained 62.6% of the variance in attitudes toward aging ( kaiser meyer , the results from the efa also revealed one factor with an eigenvalue of 2.6 ( kaiser meyer olkin = 0.77 ) ; 64.9% of the total factor variance was explained by four items . the world health organization five well - being index ( who-5 ) was used to measure psychological well - being . i woke up feeling fresh and rested ; and my daily life has been filled with things that interest me . the theoretical raw score ranges from 0 to 25 , and is transformed into a scale with scores ranging from 0 to 100 ; higher scores suggest higher levels of psychological well - being.21,22 the scale has received good reliability and validity among older malaysians.23,24 in the present study , the who-5 showed strong internal consistency , with a cronbach s alpha of 0.89 for the female respondents , and 0.90 for the males . the efa established the unidimensionality of the scale , where the first factor with an eigenvalue of 3.46 explained 69% of the total variance for females . unidimensionality of the scale was also established for males , where the first factor with an eigenvalue of 3.5 accounted for 70.3% of the total factor variance . attitudes toward aging were measured using a four - item scale developed and validated by the authors . the items included : 1 ) older adults are respected for their knowledge and wisdom ; 2 ) i will have more time with family and friends when i grow old ; 3 ) i hope to be able to do what i want when i grow old ; and 4 ) old age is time for resting . each item was rated on a three - point likert scale , with responses ranging from 13 . the internal consistency for attitude toward aging ( using cronbach s alpha ) was 0.80 for women and 0.82 for older men . the findings from the exploratory factor analysis ( efa ) showed construct validity for both groups . the results among women revealed that the four items had factor loadings of more than 0.6 , and that collectively , all four items explained 62.6% of the variance in attitudes toward aging ( kaiser meyer , the results from the efa also revealed one factor with an eigenvalue of 2.6 ( kaiser meyer olkin = 0.77 ) ; 64.9% of the total factor variance was explained by four items . the world health organization five well - being index ( who-5 ) was used to measure psychological well - being . i woke up feeling fresh and rested ; and my daily life has been filled with things that interest me . the theoretical raw score ranges from 0 to 25 , and is transformed into a scale with scores ranging from 0 to 100 ; higher scores suggest higher levels of psychological well - being.21,22 the scale has received good reliability and validity among older malaysians.23,24 in the present study , the who-5 showed strong internal consistency , with a cronbach s alpha of 0.89 for the female respondents , and 0.90 for the males . the efa established the unidimensionality of the scale , where the first factor with an eigenvalue of 3.46 explained 69% of the total variance for females . unidimensionality of the scale was also established for males , where the first factor with an eigenvalue of 3.5 accounted for 70.3% of the total factor variance . first , exploratory and confirmatory factor analysis and internal consistency reliability were applied to evaluate the psychometric properties of the scales used in the study . next , descriptive statistics ( for example , mean , standard deviation , and frequency ) were performed to provide a profile of the general characteristics of the sample . finally , the actor partner interdependence model ( apim ) using the statistical package for the social sciences ( ibm corporation , armonk , ny , usa)/amos version 20 ( europress software , cheshire , uk ) was used to perform dyadic analysis . the apim as a new statistical modeling technique , was proposed by a number of authors,14,2527 to explore how individuals independent variables simultaneously and independently contribute to their own dependent variables and to their partners dependent variables . this statistical approach is commonly used to study pairs of individuals nested within family systems , such as couples , siblings , and in parent child relationships , allowing researchers to better understand intra- and interpersonal effects simultaneously.14,26 the model allows researchers to study the impact of a individual factor on his or her own outcome variable ( actor effect ) , as well as on the outcome variable of the partner ( partner effect).28 the apim can be categorized into four types of sub - models , including the actor - only pattern , the partner - only pattern , the couple pattern , and the social comparison pattern . the actor - only pattern indicates that individuals outcomes are influenced only by their own characteristics , and that the partner does not have any effect on the outcome . the partner - only pattern suggests that individual s factors do not affect their own outcomes . in other words , the next pattern , which was adopted for the current study , is the couple pattern . this pattern suggests that the individual is as affected by his / her own characteristics as well as by his / her spouse s characteristics . another pattern that is slightly similar to the couple pattern is the social - comparison pattern . the most important difference between the couple pattern and the social - comparison pattern is that the actor effect in the social - comparison pattern is a positive predictor of a person s own outcomes , but his or her partner s effect is a negative predictor of the spouse s outcomes.29 in the present study ( couple pattern ) , the actor effect is the outcome obtained from the husbands/wives attitudes toward aging on their own psychological well - being . the partner effect is that a husbands / wife s attitudes toward aging affects the spouse s psychological well - being . the following indices were used to assess the adequacy of the model fit : the goodness - of - fit index ( gfi ) ; the root mean square error of approximation ( rmsea ) ; the comparative fit index ( cfi ) ; the standardized root square mean residual ; and the tucker the rmsea and the standard - ized root square mean residual , values greater than or equal to 0.90 for the gfi , and values greater than or equal to 0.95 for the cfi and the tucker lastly , interdependence within spousal dyads on attitudes toward aging and psychological well - being were tested using the intraclass correlation coefficient ( icc).31 the mean ages of the husbands and wives in this sample were 60.37 years ( 6.55 years ) and 56.33 years ( 5.32 years ) , respectively . the mean age difference between the spouses in this sample was 4.03 years ( 4.66 years ) , and men were , on average , older than their wives . the majority of the couples were living in traditional houses ( 82% ) . regarding educational attainment , 11.7% of women and 12.3% of their spouses had no formal education . in terms of educational homogamy , 64% of couples had the same level of education ( 6.3% no formal education ; 31% primary education ; and 26.7% secondary education ) . the mean score for attitudes toward aging was 9.53 ( 1.64 ) , with a possible range of 512 . the mean psychological well - being score among this sample was 60.49 ( 23.32 ) . actor effects were examined , while controlling for partner effects.14 the findings from amos that were used to determine the presence of actor effects revealed an acceptable model fit ( cmin / degrees of freedom [ df ] = 2.23 ; gfi = 0.91 ; cfi = 0.95 ; rmsea = 0.06 ) . significant actor effects emerged between attitude toward aging and psychological well - being for both wives ( critical ratio [ cr ] = 2.68 ; p<0.01 ) and husbands ( cr = 2.61 ; p<0.01 ) . as expected , the results that emerged when testing for actor effects revealed that attitudes toward aging significantly contributed to the psychological well - being of older men and women . the findings from amos revealed that the proposed model acceptably fits the data ( cmin / df = 2.42 ; gfi = 0.92 ; cfi = 0.94 ; rmsea = 0.07 ) . results of the apim indicated that older adults psychological well - being is significantly predicted by their spouses attitudes toward aging , among both older men ( cr = 2.75 ; p<0.01 ) and women ( cr = 2.70 ; p<0.01 ) . husbands attitudes toward aging significantly and positively predicted their wives well - being , while the wives attitudes toward aging significantly and positively predicted their husbands well - being ( table 2 ) . the results revealed a significant relationship between older adults attitude toward aging and the attitudes of their spouses ( icc = 0.59 ; p<0.001 ) , and a similar level of interdependence was noted for psychological well - being ( icc = 0.57 ; p<0.001 ) . in other words , with respect to attitudes toward aging and psychological well - being , people tended to be more like their dyad partners than to the average person in the sample as a whole . actor effects were examined , while controlling for partner effects.14 the findings from amos that were used to determine the presence of actor effects revealed an acceptable model fit ( cmin / degrees of freedom [ df ] = 2.23 ; gfi = 0.91 ; cfi = 0.95 ; rmsea = 0.06 ) . significant actor effects emerged between attitude toward aging and psychological well - being for both wives ( critical ratio [ cr ] = 2.68 ; p<0.01 ) and husbands ( cr = 2.61 ; p<0.01 ) . as expected , the results that emerged when testing for actor effects revealed that attitudes toward aging significantly contributed to the psychological well - being of older men and women . the findings from amos revealed that the proposed model acceptably fits the data ( cmin / df = 2.42 ; gfi = 0.92 ; cfi = 0.94 ; rmsea = 0.07 ) . results of the apim indicated that older adults psychological well - being is significantly predicted by their spouses attitudes toward aging , among both older men ( cr = 2.75 ; p<0.01 ) and women ( cr = 2.70 ; p<0.01 ) . husbands attitudes toward aging significantly and positively predicted their wives well - being , while the wives attitudes toward aging significantly and positively predicted their husbands well - being ( table 2 ) . the results revealed a significant relationship between older adults attitude toward aging and the attitudes of their spouses ( icc = 0.59 ; p<0.001 ) , and a similar level of interdependence was noted for psychological well - being ( icc = 0.57 ; p<0.001 ) . in other words , with respect to attitudes toward aging and psychological well - being , people tended to be more like their dyad partners than to the average person in the sample as a whole . the main purpose of this study was to examine the dyadic effects of attitudes toward aging on spouses psychological well - being . actor effects were tested while controlling for partner effects . as expected , attitudes toward aging had positive and statistically significant actor effects on psychological well - being for both husbands and wives . consistent with the results from past studies , which indicated that attitudes toward aging affect subjective well - being and life satisfaction among older adults,3,4,7,10,32,33 our study showed a positive association between attitudes toward aging and psychological well - being among older couples . attitudes toward aging appear to affect the psychological well - being of older adults through three mechanisms , including psychological , behavioral , and physiological pathways . the psychological pathway , where attitudes toward aging may negatively affect older adults well - being , is self - fulfilling prophecy ( sfp ) . the sfp is an initially false belief of a situation that may lead to a new behavior , which then renders the originally false conception as true.34 this suggests that peoples attitudes toward aging might affect their own aging process in a sort of self - fulfilling prophecy . positive attitudes about aging may be internalized , with resultant positive expectations and , in turn , positive self - perceptions.17,35 older adults with positive attitudes toward aging may assume that health problems are not a direct consequence of growing old , and that healthy practices are effective ; therefore , they are willing to seek treatment and healthy practices as a result . it is found that a negative attitude towards aging heightens cardiovascular response to stress . repeated elevations of cardiovascular response to stress increases susceptibility to cardiovascular events and consequently contributes to general low well - being.35 the partner effects of attitudes toward aging on the psychological well - being of older adults and their partners using the apim model found that there were significant partner effects of attitude toward aging on their partners psychological well - being . this finding contributed new knowledge about the impact of attitudes toward aging on psychological well - being among older adults and their partners . this finding was slightly similar to the findings obtained from a study conducted by kim et al36 on a sample of 168 married survivor the researchers found that when a couple was dealing with a major illness , the extent to which women psychologically adjusted to the situation played a key role not only in their own well - being , but also in their spouse s well - being.36 it was concluded that positive attitudes toward aging would benefit both partners . they found that one s psychological adjustment to a stressor was affected both by one s own perceptions and by the partner s perceptions . similarly , the findings from a study conducted by berg and wilson37 on a sample of 104 infertile couples showed that one s well - being could be affected not only by his / her own perceptions , but also by the partner s perceptions . the results of the interdependence analysis revealed a significant association between older adults attitudes toward aging and the attitudes of their spouses ; there was a similar level of interdependence for psychological well - being as well . these findings expand upon those of previous studies , in which researchers found evidence for interdependence within elderly spousal dyads in measures of well - being18,20 and depressive symptoms.38,39 for example , bookwala and schulz18 investigated 1,040 elderly married couples and found that indicators of well - being for one spouse , such as perceived health , depressive symptoms , feelings about life as a whole , and satisfaction with life were significantly correlated with the well - being of the other spouse . similarly , the results of a sample of 2,235 spousal dyads showed that individuals perceptions of well - being tend to be similar to those of their spouses.20 tower and kasl,40 in their study on a sample of 317 community - dwelling older couples , found that depressive symptoms in one spouse had a significant impact on depressive symptoms in the other spouse . these findings are consistent with the tenets of family system theory , where each family member has a psychological and emotional influence on other family members ; in addition , these elements reciprocally influence and are influenced by other family members thoughts , feelings , and actions.15 according to lewis et al42 and wilson,43 the interrelated psychological well - being of older spouses may come from several paths . first , people tend to marry people who are similar in health , healthy behaviors , and health determinants ( such as socioeconomic factors ) . third , couples engage in certain similar habits and behaviors that reduce their odds of developing health problems . in addition , the interdependence of spouses psychological well - being may be influenced by the events experienced by the couple as a unit.44 although the apim approach guiding this research is a notable strength of this investigation , the findings should be considered in view of some limitations . first , the cross - sectional nature of the study makes causality difficult to establish . therefore , longitudinal studies are needed to better explore the causal direction of the effects observed in this study . the second limitation that should be addressed in future research is the effect of exogenous and confounding variables , which may affect the model . although the apim approach guiding this research is a notable strength of this investigation , the findings should be considered in view of some limitations . first , the cross - sectional nature of the study makes causality difficult to establish . therefore , longitudinal studies are needed to better explore the causal direction of the effects observed in this study . the second limitation that should be addressed in future research is the effect of exogenous and confounding variables , which may affect the model . collectively , the results of the present study offer new insights into the effects of attitude toward aging and psychological well - being in later life . attitudes toward aging appear to predict not only an individual s own psychological well - being , but also that of his or her spouse . therefore , spouses should be aware that their perceptions of aging affect their partners psychological well - being . the proposed model , which hypothesized that an older adult s perception of his or her own aging significantly affected his or her own , and his or her spouse s , psychological well - being was supported . it will be valuable to extend and test the generalizability of the proposed model of the spousal attitude future research should also continue to study the dyadic effects of other factors that may affect psychological well - being in later life . despite the limitations of this investigation , the current study has implications for research and practice . the findings highlight the importance of considering both intrapersonal and interpersonal factors that may influence older adults psychological well - being . in addition , programs for older adults and their spouses should be developed in order to enhance positive attitudes toward aging ; this may facilitate successful adaptation in old age , and maximize an older adult s psychological well - being . Output:	backgroundthere is a growing body of literature indicating that attitudes toward aging significantly affect older adults psychological well - being . however , there is a paucity of scientific investigations examining the role of older adults attitudes toward aging on their spouses psychological well - being . therefore , the present study aimed to examine the dyadic effects of attitude toward aging on the psychological well - being of older couples.methodsdata for the present study , consisting of 300 couples aged 50 years and older , were drawn from a community - based survey entitled poverty among elderly women : case study of amanah ikhtiar conducted in peninsular malaysia . an actor partner interdependence model using amos version 20 ( europress software , cheshire , uk ) was used to analyze the dyadic data.resultsthe mean ages of the husbands and wives in this sample were 60.37 years ( 6.55 ) and 56.33 years ( 5.32 ) , respectively . interdependence analyses revealed significant association between older adults attitudes toward aging and the attitudes of their spouses ( intraclass correlation = 0.59 ; p<0.001 ) , and similar interdependence was found for psychological well - being ( intraclass correlation = 0.57 ; p<0.001 ) . the findings from amos revealed that the proposed model fits the data ( cmin / degrees of freedom = 3.23 ; goodness - of - fit index = 0.90 ; confirmatory fit index = 0.91 ; root mean square error of approximation = 0.08 ) . results of the actor partner independence model indicated that older adults psychological well - being is significantly predicted by their spouses attitudes toward aging , both among older men ( critical ratio = 2.92 ; p<0.01 ) and women ( critical ratio = 2.70 ; p<0.01 ) . husbands and wives own reports of their attitudes toward aging were significantly correlated with their own and their spouses psychological well-being.conclusionthe findings from this study supported the proposed spousal attitude well - being model , where older adults attitudes toward aging significantly affected their own and their spouses psychological well - being . the theoretical and practical implications of the findings are discussed .
PubmedSumm2059	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: different individuals use different strategies for coping with negative affective state and associated life problems . such coping mechanisms are important both in periods of acute stress / emergencies ( such as hurricane disasters ) as well as in patients suffering from chronic illnesses such as depression , breast cancer , and hiv / aids . for example , in a study on the coping mechanisms and depression in elderly medically ill men , a high proportion of the respondents sought comfort in religious beliefs and practices . this in turn was inversely related to their severity of depression . on the other hand , the use some of these coping styles , such as substance use , may be termed as maladaptive and may result in poorer health outcomes for the patient . in chronic diseases , such as depression and anxiety , knowledge of these coping styles by the treating clinician / psychiatrist we , therefore , studied the frequency of these different coping styles in patients with symptoms of anxiety and depression presenting to primary health care settings of pakistan . to identify and estimate the frequency of the different coping mechanisms used by patients with symptoms of anxiety and depression . to estimate the prevalence of anxiety and depression in patients attending three primary health care settings of karachi , pakistan . a cross - sectional survey was conducted at three primary health care centers between september to october 2004 . the three primary health care centers at hijrat colony , sultanabad , and rehri goth were chosen because they represent the semiurban and rural settings of karachi and pakistan . additionally easy access and complete demographic profile of the community were available because of an ongoing urban health program ( uhp ) at these centers by our hospital . a team of seven medical students were trained in the use of questionnaire for assessing depression , anxiety , and coping strategies . the questionnaire had 4 parts and collected information on basic sociodemographic characteristics , the aga khan university anxiety and depression scale ( akuads ) , 28-item brief cope inventory , and other questions relating to aspects such as substance abuse , self - medication , and religious practices . the scale is an indigenously developed screening instrument to be used in the community for the assessment of psychiatric morbidity in the population . each question gives 4 choices to the respondent ( never : 0 , sometimes : 1 , mostly : 2 , and always : 3 ) and the total score is determined by adding the scores for each of the responses . at a score of 20 the tool has been used in a variety of settings and the validity of the instrument is also more than the previously used instruments , as it includes both the psychiatric and somatic aspects of the disease , hence making it a suitable tool to be used in the community setting . coping strategies employed in the past 1 month by the participants screened as having depression and anxiety were determined with the use of brief cope inventory . the brief cope developed by carver at the university of miami is one of the most commonly used coping measures and has been cited by more than 900 articles as of august 2011 . it is the abridged version of the original cope inventory and assesses 14 coping types with 28 questions ( 2 questions per type ; see figure 1 ) . ( i have been taking action to try to make the situation better ) , religion ( i have been using alcohol or other drugs to make myself feel better ) . since most of the participants presenting to primary health care settings are not literate , these items were rephrased into questions so that the questionnaire became interviewer based . for example , for substance use , have you been using alcohol or other drugs to make yourself feel better ? these items were then translated into urdu by authors p. m. kasi and u. z. khan . a back translation was then made by fahd khalid syed and compared with the original version . the final version was then given for approval to naqvi sahab ( learning resource center , aga khan university ) to check the validity of the translated items in urdu language . the responses to these questions are measured on a 4-point likert - type scale with responses ranging from 1 ( i 've not done this at all ) to 4 ( i 've been doing this a lot ) . the scores ( ranging from 2 to 8) and the means for each coping method were then calculated . although each of these coping strategies overall can not be termed as adaptive or maladaptive and are dependent on the context and situation , the suggested grouping used in some of the previous studies is outlined in figure 2 . it is important to note that the author of the brief cope tool did not attempt to dichotomize the coping scales into positive or negative coping strategies , as this task has been left to the researchers utilizing the brief cope tool and has been reported in various ways in the literature . more women report to these primary health care centers , thus their number gets higher ( 74.1% ) . the areas interviewed represent the underprivileged class of the society , where illiteracy was seen to be high ( 52.5% ) , with more than half of the families having a monthly income of less than rs . 5,000 ( $85 us ) . other surrogate markers of socio - economic status , such as the type of house and house ownership , which have been used in the national health survey were also used and the results shown . out of 162 people interviewed , 55 individuals screened positive for anxiety and depression ( akuads score 19 ) . females were significantly more likely to have symptoms of anxiety and depression than men ( p value = 0.024 , or = 2.62 ) . on the other hand , marital status , mother tongue , and other measures of socioeconomic status out of the 55 individuals who had symptoms of anxiety and depression , 3 did not fill the cope analysis of coping styles was , therefore , carried out on the results of 52 individuals . of the 14 coping styles studied , the most frequently used strategies were religion ( 48.1% ) , acceptance ( 34.6% ) , use of instrumental support ( 32.7% ) , active coping ( 30.8% ) , planning ( 28.8% ) , and use of emotional support ( 28.8% ) , using the particular coping style on a moderate ( i do this most of the time ) to frequent ( i do this all the time ) basis ( figure 3 ) . the other 8 coping styles were used less frequently by individuals with symptoms of anxiety and depression , with humor ( 9.6% ) , behavioral disengagement ( 7.7% ) , and substance use ( 5.8% ) being used least frequently ( see also table 3 ) . with respect to religion as a coping mechanism , we also inquired on the particular method used . as shown in figure 4 , 75% of the individuals with anxiety and depression found comfort or were praying to feel better , and more than half sought comfort by recitation of the holy book ( quran ) or by frequent remembrance of god ( tasbeeh ) . it is also interesting to note that a quarter of the individuals had also visited a faith healer for their symptoms . taweez , also a remedy to problems given by faith healers , was sought by 27% of the individuals with symptoms of anxiety and depression . across the three primary health care centers , we found the prevalence of anxiety and depression to be around 34% . this is similar to studies done in similar settings in pakistan , with the prevalence ranging between 30.4 to 38.4% [ 6 , 7 ] . a recent review in bmj on anxiety and depressive disorders in pakistan also estimated the overall prevalence to be around 34% ( 2966% for women and 1033% for men ) . we studied the frequency of different coping styles and found religious coping as the most common behavior , with significant proportion of individuals finding comfort and relief in different religious practices . a probable explanation for lowest scores on questions of substance abuse , that is , use of alcohol or other drugs to feel better or help the individual get through his / her symptoms , is first , and the data represents mainly women of low socio - economic strata of pakistan . men are more likely to be involved in substance abuse , and alcohol in particular is more in use in the higher socioeconomic classes of pakistan . islam also prohibits alcohol , and all individuals , except one , in our study were muslims . it is , however , interesting to note that there is much benzodiazepine abuse , where most of these drugs are available over the counter , with many individuals self - medicating . when asked about whether or not the person self - medicated to feel better , we found 30.8% have been doing so , with 19.2% of the individuals doing this on a moderate to frequent basis we have not looked into the drugs they were using but a center - based study in our hospital by khawaja et al . ( personal communication ) found this to be close to 3040% , which is also consistent with our findings [ 9 , 10 ] . it can , therefore , be suggested that in our context self - medication should be studied as a separate coping mechanism and the treating physician / psychiatrist should be aware if his patient is doing so . coping mechanisms are also associated with the patient 's understanding of his disease / symptoms and the ways in which he manages his illness . for example , endorsement of depressive symptoms was found to be associated with more self - blame and emotional venting . perceived negative consequences of depression led to more active coping , religious coping , and self - blame , whereas perception of disease as chronic led to less planning on the part of the patient . coping behaviors are thus affected by a number of parameters and being aware about your patients ' coping styles to manage his depression could in turn guide you while providing him with treatment . similarly , active coping was associated with lower levels of anxiety in accident and emergency house officers . coping by venting , on the other hand , produced greater levels of anxiety and depression . the weaknesses of the current study include the cross - sectional design of the study and employing tools that use self - reported data to assess the severity of symptoms and the frequency of usage of a particular coping strategy . use of screening tool ( akuads ) to establish depression and further assess its severity is itself another limitation . currently in developing countries like pakistan , there is dearth of mental health professionals and services . for an estimated population of more than 150 million , which is mostly rural , there are only 342 practicing psychiatrists , with most of them located in major cities . majority of patients present to general physicians and other specialists . in the light of previous studies , it may , therefore , be important to empower patients with positive coping styles or discourage negative coping styles to improve their overall quality of life , thus providing clinicians / psychiatrists with a useful tool as coping behaviors are modifiable . Output:	different individuals use different coping styles to cope with their problems . in patients with anxiety and/or depression , these have important implications . the primary objective of our study was to estimate the frequency of different coping mechanisms used by patients with symptoms of anxiety and depression . a descriptive , cross - sectional survey was conducted and patients with symptoms of anxiety and depression were identified using the aga khan university 's anxiety and depression scale ( akuads ) . coping styles were determined by using the 28-item brief cope inventory . we were able to recruit 162 people . the prevalence of anxiety and depression was found to be 34% . females were more than 2 times likely to have anxiety and depression ( p value = 0.024 , or = 2.62 ) . in patients screening positive for akuads , religion was the most common coping mechanism identified . acceptance , use of instrumental support , and active coping were other commonly used coping styles . our findings suggest that religious coping is a common behavior in patients presenting with symptoms anxiety and depression in pakistan . knowledge of these coping styles is important in the care of such patients , as these coping methods can be identified and to some extent modified by the treating clinician / psychiatrist .
PubmedSumm2060	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: pneumonia is the second cause of hospital acquired infections after catheter site infection that leads to prolonged hospital stays , increased morbidities and higher expenses[1 , 2 ] . ventilator associated pneumonia ( vap ) is the most common kind ( 86% ) of hospital acquired pneumonia . according to cdc definitions , vap is an episode of pneumonia that develops more than 48 hours after initiation of mechanical ventilation[3 , 4 ] . since the most important mechanism of vap is aspiration of colonized secretions in the nasopharynx , many attempts have been done to reduce colonization in trachea and nasopharynx[512 ] . in general , diagnosis of vap other diagnostic tools like clinical , laboratory , imaging findings and examining the sputum qualitatively and quantitatively are limited too . thus neonatologists utilize a combination of these techniques to diagnose vap according to their experience and setting . newborns are more prone to vap because their mucosal layers are delicate , and their immune system is not completely functioning[2 , 13 ] . it is stated that low birth weight , ventilators , frequent tracheal suctioning , sedation with opioids , and reintubation are directly associated with the incidence of vap . recent data suggest that during mechanical ventilation , lateral positioning ( in which the endotracheal tube is horizontal ) may decrease the incidence of bacterial colonization of endotracheal tube[1619 ] . the objective of this study was to evaluate the influence of lateral and supine position on bacterial colonization of endotracheal tube in neonates . we conducted a prospective , randomized , clinical trial in the neonatal intensive care unit of 17 shahrivar children 's hospital of rasht ( rasht iran ) . thirty - one intubated neonates ( intubated within 48 hours after birth ) were included ; 16 neonates were positioned supine ( supine group ) , and 15 were maintained in the lateral position ( lateral group ) . sample size was chosen based on a similar study , indicating that 87% of neonates in supine position versus 30% of neonates in lateral position had bacterial colonization after 5 days . so , two groups of 10 neonates would suffice to draw a conclusion . to increase the accuracy of the study , by increasing attrition rate , 31 newborns ( 16 in the supine and 15 in the lateral group ) participants were randomized by random fixed block method into two groups of a ( neonates in supine position ) and b ( neonates in lateral position ) . inclusion criteria of this study were : 1 ) gestational age 28 weeks from the last menstrual period , 2 ) tracheal intubation at postnatal age of < 48 hours , 3 ) absence of congenital malformations such as tracheoesophageal fistulae , chest deformities or diaphragmatic hernia . patients were excluded if they were diagnosed as congenital sepsis or pneumonia or they could not be maintained on mechanical ventilation for 5 full days . we recorded demographic data and the results of routine laboratory tests that were performed for all neonates at the admission time in nicu from their files such as cbc , platelets , crp , abg ( arterial blood gas ) , blood cultures , and cxr . patients were ventilated in simv mode using babylog 8000-plus drager ventilator with heated humidification system . open method of suction was used for suctioning secretions as needed and no prophylactic topical oropharyngeal antibiotics and selective gut decontamination was done in any of the patients . neonates in group a were maintained on their backs and endotracheal tube was held upright in a vertical position and their bed was kept horizontal with 10 degree head tilt ( routine care in our nicu ) . group b subjects were positioned on their sides with the back supported by a rolled soft towel and they were alternatively rotated to left or right lateral position by trained nurses every two hours . tracheal aspirates of both groups were cultured in second and fifth days of mechanical ventilation . for this purpose the researcher wore sterile gloves and suctioned the endotracheal tube , using feeding tubes no 5 or 6 , without injecting any solution into the endotracheal tube and instantly transferred the samples into an emb agar and blood agar medium . cultures were incubated for 24 hours in 37c . a single microbiologist blind to the grouping analyzed the dishes and studied the antibiogram . abg variables and ventilator setup were recorded at second and fifth days of intubation from patients files too ( abg was performed for all intubated neonates daily in our setup ) . data were analyzed with spss version 16 by tests of chi square , t - test and repeated measure anova . the study protocol was approved by the ethical committee of guilan university of medical sciences . in the second day of ventilation positive cultures were recognized in 6.2% ( 1 neonate ) of supine group and 6.7% ( 1 neonate ) of lateral group . after 5 days tracheal cultures were positive in 25% ( 4 neonates ) of supine group and 13.3% ( 2 neonates ) of lateral group that was n't statistically significant ( p=0.9 in second and fifth day ) . the most common organisms isolated from tracheal aspirates were gram - negative rods ( klebsiella ) . table one gives comparison of weight , gestational age , and hours after birth , in the two groups . there was not statistically difference between the two groups about initiation of feeding ( p=0.3 ) , use of h2 blocker ( p=0.9 ) and rate of reintubation ( p=0.6 ) . because all the patients had respiratory distress at the time of admission , they all received antibiotic treatment according to unit policy . except one patient with pneumothorax , other patients were intubated because of hyaline membrane disease . comparison of interfering parameters between supine and lateral position groups sd : standard deviation table 2 shows the means of lab data of both groups in the first day of hospitalization . we compared some abg variables and ventilator setup at the beginning , second day , and fifth day of mechanical ventilation in the two groups by t - test . we also compared the trend in change of abg variables and ventilator setup in both groups using repeated measure anova . the only exception was the means of po2 of the second and fifth day of ventilation that was higher in the supine group ( table 3 ) . means of lab data of both groups in the first day of hospitalization sd : standard deviation means of abg variables of second and fifth day after the initiation of mechanical ventilation abg : atrial blood gas ; sd : standard deviation in our study bacterial colonization in endotracheal tube was not different in the two groups after two days . colonization increased by the fifth day in both groups ( 6.7% to 13.3% in group b and 6.2% to 25% in group a ) . the rate of increase in colonization from the second to the fifth day is higher in the supine positioned group ( group a ) . aly et al also found an increased rate in the supine group yet it was statistically significant . they selected 50 newborns with day five colonization count of 87% in the supine group and 30% in lateral positioned group . our selection was 31 newborns with five day colonization count of 25% ( a ) versus 13% ( b ) . the reason of this discrepancy is probably due to difference in study setting in terms of hand washing routines , lab techniques and hygiene . it is to be noticed that some of our participants with the second day positive endotracheal cultures could not finish the study because of subsequent sepsis or pneumonia and death . therefore , the real rate of colonization is expected to be higher than we found . when the endotracheal tube is in vertical position , respiratory secretions passively enter to the lower airways . therefore , eliminating the effect of gravity by maintaining the endotracheal tube in horizontal position should make respiratory secretions less likely to move down the airway . in addition , we think that such positioning of the trachea facilitates sliding of respiratory secretions along the endotracheal tube wall and decreases the need for tracheal suctioning . aly et al stated that reintubation rate in the supine group was more in their study and they mentioned this as one of the limitations of their study . we can consider our groups to be more homogeneous in terms of newborns sex , hours from birth , gestational age , weight , type of delivery , and all laboratory indices . the only exception was hb level that was more in the supine group ( group a ) ( mean hb level in group a was 15.88 g / dl and it was 14.36g / dl in group b which was significantly different ( p<0.02 ) ) . in other words , positioning did not have any effect on po2 level because the supine group started the study by a higher level of hb . differences in initiation of feeding and h2 blocker would adversely affect the result of the study , our groups were homogeneous in terms of initiation of feeding and h2 blocker . balagur et al by a metaanalysis of 11 studies on the effect of safety in different positioning concluded that lateral position did not have any advantage over other positions , yet prone position had a better po2 level when compared with supine position . ventilator setup was the same in both groups and fio2 was reduced as time passed and the trend was the same in both groups . the most prevalent bacteria found in our study were gram negative rods among which klebsiella took precedence over pseudomonas and enterobacter . afjeh from iran also reported that vap related organism was gram negative rods ( e. coli and then klebsiella ) more than other bacteria . frequency of bacterial growth in endotracheal tube in the two groups this was in accordance with tripathi et al and aly et al study[3 , 16 ] . this reports indirectly confirm the importance of hand - washing in neonatal intensive care units , because the source of gram - negative rods is mainly environmental , primarily from the hands of staff members . although we chose the sample size according the statistical formulas used in a similar study by aly et al , the frequency of colonization in our study was lower . so , further studies with larger sample size are necessary to determine physiologic or clinical benefits of lateral position . because endotracheal cultures were performed qualitative , we could not show the correlation of position and intensity of colonization . in our study we surveyed the effect of position on bacterial colonization but not vap . colonization of the endotracheal tubes is the main risk factors for vap , but it may occur separately with no symptoms or signs of vap . although the colonization rate in lateral position group was 2 times less than in supine position group , because the difference was not statistically significant it needs to be confirmed by a larger sample study . incidentally there is a negative attitude toward lateral positioning because the traditional position in all nicus is supine so some neonatologists were reluctant to cooperate with the intervention . Output:	objectiverecent data suggest that during mechanical ventilation , lateral patient position ( in which the endotracheal tube is horizontal ) decreases the incidence of bacterial colonization of ventilated neonates . the objective of this study was to evaluate the influence of lateral and supine position on bacterial colonization of endotracheal tube in neonates.methodswe conducted a prospective , randomized , clinical trial with 31 intubated neonates ( intubated within 48 hours after birth ) ; sixteen neonates were positioned supine ( supine group ) , and fifteen were maintained in the lateral position ( lateral group).tracheal aspirates were cultured in second and fifth days of mechanical ventilation . data were analyzed with spss version 16.findingsin the second day of ventilation , positive cultures were recognized in 6.2% of supine group and 6.7% of lateral group . after 5 days , tracheal cultures were positive in 25% ( 4 neonates ) of supine group and 13.3% ( 2 neonates ) of lateral group that was n't statistically significant ( p=0.9 in second day and p=0.9 in the fifth day ) . the most common organisms isolated from tracheal aspirates were gram - negative rods ( klebsiella).conclusionsince respiratory contamination is very common among ventilated neonates and the effect of lateral position on bacterial colonization of endotracheal tubes of intubated neonates was n't established in our study , further studies are required to suggest ways to decrease bacterial colonization of intubated neonates .
PubmedSumm2061	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: lupus nephritis ( ln ) is a major clinical manifestation of systemic lupus erythematosus ( sle ) ; it occurs in up to 50% of patients at onset of the disease and over 60% of patients during the disease . clinical course ranges from asymptomatic urinary occult blood to nephrotic syndrome or acute kidney injury since kidney injuries in ln are so variable . the patients of the who class iv ( proliferative glomerulonephropathy ) at initial renal biopsies show higher rate of end - stage renal failure ( esrf ) compared with those of the other classes . the mean 50% renal survival time of class iv is 189 months in japanese patients . to understand the pathogenesis of cytokines on ln , murine models of sle fifty percent survival is 5 - 6 months of age , and the cause of death is renal failure . in nzbxnzw mice , ln becomes apparent at 5 - 6 months of age , leading to renal failure and death at 1012 months of age . although numerous underlying mechanisms are reported , cytokine plays a key role in disease initiation and progression . this paper focuses on the contribution of cytokine , cytokine receptors , and intracellular signaling in ln . the deposit of immune complexes ( ics ) has been regarded as responsible for the initiation of ln . ic deposition activates complement cascade , leading to mesangial cell activation and proliferation . once activated , mesangial cells produce various types of cytokines and chemokines , leading to amplification of glomerular disease . in addition to ic - mediated glomerular injury , auto - antibodies may also promote proliferation and activation in kidney resident cells . demonstrated that anti - dna antibody induced the secretion of interleukin ( il- ) 1 , il-6 , and tumor necrosis factor ( tnf- ) in human cultured mesangial and tubular epithelial cell . these observations suggest that renal resident cells activated by ics and/or auto - antibodies secret the cytokines , which may further amplify inflammatory processes . they also demonstrated that anti - dna antibody induced protein kinase c activation , which is a signal pathway causing the synthesis of cytokines in human mesangial cell . several groups showed the beneficial effects of tnf in nzbxnzw mice [ 811 ] , whereas some groups reported the adverse effects in mrl - fas mice [ 1217 ] . the protective effect is specific to nzbxnzw strain , and the mechanism is not clear . as for the human ln , yokoyama et al . showed that serum levels of tnf- are correlated to glomerular icam-1 expression , which is associated with endocapillary lesions in renal biopsy specimen . summarized the reported 12 cases , who were treated with tnf blockers . in 9 out of 12 patients , tnf blocker therapy led to the improvement of ln and the long - term renal responses . matsumura et al . also reported that 6 out of 8 patients showed improved urinary protein and sle activity by the anti - tnf therapy . these data suggest that anti - tnf- therapy may have therapeutic potentials in human ln . however , some groups reported that anti - tnf- therapy in rheumatic disease induce autoantibodies formation and lead to sle including ln [ 22 , 23 ] . we should be aware that anti - tnf- therapy could induce sle as well . , several studies demonstrated that il-6 contributes to the production of anti - dna antibody from b cells [ 24 , 27 ] . reported that il-6 inhibits the function of regulatory t cells in lupus model mice . in human samples , il-6 mrna level in peripheral blood mononuclear cells is higher in patients with active ln than that in those with inactive ln . as for the clinical therapy , they reported that arthritis improved all 7 patients with arthritis at base line , but there was no change of proteinuria during the study in all 5 patients with ln at the base line . il-1 induces endothelial adhesion molecules and increases the production of igg and anti - dna antibody from b cell in mrl - fas mice . anti - dsdna antibody induces il-1 production in mesangial cells , which lead to the overexpression of extracellular matrix , hyaluronan . in human ln however , il-1 receptor antagonist therapy does not improve ln in mrl - fas mice . several protein kinase cascades mediate the intracellular cytokine signal transduction , leading to various types of cell response , such as cell migration , proliferation , and inflammatory response . p38 mitogen - activated protein kinase ( mapk ) is responsible for the production and signal transduction of cytokines . we found that pharmacologic inhibition of p38 mapk significantly reduced cytokine expression and improved the renal injury in mrl - fas mice . in addition , the inhibition of p38 mapk also reduced the number of mature dcs within injured kidney and decreased il-12 and il-23 expression on dcs ( figure 1 ) . once inflamed renal resident cells produce cytokines and chemokines , leukocytes migrate to glomerulus and interstitium . in human ln , most infiltrating mononuclear leukocytes are t lymphocytes , with lesser numbers of macrophages ( m ) , b lymphocytes , and natural killer cells . infiltrate m and dendritic cells ( dcs ) secrete a variety of cytokines and activate nave t cells , leading the cytokine profile towards th1 , th2 , and/or th17 . recent studies suggest that th17 cells play a crucial role in the pathogenesis of ln . zhang et al . demonstrated that il-17-producing cd3 cells from lupus prone mice induce nephritis when transferred to nonautoimmune , lymphocyte - deficient rag-1 mice . steinmetz et al . showed that cxcr3 , which is expressed on th1 and th17 cells , deficient lupus prone mrl - fas mice ameliorate ln accompanied by the reduction of interferon ( ifn)- and il-17 producing t cells . furthermore , il-23 receptor b6/lpr mice are protected from the development of ln , followed by the decrease of il-17-producing t cells . in human ln reported that cd4cd8 double - negative t cells produce il-17 and infiltrate the kidneys in ln patients . interestingly , double - negative t cells have been reported as a major source of il-17 in mrl - fas mice as well . il-12 mrl - fas mice are protected from ln followed by the reduced production of ifn- . moreover , ifn- receptor mrl - fas mice showed the decreased renal pathology and extended survival . in contrast , the role of type i ifn ( ifn-/ ) , which is classically thought to induce th1 type inflammation , is equivocal . the administration of ifn- accelerates the development of lupus in lupus - prone mice [ 46 , 47 ] . moreover , type i ifn receptor ( ifnar ) nzbxnzw is protected from ln . as opposed to this study , hron and peng reported that ifnar mrl - fas mice showed increased lymphadenopathy , autoantibody production , and ln . of note , schwarting et al . reported that the ifn- therapy reduces the activity of ln in mrl - fas mice . these results indicate the different role of ifn- and ifn- in ln though ifnar is the common receptor for both ifns . supporting this notion , satchell et al . reported that ifn- had an effect on barrier properties , increasing electrical resistance across monolayers of either glomerular endothelial cells or podocytes and decreasing transmonolayer passage of albumin . il-18 , which is a strong inducer of ifn- , is upregulated in mrl - fas mice . interestingly , docosahexaenoic acid in fish oil decreased serum levels of il-18 and attenuated lupus nephritis in nzbxnzw mice . in human ln , they also demonstrated that ifn-/il-4 ratio in peripheral blood cd4 cell was correlated with pathological activity index . yokoyama et al . revealed that serum level of ifn- is related to proliferative and active lesions , and the level is decreased by methylprednisolone pulse therapy . chan et al . reported the correlation between glomerular expression of the th1 transcription factor ( t - bet ) , ifn- , and il-2 with serum c3 , c4 and anti - double - strand - dna antibody level . tucci et al . demonstrated the il-18 expression within glomeruli in patients with severe ln . charles et al . revealed that autoreactive ige and il-4 are essential for lupus model mice and sle patients . they showed that activated basophils secret il-6 and il-4 , which promote th2 response and b cell activation , resulting in autoantibodies production [ 62 , 63 ] . il-27 receptor mrl - fas mice showed membranous glomerulonephritis with the predominance of th2 systemic reaction . in lupus patients with who type v ( membranous nephropathy ) , ifn-/il-4 expression ratio was lower in peripheral blood t cell , whereas ifn-/il-4 expression ratio was higher in those with who type iv ( proliferative glomerulonephropathy ) . furthermore , th2 cytokine dominance is also reported in the kidney tissue from the patients with who type v . originally , th2 cells and antigen - presenting cells have been reported as a source of il-10 . however , recent reports show that th1 cells and th17 cells in addition to th2 cells produce il-10 [ 6669 ] . ishida et al . reported that anti - il-10 therapy delayed the onset of lupus nephritis in nzbxnzw mice . interestingly , they showed that anti - il-10 therapy increased the serum levels of tnf- , which contributed to the protection from autoimmunity . also described that anti - il-10 therapy reduced proteinuria in human ds - dna ab - induced lupus model mice . continuous overexpression of low levels of il-10 delayed the production of autoantibodies and decreased the severity of ln . these differences may be related to the mice strain , disease models , and/or the amount of il-10 expression . in sle patients , anti - il-10 therapy ameliorates skin and joint lesions in all 6 patients . however , the effect of anti - il-10 therapy on ln is not clear in this study . cytokine is upregulated by the immune deposits and/or autoantibodies in disease initiation phase , leading to inflammatory cytokine / chemokine expression and leukocyte infiltration and activation . activated leukocytes produce cytokines , which amplify the inflammatory response . then , sustained cytokine production by multiple triggers is associated with progression of ln . thus , cytokine is essential from the initiation to progression phase of ln ( figure 2 ) . . however , sufficient evidence is not yet available to clarify the efficacy of anticytokine therapy for human ln . Output:	lupus nephritis ( ln ) is a major clinical manifestation of systemic lupus erythematosus ( sle ) . although numerous abnormalities of immune system have been proposed , cytokine overexpression plays an essential role in the pathogenesis of ln . in the initial phase of the disease , the immune deposits and/or autoantibodies induce cytokine production in renal resident cells , leading to further inflammatory cytokine / chemokine expression and leukocyte infiltration and activation . then , infiltrate leukocytes , such as macrophages ( m ) and dendritic cells ( dcs ) , secrete a variety of cytokines and activate nave t cells , leading the cytokine profile towards t helper ( th)1 , th2 , and/or th17 . recent studies revealed these inflammatory processes in experimental animal models as well as human ln . the cytokine targeted intervention may have the therapeutic potentials for ln . this paper focuses on the expression of cytokine and its functional role in the pathogenesis of ln .
PubmedSumm2062	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: Output:	when infant rhesus monkeys were exposed to lead via the addition of lead acetate ( 0.59 mg / kg body weight ) to their formula or by the consumption of lead particles from lead - based surrogate mothers , they developed symptoms of lead intoxication within 6 weeks . seizures , muscular tremors , and altered social interaction were the predominant changes . visual impairment was also apparent in the more severely affected animals . in the animals showing obvious symptoms lead levels varied between 300 to 500 g/100 ml of blood . even in those animals having blood lead levels below 100 g , hyperactivity and insomnia were observed . when the exposure to lead was eliminated , seizures subsided and visual impairment was reduced ; however , the abnormal social interaction persisted . these animals also experienced a gradual decline in hematocrit and hemoglobin values during the period of examination . liver and kidney biopsies obtained from these lead - exposed animals revealed characteristic intranuclear inclusions . when adolescent and adult monkeys were exposed to doses of lead acetate similar to those employed in the infant experiments , lead levels in excess of 200 g/100 ml of blood were recorded . however , there were no obvious behavioral abnormalities observed . there were , however , numerous lead inclusion bodies in kidney biopsy specimens from these animals . these data suggest that , like man , the infant nonhuman primate is much more susceptible to lead intoxication than is the adult . the clinical and behavioral changes recorded in these infant rhesus monkeys suggest their use as an experimental model to evaluate lead intoxication . imagesfigure 6 .
PubmedSumm2063	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the database of the korea university anam hospital , seoul , korea was searched for all crc patients who underwent surgical resection between 2002 and 2013 . a cohort of patients who underwent hepatic resection or pulmonary resection for crc metastases was also identified . sections were incubated for 15 minutes with antibodies against mlh1 protein ( 1:200 , es05 , leica , newcastle upon tyne , uk ) , msh2 protein ( 1:100 , g219 - 1129 , cell marque , rocklin , ca , usa ) , msh6 protein ( 1:50 , 44 , cell marque ) , and pms2 ( 1:50 , mrq-28 , cell marque ) . tumors were considered deficient in mlh1 , msh2 , msh6 , and pms2 expression when there was a complete absence of detectable nuclear staining in neoplastic cells . intact nuclear staining of the adjacent non - neoplastic epithelium , stromal cells , or lymphocytes served as an internal positive control ( appendix 1 ) . whole sections were stained and reviewed for cases that showed a loss of expression on a microarray . clinicopathological features , including age , size , gender , site of primary crc , lymph node status , lymphovascular invasion , perineural invasion , carcinoembryonic antigen ( cea ) levels at diagnosis , t category , tumor differentiation , and metastatic site , were compared between dmmr and proficient mmr ( pmmr ) patients used fisher exact , tests and mann - whitney test . this case was reviewed by the institutional review board of korea university medical center ( an15349 - 00 ) . the database of the korea university anam hospital , seoul , korea was searched for all crc patients who underwent surgical resection between 2002 and 2013 . a cohort of patients who underwent hepatic resection or pulmonary resection for crc metastases was also identified . to characterize the mmr system , ihc was performed using 4-m - thick paraffin tissue sections . sections were incubated for 15 minutes with antibodies against mlh1 protein ( 1:200 , es05 , leica , newcastle upon tyne , uk ) , msh2 protein ( 1:100 , g219 - 1129 , cell marque , rocklin , ca , usa ) , msh6 protein ( 1:50 , 44 , cell marque ) , and pms2 ( 1:50 , mrq-28 , cell marque ) . tumors were considered deficient in mlh1 , msh2 , msh6 , and pms2 expression when there was a complete absence of detectable nuclear staining in neoplastic cells . intact nuclear staining of the adjacent non - neoplastic epithelium , stromal cells , or lymphocytes served as an internal positive control ( appendix 1 ) . whole sections were stained and reviewed for cases that showed a loss of expression on a microarray . clinicopathological features , including age , size , gender , site of primary crc , lymph node status , lymphovascular invasion , perineural invasion , carcinoembryonic antigen ( cea ) levels at diagnosis , t category , tumor differentiation , and metastatic site , were compared between dmmr and proficient mmr ( pmmr ) patients used fisher exact , tests and mann - whitney test . this case was reviewed by the institutional review board of korea university medical center ( an15349 - 00 ) . the clinicopathological features of 61 patients included in the study are detailed in table 1 . the mean patient age was 58 years ( range , 31 to 78 years ) ; 26% of the patients were female and 73% were male . the mean tumor size was 5.4 cm ( range , 1.8 to 15 cm ) . the anatomical location of the primary tumor was classified as left ( rectum , rectosigmoid colon , splenic flexure , and descending colon ) or right side ( ascending colon , hepatic flexure , and cecum ) . in total , 55/61 ( 90.1% ) were t3 cancers that invade through the muscularis propria into pericolorectal tissues . neither patient had a family history of cancer . using fisher exact , tests and mann - whitney tests to analyze the relationship between mmr status and clinicopathological features , significant relationships were not detected for age ( p=.58 ) , size ( p=.14 ) , sex ( p=.34 ) , site ( p=.96 ) , t stage ( p=.36 ) , tumor differentiation ( p=.32 ) lymph node metastasis ( p=.13 ) , lymphovascular invasion ( p=.75 ) , perineural invasion ( p=.30 ) , or cea level ( p=.49 ) . of the primary tumors , 7/61 ( 11.4% ) showed a dmmr system based on ihc , while secondary tumors showed a deficiency in 13/61 cases ( 21.3% ) . in total , three cases showed deficiencies in both the primary and the paired metastatic lesions . in 10 cases , proficient expression was found in the primary lesions , but not in the corresponding metastatic lesions . in four cases , proficient expression was detected in the secondary tumor , but not in the primary tumor . in total , 14 patients died of cancer , and the median survival was 33 months from the date of initial diagnosis . a survival analysis was performed assuming that the primary and metastatic tumors showing intact expression using ihc group are pmmr and the loss of expression in either tumor indicates dmmr ( fig . this result was not statistically significant ( log - rank test , p=.22 ) , but the pmmr group showed a more favorable prognosis compared to the dmmr group . the clinicopathological features of 61 patients included in the study are detailed in table 1 . the mean patient age was 58 years ( range , 31 to 78 years ) ; 26% of the patients were female and 73% were male . the mean tumor size was 5.4 cm ( range , 1.8 to 15 cm ) . the anatomical location of the primary tumor was classified as left ( rectum , rectosigmoid colon , splenic flexure , and descending colon ) or right side ( ascending colon , hepatic flexure , and cecum ) . in total , 55/61 ( 90.1% ) were t3 cancers that invade through the muscularis propria into pericolorectal tissues . using fisher exact , tests and mann - whitney tests to analyze the relationship between mmr status and clinicopathological features , significant relationships were not detected for age ( p=.58 ) , size ( p=.14 ) , sex ( p=.34 ) , site ( p=.96 ) , t stage ( p=.36 ) , tumor differentiation ( p=.32 ) lymph node metastasis ( p=.13 ) , lymphovascular invasion ( p=.75 ) , perineural invasion ( p=.30 ) , or cea level ( p=.49 ) . 7/61 ( 11.4% ) showed a dmmr system based on ihc , while secondary tumors showed a deficiency in 13/61 cases ( 21.3% ) . in total , three cases showed deficiencies in both the primary and the paired metastatic lesions . in 10 cases , proficient expression was found in the primary lesions , but not in the corresponding metastatic lesions . in four cases , proficient expression was detected in the secondary tumor , but not in the primary tumor . in total , 14 patients died of cancer , and the median survival was 33 months from the date of initial diagnosis . a survival analysis was performed assuming that the primary and metastatic tumors showing intact expression using ihc group are pmmr and the loss of expression in either tumor indicates dmmr ( fig . this result was not statistically significant ( log - rank test , p=.22 ) , but the pmmr group showed a more favorable prognosis compared to the dmmr group . in this study , mmr proteins were evaluated by ihc for the following reasons : ( 1 ) ihc is less time consuming than pcr , ( 2 ) is easier to implement , and ( 3 ) enables the simultaneous identification of affected mmr genes . first , it is well known that the ihc staining patterns for msh6 ihc are variable . for instance , tissue hypoxia and delayed fixation reduce the sensitivity of detection of mmr gene expression or suppress mmr . third , some mlh1 mutation - positive cases or even cases with mlh1 promoter methylation show false - positive nuclear staining . a missense mutation or an in - frame insertion / deletion mutation in mlh1 does not affect mlh1-pms2 interactions , and thus the protein reacts with the antibody used for ihc . lastly , interpretation of mmr ihc results requires consideration that the mmr proteins act as heterodimers . for example , crcs that show loss of mlh1 and pms2 expression , but intact msh2 and msh6 expression indicate an mlh1 deficiency . the loss of pms2 expression results from mlh1 mutations . in this study , our major interest was investigating the concordance in mmr deficiencies between the primary crcs and its corresponding metastases . reported matching ihc results between the primary tumor and metastatic tissue for all cases examined . however , agoston et al . found discordance in the mmr status between primary tumors and metastases in 20.2% of cases . haddad et al . found a very low frequency of msi using pcr in resected crc hepatic metastases in 190 patients . unlike in primary crcs , the rate of msi in resectable crc hepatic metastases is approximately 2.5% . first , high - frequency msi primary crcs do not frequently metastasize to the liver . second , high - frequency msi primary crcs spread to the liver extensively and therefore are considered unresectable . however , the incidence was somewhat different between the primary ( 11.5% ) and the metastatic ( 21.3% ) cancer groups . in this study , among the 36 hepatic metastatic cancers , five cases showed dmmr in primary crcs . this frequency of approximately 14% was not as low as that reported in the previous study . the clinicopathological features and each mmr protein expressions of the cases with either primary or metastatic dmmr are summarized in tables 4 and 5 . except one case , based on ihc , primary and metastatic lesions of all the cases showed intact mlh1 expression . in 10 cases , proficient expression was found in the primary tumor , but not in the corresponding metastatic lesions . in four cases , proficient expression was detected in the secondary tumor , but not in the primary tumor . furthermore , the precise mmr proteins that showed a loss of expression were not exactly matched in the primary and the metastatic tumors . to further examine the data , we divided the dmmr cases into five groups . during heterodimeric complex formation , the loss of pms2 is followed by the loss of mlh1 expression owing to functional dimerization . similarly , the loss of msh2 is accompanied by the loss of msh6 ( fig . 2 ) . however , the loss of isolated msh6 or pms2 is not accompanied by mlh1 or msh2 , reflecting mutations in msh6 or pms2 . although three cases were classified as a combined group that do not belong to the above four groups , most of the dmmr cases fit the classification , indicating that the ihc expression is consistent . mlh1 mutations are the most common type among the mmr genes . however , the detected loss of the mlh1 protein was significantly less frequent than the loss of the msh2 protein in this study . as stated above , we attributed this difference to the ihc technique . most mutations in msh2 result in truncated proteins , which consequently showed a loss of expression in ihc analyses . mlh1 mutations are non - functional missense mutations , and mutated - mlh1 cases also show proficient expression . a meta - analysis showed that using ihc , only 74% of mlh1 losses were detected in the mlh1 mutation - positive cases , compared to the 91% detected using msi testing . however , msh2 mutations could be detected in up to 94% of the msh2 mutation - positive cases by ihc . we infer that there may be false - positive mlh1 cases , and additional pcr studies are needed to identify the precise mlh1 mutations . in our study , cases were considered deficient if any target locus ( e.g. , mlh1 , msh2 , msh6 , or pms2 ) was not expressed . this is a limitation of our study , and variable patterns could lead to erroneous interpretations . in one case , we detected the losses of mlh1 , msh6 , and pms2 expression , suggesting a similar scenario . we attributed this discordance in ihc results to the variable reactivity and subjective data interpretations . all cases received adjuvant chemotherapy after resection of the primary tumor , because all cases included in this study were stage iv . most of the patients ( 56 cases ) were treated with folfox regimens ( combination of folic acid , 5-fluorouracil [ 5-fu ] and oxaliplatin ) . three cases were treated only with 5-fu based chemotherapy . among the 61 cases , 50 cases received chemotherapy before metastatectomy , where s 11 cases received chemotherapy after metastatectomy . of the 50 cases that received chemotherapy before metastatectomy , 11 cases ( 22% ) showed discordance in ihc . in seven cases , proficient expression was found in the primary lesions , but not in the corresponding metastatic lesions . we assume that these losses of expression could be associated with chemotherapy - related effect . since all cases were stage iv tumors , the low survival rates were expected . despite these limitations , we detected a correlation between the mmr status of primary and metastatic crcs using only ihc . further research should focus on the modifications in the biological and biochemical properties of dna mmr proteins during metastatic processes . Output:	backgroundcolorectal cancer ( crc ) is one of the most common malignancies worldwide . approximately 10%15% of the crc cases have defective dna mismatch repair ( mmr ) genes . although the high level of microsatellite instability status is a predictor of favorable outcome in primary crc , little is known about its frequency and importance in secondary crc . immunohistochemical staining ( ihc ) for mmr proteins ( e.g. , mlh1 , msh2 , msh6 , and pms2 ) has emerged as a useful technique to complement polymerase chain reaction ( pcr ) analyses . methodsin this study , comparison between the mmr system of primary crcs and paired liver and lung metastatic lesions was done using ihc and the correlation with clinical outcomes was also examined . resultsbased on ihc , 7/61 primary tumors ( 11.4% ) showed deficient mmr systems , while 13/61 secondary tumors ( 21.3% ) showed deficiencies . in total , 44 cases showed proficient expression in both the primary and metastatic lesions . three cases showed deficiencies in both the primary and paired metastatic lesions . in 10 cases , proficient expression was found only in the primary lesions , and not in the corresponding metastatic lesions . in four cases , proficient expression was detected in the secondary tumor , but not in the primary tumor . conclusionsalthough each ihc result and the likely defective genes were not exactly matched between the primary and the metastatic tumors , identical results for primary and metastatic lesions were obtained in 77% of the cases ( 47/61 ) . these data are in agreement with the previous microsatellite detection studies that used pcr and ihc .
PubmedSumm2064	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: refractive accommodative esotropia ( rae ) is an esotropia that is restored to orthotropia at all fixation distances and all gaze positions by optical correction of the underlying hypermetropic refractive error . retinal image blur induces accommodation to focus the image ; then , this accommodative effort elicits accommodative convergence when fixating at distance or near . when fusional divergence is insufficient to compensate for the excessive convergence innervation , esotropia develops . vergence movements including divergence require the cooperation of the cerebral cortex , in particular , a state of visual attention called the psychooptical reflex . the child behavior checklist ( cbcl ) is widely used in the field of psychology and consists of 113 questions used to rate the behavioral and emotional problems of children . there are two dimensions : internalizing symptoms ( withdrawn , somatic complaints , and depression / anxiety ) and externalizing symptoms ( social problems , thought problems , attention problems , delinquent behavior , and aggressive behavior ) . therefore , i evaluated the relationships between rae and psychological symptoms as measured by the cbcl . i prospectively recruited patients aged 36 years with more than 3.0 diopters of hyperopia who presented at kim 's eye hospital from january 2011 to march 2013 . this research study was reviewed and approved by the institutional review board ( irb ) of kim 's eye hospital , and all procedures conformed to the guidelines of the declaration of helsinki . all patients underwent a full ophthalmologic examination including slit - lamp biomicroscopy , fundoscopy , alternate prism cover testing , and cycloplegic refraction using 1% cyclopentolate . refractive accommodative esotropia was defined as an esotropia that was corrected to > 10 prism diopters ( pd ) of orthotropia at both far and near distance with use of the full cycloplegic hyperopic correction . the control group had hyperopia of > 3 diopters ( d ) and orthotropia at near and far distance . subjects were excluded if they had worn glasses previously or had a history of ophthalmic surgery . further , children with anisometropia > 1.0 d and astigmatism > 1.5 d were excluded . the angle of esodeviation was measured by alternate cover test , and the mean deviation of two consecutive measurements was used . the k - cbcl was completed by parents at the first visit before glasses were prescribed . the child behavior checklist ( cbcl ) is a parent - report questionnaire in which the child is rated on various behavioral and emotional behaviors . the reliability and validity of the korean version of the cbcl ( k - cbcl ) are well established in korean children and adolescents . the checklist assesses internalizing ( i.e. , anxious , depressive , and overcontrolled ) and externalizing ( i.e. , aggressive , hyperactive , noncompliant , and undercontrolled ) behaviors . several subareas are measured , including internalizing factors ( social withdrawal , somatic complaints , anxiety , and depression ) and externalizing factors ( social problems , thought problems , attention problems , delinquent behavior , and aggressive behavior ) . the k - cbcl score was computed based on korean normative samples , with the total problem behavior score computed by summing the scores obtained for each item . raw scores for each clinical factor were transformed into t - scores based on published norms . i investigated the relationship between the k - cbcl score and the amount of esodeviation in rae children . independent student 's t - test was performed to investigate the differences in k - cbcl indexes between groups . the correlation between k - cbcl score and the angle of esodeviation was measured using the pearson correlation method . a total of 20 rae children and 34 hyperopic children were included in the study . the mean ages of rae children and controls were 5.0 1.7 and 5.4 0.8 , respectively ( p = 0.233 ) . mean refractive errors were slightly higher in the control group ( rae children , 4.6 d ; control group , 5.3 d ) ; however , there was no significant difference between groups ( table 1 ) . two out of three internalizing indexes were significantly different between groups ( somatic complaints : rae children 50.1 4.6 and controls 46.6 5.8 , p = 0.026 ; depression / anxiety : rae children 48.8 7.9 and controls 43.9 6.8 , p = 0.024 ) . although there was no significant difference , rae children scored slightly higher on the externalizing behavior index . regarding the relationship between the angle of esodeviation and the k - cbcl score , although the frequency of withdrawn behavior did not differ between groups , the far angle of esodeviation showed a moderate correlation with withdrawn behaviors ( table 2 ) . among the rae children who scored higher on indices related to social and attention problem , the near angle of esodeviation was larger than the far angle of esodeviation ( r = 0.725 and r = 0.650 ; p = 0.018 and p = 0.042 , resp . ) . the present study revealed that psychosocial problems might be related to the pathogenesis of refractive accommodative esotropia . hypermetropic children with several characteristics including a positive family history , low binocular sensory function , low hypermetropia ( < + 3.0 d ) , and significant anisometropia had a high risk of developing refractive accommodative esotropia . unfortunately , i did not investigate sensory function and family history in the present study . mean refractive error was slightly reduced in the rae group ( rae children 4.6 d and control group 5.3 d ) , though this trend was not significant . the present study showed that rae children scored higher than controls on the indexes for somatic complaints and depression / anxiety . somatic complaints include fatigue , aches , nausea , vomiting , headaches , dizziness , and skin , stomach , or eye problems . children with significant medical disorders tend to score high on the internalizing subscale and particularly on the somatic complaints subscale of the cbcl . somatic complaints are a predominant characteristic of anxiety disorders and are believed to play an important role in anxious states [ 7 , 8 ] . furthermore , children with anxiety disorders exhibit more somatic complaints compared to normal control children . control subjects tended to score higher on other externalizing indexes , though these trends were not significant . internalizing factors such as withdrawn behavior , somatic symptoms , and depression / anxiety refer to a broad class of behaviors in which children direct feelings and emotions inward ; these are commonly in contrast to externalizing factors . as shown in figure 1 , although the associations with externalizing factors had no statistical significance , the rae group scored higher on internalizing indexes and lower on externalizing indexes than the control group . the angle of deviation in rae is usually variable . to minimize deviation variability , i used the mean of two consecutive deviation measurements for all analyses . the angle of esodeviation correlated with withdrawn behaviors on the cbcl , but the near angle was not associated with the score on any subsection of the cbcl . withdrawn behavior is defined as the behavioral tendency to isolate oneself from one 's peers ; nine items on the cbcl indicate withdrawn behavior such as a preference for being alone , sadness , unwillingness to talk , shy demeanor , and withdrawn or underactive behavior . withdrawn behavior was negatively correlated with the angle of deviation . in other words , rae children with lower scores on indexes of withdrawn behavior , who are outgoing and active , are more likely to have larger angles of esodeviation . as mentioned earlier , however , there was no significant correlation between either of these measures and the angle of esodeviation . therefore , somatic complaints and depression / anxiety might be related to the pathogenesis of rae . as i mentioned earlier , i did not evaluate related factors such as binocular sensory function or family history . however , these biases can be excluded because the study was designed as a prospective study . in conclusion , hypermetropic children with high scores on the somatic complaint and depression / anxiety subscales of the cbcl could be at high risk for developing rae . Output:	purpose . to investigate the psychophysiologic aspects of refractive accommodative esotropia ( rae ) . methods . i prospectively recruited patients aged 36 years with more than 3.0 diopters of hyperopia who presented at kim 's eye hospital from january 2011 to march 2013 . i compared korean child behavior checklist ( k - cbcl ) which consists of internalizing factors ( social withdrawal , somatic complaints , anxiety , and depression ) and externalizing factors ( social problems , thought problems , attention problems , delinquent behavior , and aggressive behavior ) between rae group and control group . results . two out of three internalizing indexes were significantly different between groups ( somatic complaints : rae children 50.1 4.6 and controls 46.6 5.8 , p = 0.026 ; depression / anxiety : rae children 48.8 7.9 and controls 43.9 6.8 , p = 0.024 ) . although there was no significant difference , rae children scored slightly higher on the externalizing behavior index . in the rae group , the far angle of esodeviation showed a moderate correlation with withdrawn behaviors . conclusion . hypermetropic children with high scores on the somatic complaint and depression / anxiety subscales of the cbcl could be at high risk for developing rae . psychosocial problems might be related to the pathogenesis of refractive accommodative esotropia .
PubmedSumm2065	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the management of unowned domestic cat ( felis catus ) populations is a global problem that raises issues for individual cat welfare and conservation of native wildlife . the remarkable reproductive success of this species is a major factor in the ability of cats to colonise diverse environments . the onset of puberty in females can be as early as 12 weeks and a single female cat can produce 40 kittens per year . it is estimated that between 75% and 97% of cats in a population need to be desexed to stabilise numbers . early - age desexing removes the responsibility for desexing from owners who , despite financial incentives , frequently fail to return the cat for surgery , while still allowing rehoming at a young age to optimise socialisation . there is increasing evidence that early - age desexing is not only safe in the short and long term , but also offers advantages , including reduced surgical time and rapid recovery , over the traditional age of 6 months or older . many national and international shelter , welfare and veterinary organisations now endorse early - age desexing , including the royal society of the prevention of cruelty to animals , the international society of feline medicine and the american veterinary medical association . despite these factors , recent surveys indicate that attitudes within the veterinary profession towards early - age desexing are variable . the choice of surgical approach to ovariohysterectomy ( ohe ) shows regional variation , with a flank approach favoured in the uk ( 96% ) , and a midline approach in the us . benefits described for midline ohe include better visualisation and reduced postoperative pain scores , whereas a flank approach may offer shorter surgery time and reduced risk of hypothermia , herniation or evisceration . evidence to support one approach over the other is limited , particularly in prepubertal cats . the aim of this study was to determine whether there was an association between age and surgical approach , individually or combined , and perioperative wound complication in cats undergoing ohe . cases were recruited from the cat protection society ( cps ) of new south wales desexing program . surgeries were performed at four veterinary clinics ( a d ) . a single surgical approach , either flank ( clinic a ) or midline ( clinics b d ) was used at each location . only healthy cats with a body weight of 1 kg or more were accepted in this program . cats were examined twice daily and findings , including wound assessment , were recorded in a manual database . the database was searched retrospectively to identify cats that underwent ohe between 1 june 2010 and 31 december 2012 inclusive . cases were included if the following information was available : age at the time of surgery ; clinic where surgery was performed ; and complete records from at least 1 day prior to surgery to 5 days postsurgery inclusive . cases were excluded if surgery was performed by a veterinary student under the supervision of a veterinarian . a wound complication was recorded when the description of the surgical site included any of the following terms : redness ; swelling ; heat ; pain ; exudation ( grouped as inflammation ) ; or wound dehiscence in the 5 day postoperative period . data were exported into a csv file and analysed using statulator , an online statistical program ( http://statulator.com/ ) . initially , descriptive analyses were conducted , including the creation of frequency tables for categorical variables and the calculation of summary statistics for age to understand the distribution of variables . to evaluate the association of age with wound complication , cats were classified into groups according to age : those aged 12 weeks and those aged > 12 weeks . contingency tables of age and surgical approach with wound complication ( yes / no ) were created and analyses were conducted to evaluate crude associations between them . stratified analyses were then conducted to evaluate association of surgical approach with wound complication after adjusting for age . , 194 were excluded because the clinic was not identified ( n = 145 ) , recording of wound status was incomplete ( n = 45 ) or surgery was performed by a student under supervision ( n = 4 ) . in total , 312 cases met the inclusion criteria ( table 1 ) . the mean age at the time of surgery was 41 weeks ( median 12 weeks , range 5468 weeks ) . wound complication was identified in 19 cases ( 6.09% ) . in 18 of these , signs were consistent with inflammation at the surgical site , while wound dehiscence was identified in a single 12-week - old patient that had an ohe via a midline approach . age at desexing was not a risk factor for wound complication ; there was no significant difference in the development of wound complications in cats up to 12 weeks of age compared with older cats ( p = 0.38 ) ( table 2 ) . considering all cases , regardless of age , a midline approach was associated with 2.95-fold increased risk of wound complication compared with a flank approach ( p = 0.011 ) . when a stratified analysis by age was performed ( table 3 ) , a midline approach was 4.59-times more likely than a flank approach to result in wound complication in cats up to 12 weeks of age ( p = 0.015 ) . however , in cases over 12 weeks of age , there was no association between surgical approach and the risk of wound complication ( p = 0.22 ) . frequency tables of categorical variables and odds ratios based on crude association of each explanatory variable with development of wound complication ci = confidence interval stratified analysis of association between surgical approach and wound complications , after adjusting for age adjusted risk ratio and 95% confidence interval ( ci ) 2.75 ( 1.186.43 ) ; cochran mantel haenszel statistic 5.80 ; p value 0.016 we used a cut - off of 12 weeks to ensure that cats were prepubertal ; of 33 cats < 1 year of age that were pregnant at desexing in a uk study , two were < 4 months old . others have demonstrated that early - age desexing from 6 weeks of age carries no increased risks to the patient . there are several factors that require particular attention when performing ohe in paediatric patients compared with older cats . these include avoiding hypoglycaemia by fasting for a much shorter period ( 34 h ) and offering food immediately on recovery , as well as preventing hypothermia . a comprehensive review of early - age desexing , including suitable anaesthetic protocols , has recently been published . the overall incidence of perioperative wound inflammation ( 6% ) or dehiscence ( < 0.003% ) in this study was low , as expected for a routine procedure performed in healthy animals . our findings support previous studies that demonstrate no increase in the short - term complication rate following the desexing of young cats compared with older cats . in fact , higher overall complication rates are reported in cats > 6 months compared with those aged < 12 weeks . previous prospective , randomised studies comparing flank vs midline approaches to ohe have produced conflicting results and none have investigated prepubertal patients specifically . some of the suggested pros and cons of a flank vs a midline approach to ohe may become redundant when they are applied to prepubertal patients . for example , reduced abdominal and bursal fat allows enhanced visualisation and more accurate vessel haemostasis during ohe by either approach in kittens . the observation here that a midline approach increased the risk of wound complication in cats up to 12 weeks , but not in older cats , is interesting . the first , investigating ovariectomy in 38 cats of any age , found that a midline approach carried a higher risk of wound swelling , while a flank approach was more painful on palpation . a second study of 66 cats aged 6 months to 10 years found a greater risk of wound tenderness and discharge with a flank compared with a midline approach . surgeries were performed by veterinary students and the overall wound complication rate , as reported by owners , was high ( 42% ) with severe complications noted only after midline surgery . in a separate study , burrow et al compared flank and midline approaches to ohe in 20 cats ( median age 10 months ) performed by a single experienced surgeon and found that the flank approach was marginally quicker but was associated with a tendency to be more painful than a midline approach . it is important to note that kittens are less likely than adult cats to demonstrate visual indicators of pain , so early - age desexing protocols should include interactive pain assessment such as wound palpation . a flank approach was used by a single clinic so we can not exclude that factors other than approach contributed to our observations . however , the overall wound complication rate was low and ohe is a standard procedure so any effect from other factors is suggested to be low . these findings add to the evidence base supporting a cultural change to embrace the practice of early - age desexing as standard for feline patients . Output:	objectivesthe aim of the study was to investigate the effects of age and/or surgical approach on perioperative wound complication following ovariohysterectomy ( ohe).methodsa retrospective search of perioperative monitoring records from a shelter desexing program was conducted to identify cats that underwent ohe between 1 june 2010 and 31 december 2012 inclusive . a wound complication was defined as gross observation of inflammation or wound dehiscence at the surgical site in the 5 day postoperative period . cases were grouped according to age ( 12 weeks or > 12 weeks ) and surgical approach ( flank or midline ) . stratified analyses were conducted to evaluate the association between surgical approach and wound complications , after adjusting for age . mantel haenszel adjusted risk ratio , cochran mantel haenszel test statistic and their 95% confidence intervals were presented.resultsa total of 312 cases met the study criteria . the overall wound complication risk was low ( 6.09% ) and was not related to age . a midline approach was associated with a 4.59-times increased risk of wound complication , compared with a flank approach in cats up to 12 weeks of age ( p = 0.015 ) but not in older cats.conclusions and relevancethese findings support the practice of prepubertal desexing for cats .
PubmedSumm2066	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: osteoporosis and hip and vertebral osteoporotic fractures among an ageing population are a worldwide health problem that lead to a decreased quality of life ( 1 ) . body weight and body mass index ( bmi ) are considered potentially modifiable determinants of bone mass ( 2 , 3 ) . several studies have demonstrated an association of low body weight and bmi with low bmd and fractures ( 4 ) , while several other studies showed a protective effect of higher bmi on bmd ( 5 - 8 ) . among most frequently cited mechanisms to explain this positive association of weight with bmd is weight - related loading on bones ( 9 ) , though higher estrogen and insulin synthesis and higher plasma leptin levels are considered to play an important role ( 10 ) . on the other hand , it should be acknowledged that there are studies that did not demonstrate a significant positive association between weight and bmi and bmd or demonstrated a negative association of obesity and bmd ( 11 - 13 ) ; interaction between fat mass and bones through pro - inflammatory cytokines or low levels of adiponectins are some of the proposed mechanisms ( 14 - 16 ) . the aim of this study was to explore the relationship between body weight and body mass index ( bmi ) with total hip and lumbar spine bone mineral density ( bmd ) in a population of 100 women and 32 men from kosovo . sixty postmenopausal and forty premenopausal women , and thirty two men were prospectively enrolled in this cross - sectional study . the study protocol was approved by the ethics committee , and written informed consent was obtained from all study participants before enrollment . baseline anthropometric data ( height and weight ) were measured in light indoor clothing without shoes before performing dxa , and body mass index was calculated as kg / m . the study population was classified in three categories on the basis of world health organization ( who ) body mass index ( bmi ) criteria : normal weight ( n=28 , bmi < 24.9 kg / m ) , overweight ( n=41 , 25.0 - 29.9 kg / m ) and obese ( n=63 , bmi>30 kg / m ) . quantitative assessment of bone mineral density ( bmd ) at the lumbar spine and total hip was performed using dual - energy x - ray absorptiometry ( dxa ; stratos 800 ) technique by trained examiners . the diagnosis of osteoporosis was done according to who t - score criteria ( osteoporosis was considered if t - score -2.5 sd , low bone mass if t - score between -2.5 sd and -1 sd , and normal bmd if t - score > -1sd ) . continuous variables are expressed as means standard deviations ( median ) , while categorical data are presented as percentages . as the data were not normally distributed , the kruskal - wallis test was used to compare characteristics of the groups , while chi square test was used for categorical variables age - adjusted multiple linear regression analysis of independent associates ( weight and bmi ) of total lumbar spine and total hip bmd was calculated . continuous variables are expressed as means standard deviations ( median ) , while categorical data are presented as percentages . as the data were not normally distributed , the kruskal - wallis test was used to compare characteristics of the groups , while chi square test was used for categorical variables age - adjusted multiple linear regression analysis of independent associates ( weight and bmi ) of total lumbar spine and total hip bmd was calculated . the study population consisted of 60 menopausal women with a mean age of 63.689.27 ( sd ) years , 40 premenopausal women with a mean age of 43.447.42 ( sd ) years and 32 men with mean age of 66.3414.97 ( sd ) years . distribution of the total hip and total lumbar spine bmd according to bmi groups in menopausal and premenopausal women and men is presented in figure 1 . the descriptive characteristics of the study population groups are shown in table 1 , which includes age , bmi , height , weight , lumbar spine and total bmd , as well as information regarding bmi groups and t - score groups . there were no significant differences between study groups regarding body weight ( p=0.108 ) , while bmi score was significantly higher among menopausal women compared to premenopausal women or men ( p=0.001 ) . there were no statistically significant differences in lumbar spine or total hip bmd values between three study groups . there were significantly more obese and osteoporotic subjects among menopausal women compared to premenopausal women or men ( p<0.001 and p=0.002 , respectively ) . distribution of total hip and lumbar spine bmd according to bmi groups descriptive characteristics of the study subjects . parameters are given as mean standard deviation , median ( in the brackets ) , and number of cases ( percentage ) as appropriate ; a p - values of kruskal - wallis test or chi - square test as appropriate age - adjusted multiple linear regression analysis of independent associates of lumbar spine and total hip bmd . bmd : bone mineral density , bmi : body mass index , : unstandardized regression coefficient a statistically significant difference was observed in menopausal women for both total hip and lumbar spine bmd values among bmi groups ( p=0.010 and p=0.003 , respectively ) . total hip and lumbar spine bmd values in obese women at menopause were significantly higher compared to normal weight ( p=0.008 and p=0.035 , respectively ) . lumbar spine bmd values in obese women at menopause were significantly higher compared to overweight women at menopause ( p=0.002 ) , while no statistically significant difference was observed in total hip bmd values between obese and overweight subjects ( p=0.061 ) . no significant difference was observed between overweight and normal weight subjects in total hip and lumbar spine bmd values ( p=0.091 and p=0.783 , respectively ) . age - adjusted multiple linear regression analysis showed that both weight and bmi were significant independent associates of lumbar spine and total hip bmd , with highest adjusted squared values in total hip bmd , where weight and bmi explained 37.1% and 37.2% of the variation , respectively . a statistically significant difference was observed in premenopausal women for total hip bmd values among bmi groups ( p=0.023 ) . total hip bmd values in obese premenopausal women were significantly higher compared to normal weight and overweight subjects ( p=0.007 and p=0.043 , respectively ) , while no statistically significant difference was observed in total hip bmd values between overweight and normal weight subjects ( p=0.458 ) . no statistically significant difference was found for lumbar spine bmd values among bmi groups ( p=0.477 ) . according to age - adjusted multiple linear regression analysis no significant association of weight and bmi as independent variables with total hip and lumbar spine bmd a statistically significant difference was observed in men for total hip bmd values among bmi groups ( p=0.007 ) . total hip bmd values in obese male subjects were significantly higher compared to normal weight and overweight subjects ( p=0.03 and p=0.002 , respectively ) , while no statistically significant difference was observed in total hip bmd values between overweight and normal weight subjects ( p=0.938 ) . no statistically significant difference was found for lumbar spine bmd values among bmi groups ( p=0.060 ) . nevertheless , age adjusted multiple linear regression analysis showed a significant positive effect of the bmi on lumbar spine bmd in men ( p=0.040 ) , where it explains 12.0% of the variation . though there was a positive tendency , no significant effect of weight was observed on lumbar spine bmd in men ( p=0.058 ) , while both weight and bmi showed a significant positive effect on total hip bmd in men ( p=0.042 and p=0.028 , respectively ) according to multiple linear regression analysis . in this cross - over study , we have demonstrated that total hip bmd levels of obese menopausal and premenopausal women and men were significantly higher compared to overweight or normal weight subjects , while lumbar spine bmd levels of only menopausal women and men were higher among obese subjects . we also found evidence to suggest that bmi is a significant independent associate of lumbar spine and total hip bmd in both menopausal women and men . every unit increase in bmi among menopausal women was associated with an increase of 0.1 g / cm2 in l1-l4 bmd , and 0.01 g / cm2 in total hip bmd , while among men every unit increase in bmi was associated with an increase of 0.012 g / cm2 in l1-l4 bmd , and 0.018 g / cm2 in total hip bmd . our results are in agreement with some of the studies that demonstrated positive association of bmi with bmd . in this regard , lloyd et al ( 5 ) examined variation in bmd across bmi spectrum using data from the national health and nutrition examination survey ( nhanes ) and found positive association of bmi with bmd in the respective sample of elderly subjects , without gender differences . similarly , in a large retrospective cohort study in women aged 40 to 59 , morin et al ( 3 ) concluded that low weight and bmi predict osteoporosis and are associated with increased fracture risk . our results are fully consistent with the results of doan et al ( 6 ) that conducted a similar study among 900 geriatric subjects and demonstrated a statistically significant difference for the lumbar spine and femur neck bmd levels among the bmi groups as well as a significant relationship between both the lumbar and the femoral bmd measurements and the bmi in women , and similarly found no significant differences in lumbar spine levels among bmi groups in men . another recently conducted study of hoxha et al ( 7 ) in 64 males from kosovo compared bmi groups for l1-l4 bmd , femur neck bmd and total hip bmd levels , and reported no differences among them ; nevertheless , they demonstrated a positive correlation between bmi and femur neck and total hip bmd as well as positive association of bmi with all measurement sites bmd . due to differences in the study populations , our results are partly consistent with them , as far as we found significant differences in total hip bmd values among bmi groups in men while we failed to demonstrate a significant association of bmi with lumbar spine bmd . on the other hand , it should be acknowledged that there are studies that did not demonstrate a significant positive association between weight and bmi and bmd , or to the contrary demonstrated a negative association of obesity and bmd , thus seriously challenging the widespread belief that obesity is protective . paniagua et al ( 11 ) found a high prevalence of osteopenia and osteoporosis among male nursing home residents who were more likely to be classified as overweight and obese according to bmi . similarly , greco et al ( 12 ) demonstrated that obesity was associated with a osteoporosis and provided clinical evidence that obesity should not be considered as a protective factor . in conclusion , total hip bmd values in obese menopausal and premenopausal women as well as men were significantly higher compared to overweight or normal weight subjects . bmi was shown to be a significant independent associate of lumbar spine and total hip bmd in both menopausal women and men , while such a relationship among premenopausal women did not exist . nevertheless , significantly more obese and osteoporotic subjects among menopausal women represent a population at risk for fractures because of poor balance and frequent falls ; therefore , both obesity and osteoporosis prevention efforts should begin early on in life . Output:	background and objective : body weight and body mass index ( bmi ) are considered potentially modifiable determinants of bone mass . therefore , the aim of this study was to explore the association between body weight and body mass index ( bmi ) with total hip and lumbar spine bone mineral density ( bmd).methods : this cross - sectional study included a population of 100 women and 32 men from kosovo into three bmi groups . all the study subjects underwent dual - energy x - ray absorptiometry ( dxa ) measurements.results:total hip bmd levels of obese menopausal and premenopausal women and men were significantly higher compared to overweight or normal weight subjects , while lumbar spine bmd levels of only menopausal women and men were higher among obese subjects . age - adjusted linear regression analysis showed that bmi is a significant independent associate of lumbar spine and total hip bmd in menopausal women and men.conclusion:despite positive association between bmi and lumbar spine and total hip bmd in menopausal women , presence of more obese and osteoporotic subjects among menopausal women represent a population at risk for fractures because of poor balance and frequent falls ; therefore , both obesity and osteoporosis prevention efforts should begin early on in life .
PubmedSumm2067	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: principal neurons of the ca1 region receive input from the entorhinal cortex ( ec ) via two pathways : ( i ) the direct temperoammonic ( ta ) pathway , which consists of afferents from layer iii of ec and forms synapses with ca1 neurons in the stratum lacunosum moleculare ; and ( ii ) the indirect pathway , which originates from layer ii of ec and forms part of the trisynaptic pathway that terminates in schaffer collateral ( sc ) synapses on ca1 neurons in the stratum radiatum . nmdars are heteromeric assemblies of two essential glun1 subunits and two glun2 or glun3 subunits . there are four glun2 subunits ( a d ) of which the glun2a and glun2b predominate in the forebrain . nmdars are generally thought to be minimally active during basal synaptic transmission , but they have a key role to play during the induction of synaptic plasticity . although classically regarded as relatively stationary at the synapse , nmda receptors undergo lateral diffusion and various forms of plasticity [ 5 - 8 ] . furthermore , the subunit composition of nmda receptors can vary with development [ 9 , 10 ] and following different forms of synaptic activity [ 11 , 12 ] . recently we have shown that ltd of nmda receptor - mediated synaptic transmission can be induced by theta frequency stimulation at sc but not ta synapses and involves activation of group i mglurs ( termed mglur - ltd ; ) . the reason for the lack of activity - dependent mglur - ltd at ta - ca1 synapses is not clear but could be due to the afferent stimulation patterns in ta - ca1 not being ideal to activate group i mglurs . therefore to overcome this issue we have now used pharmacological activation of group i mglurs to investigate whether ltd can be induced at ta - ca1 hippocampal synapses and furthermore to probe the role of glun2b subunits in dhpg - ltdnmda . here we demonstrate that pharmacological activation of group i mglurs induces synaptic plasticity of nmda receptors at the sc pathway but fails to induce ltd in the ta - ca1 pathway . in addition , the nr2b - selective antagonist ro25 - 6981 was used to show that the subunit composition of nmdars remains unchanged following group i mglur - induced ltd . p14 male wistar rats were killed by cervical dislocation in accordance with united kingdom animal ( scientific procedures ) legislation . brains were removed and placed in ice - cold acsf consisting of the following ( in mm ) : nacl 124 , kcl 3 , nahco3 26 , nah2po4 1.25 , cacl2 2 , mgso4 1,d - glucose 10 ( bubbled with 95% o2/5% co2 ) . standard techniques were used to record excitatory postsynaptic potentials ( epsc ) in response to stimulation ( 100 s , 310 v ) of the sc or ta pathways . in those experiments in which both sc - ca1 and ta - ca1 inputs were studied , alternate stimulation of both pathways was conducted in each slice . electrodes ( 4 7 m ) were filled with ( in mm ) : csmeso4 , 130 , nacl 8 , mg - atp 4 , na - gtp 0.3 , egta 0.5 , hepes 10 , qx-314 5 , ph adjusted to 7.27.3 using csoh and osmolarity to 275290 mosm with sucrose . picrotoxin ( 50 m ) and nbqx ( 5 m ) were applied to isolate epscnmda . ( rs)-dhpg ( 100 m ; tocris bioscience ) was used to activate group i mglurs and ro25 - 6981 (; 5 m ; tocris bioscience ) as an antagonist of glunb receptors . changes in synaptic strength were expressed relative to the normalized baseline ( mean sem ) and significance tested using student s t - tests 30 min after ltd induction or 50 min following application of ro25 - 6981 . the group ii mglur agonist dcg - iv was applied at the end of experiments to ensure selective stimulation of ta and sc inputs . after a baseline period of 10 minutes , the group i mglur agonist ( rs)-dhpg ( 100 m ; 10 minutes ) was applied . an initial depression of nmda receptor - mediated epscs ( epscnmda ) was observed in both pathways ( at the end of dhpg application responses were : sc 53.1 3.3% ; ta 33.5 4.4% below baseline ; p < 0.05 compared to baseline ; n = 18 ; fig . thirty minutes following washout of dhpg , ltd of epscnmda ( ltdnmda ) was observed at the sc pathway ( 34.9 2.7% depression ; p=0.0012 ; n=18 ) , which was consistent with published findings [ 14 , 18 , 19 ] . however , epscnmda evoked by ta pathway stimulation recovered to baseline levels following dhpg washout ( 15.2 4.2% below baseline ; p = 0.764 ; n = 18 ) . after a baseline period of 10 minutes , the group i mglur agonist ( rs)-dhpg ( 100 m ; 10 minutes ) was applied . an initial depression of nmda receptor - mediated epscs ( epscnmda ) was observed in both pathways ( at the end of dhpg application responses were : sc 53.1 3.3% ; ta 33.5 4.4% below baseline ; p < 0.05 compared to baseline ; n = 18 ; fig . thirty minutes following washout of dhpg , ltd of epscnmda ( ltdnmda ) was observed at the sc pathway ( 34.9 2.7% depression ; p=0.0012 ; n=18 ) , which was consistent with published findings [ 14 , 18 , 19 ] . however , epscnmda evoked by ta pathway stimulation recovered to baseline levels following dhpg washout ( 15.2 4.2% below baseline ; p = 0.764 ; n = 18 ) . the difference in ltdnmda between sc - ca1 and ta - ca1 might be due to differences in nmdar subunits at the different synapses . different nmda receptor subunits are associated with different epsc characteristics and the epsc decay time constant ( ) can therefore be used as an indication of receptor subunit population at the synapse . we have previously shown that the decay constant of epscnmda in ta and sc synapses is similar , indicating a similar population of nmda receptor subunits at the two synapses . in order to clarify this further , ro25 - 6981 ( ro ; a potent and selective glun2b receptor antagonist ) was applied ( 1 m ) and a depression of epscnmda was observed in both pathways ( fig . 2 ) . after a period of 1 hour , epscnmda at the sc and the ta pathways had reduced by similar amounts ( sc : 55.8 7.0% ; ta : 43.3 5.2% ; p = 0.18 , n = 6 ) , confirming our previous work suggesting that a similar population of nmda receptor subunits are expressed at these two synaptic populations . the above result suggests that it is unlikely that differences in nmdar subunit composition within the ta - ca1 and sc - ca1 synapses explain the difference in ltdnmda between the two inputs . however , the role of nmda receptor subunits in the induction of ltd was not clear . therefore , to examine directly whether the induction of ltd in the sc pathway was dependent on glun2b activation , dhpg was applied in the presence of ro ( after 1 hour ro application ) . dhpg again caused a depression of the sc pathway ( 30.5 9.6% ; fig . 3 ) that was not significantly different from dhpg - ltdnmda induced under control conditions ( p = 0.617 ) . this suggests that dhpg - ltdnmda in the sc pathway is independent of glun2b subunit activation . ro25 - 6981 causes a similar depression of both pathways following dhpg application to examine whether there was a change in nmdar subunit composition following ltd at the sc synapse , we compared the effect of ro before and following dhpg - ltd ; a change in efficacy of glun2b - selective antagonists is often taken as an indication of a change in subunit composition of nmdars ( i.e. an increase or decrease in the complement of glun2b receptors ; e.g. ) . thus , thirty minutes following dhpg application ( 100 m ; 10 minutes ) when ltd was stably expressed , ro25 - 6981 ( 1 m ; 1 hour ) was bath applied . application of dhpg caused ltdnmda ( 34.0 4.3% depression ) . following ro25 - 6981 application , a further depression of epscnmda was observed ( 63.5 4.2% of pre - dhpg level ; fig . 4 ) that was not significantly different to the depression caused by ro25 - 6981 under control conditions ( p = 0.589 ) , indicating that a change in glun2b subunit composition does not occur as a result of dhpg - ltdnmda . recently we have demonstrated a form of ltdnmda that can be induced by synaptic stimulation at sc but not ta synapses in the hippocampus , using a short theta frequency stimulation ( tfs ; ) . this form of ltd involves activation of group i mglurs and produces metaplasticity such that the induction of subsequent ltp at the sc input is inhibited . in the current study we have utilised pharmacological activation of group i mglurs using the agonist dhpg to elucidate whether the lack of mglur - ltd of epscnmda at the ta input to ca1 in p14 animals is due to insufficient activation of mglurs by tfs . we now show that even with agonist activation of group i mglurs , ltd of epscnmda is not seen at the ta input . instead , despite an initial transient depression , responses recover to baseline levels upon washout of the agonist . a previous study , using 4 - 6 week old animals , has shown that ltd can be induced by dhpg at ta - ca1 input ( although of smaller magnitude than at sc synapses ; ) . this raises the possibility that either developmental changes may impact on the ability to induce mglur - ltd of epscnmda at ta - ca1 synapses or that there is a difference in the ability to induce ltd of epscnmda versus ltd of epscampa at ta - ca1 synapses . however , it is not possible at this stage to be able to make conclusive statements as to the impact of development on mglur - ltd of epscnmda or differences between ltd of epscnmda versus ltd of epscampa at ta - ca1 synapses . plasticity of epscnmda may involve a subunit switch in nmdar subunits [ 11 , 21 ] . as such , the ability to induce such plasticity may be dependent on the subunit composition of nmdars in the basal state . use of glun2b subunit - selective antagonists , such as ifenprodil and ro25 - 6981 , readily allows comparison of subunit composition [ 11 , 21 ] . we found that ro25 - 6981 produced the same the magnitude of depression in sc and ta inputs . similar to our previous work where we found no difference in the decay time constant of epscnmda at the two inputs to ca1 , this suggests that the inability to induce ltdnmda at ta - ca1 synapses is not due to differences in glun2b nmda subunit composition . the sensitivity of epscnmda to ro25 - 6981 in the sc input was not altered following induction of ltdnmda by dhpg . thus , a change in relative glun2a / glun2b subunit expression , which is seen in some forms of plasticity , does not underlie ltd . mglur - ltd of epscampa has been extensively investigated although there is no consensus on the precise mechanism of ltd and multiple signalling cascades have been implicated . ltdnmda induced by tfs is expected to be postsynaptically expressed , as this form of ltd is specific for epscnmda vs epscampa ; postsynaptic expression of pharmacologically - induced ltdnmda has also been demonstrated [ 14 , 18 ] . however , it affects subsequent induction of long - term potentiation ( ltp ; ) and is therefore a form of metaplasticity that will regulate the acquisition of hippocampal - dependent spatial information . future work will aim to elucidate the precise signalling and expression mechanisms underlying this ltd as well as its role in learning and memory . Output:	nmda receptors are composed of multiple subunits and are crucial in the induction of synaptic plasticity and learning and memory . in this study , application of the group i mglu receptor agonist , dhpg , caused ltd of nmda - epscs ( dhpg - ltdnmda ) of the schaffer collateral , but not of nmda - epscs of the temperoammonic pathway onto ca1 neurons of the hippocampus . dhpg - ltdnmda did not alter the sensitivity of nmda - epsc to the glun2b - antagonist , ro25 - 6981 , indicating that the postsynaptic nmda receptor subunit composition remained unchanged following dhpg - ltdnmda . furthermore , blockade of glun2b receptors did not affect the induction of dhpg - ltdnmda . these results demonstrate a difference in the plasticity of nmda receptors between two synapses onto the same ca1 neuron , but indicate that the subunit composition of nmda receptors does not account for this difference .
PubmedSumm2068	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: among stage iiia non - small cell lung cancer patients , t4n0 - 1 cases can be good candidates for surgery and have a relatively favorable postoperative prognosis [ 1 , 2 ] . however , most t4 cases with thoracic vertebral invasion may not be operated as complete resection with vertebrectomy is challenging . we report the case of a male patient with primary lung pleomorphic carcinoma invading the thoracic vertebra who underwent complete resection with en bloc total vertebrectomy after embolization of the tumor - feeding vessels . a 35-year - old japanese man presented with a 6-month history of left back pain . chest x - ray and ct revealed that a tumor ( 43 34 mm ) in s1 + 2 of the left lung had invaded the third thoracic vertebra ( fig . an mri revealed that the tumor had invaded the third thoracic vertebra and the third rib ( fig . 1c ) . as no nodal or distant metastasis was identified by whole - body ct and bone scintigram , we planned a primary surgery with a clinical diagnosis of lung cancer invading thoracic vertebrae ( ct4n0m0 , stage iiia ) . to prevent massive bleeding during resection of the tumor with vertebral invasion , arterial embolization was performed prior to surgery , and the first , second , and third intercostal arteries feeding the tumor ( fig . we performed a complete resection consisting of left upper lobe lobectomy and lymph node resection ( nd2a-2 ) , partial resection of the second and third ribs , and total en bloc spondylectomy of the third thoracic vertebra . pathological examination showed that all resection margins were free from malignant cells , and the final pathological diagnosis was pleomorphic carcinoma of the lung ( pt4n0m0 , stage iiia ) . we conducted 3 cycles of adjuvant chemotherapy ( cisplatin , 80 mg / m ; docetaxel , 60 mg / m ) followed by oral administration of s-1 [ 4 , 5 ] and radiation to the primary tumor field . the patient is alive at 14 months after surgery and without any evidence of tumor recurrence . patients with primary lung cancer invading the vertebral column are rarely operated [ 6 , 7 ] as the postoperative survival is poor ( 5-year survival rates , 1620% ) [ 6 , 8 ] . however , when complete resection is achieved , favorable prognosis might be expected for such patients [ 6 , 7 ] . thus , complete resection with vertebrectomy , especially total vertebrectomy , is challenging in the field of lung cancer surgery . in the present case , we adopted surgical treatment for the following reasons : ( 1 ) no nodal or distant metastasis was identified by whole - body ct and bone scintigram , and ( 2 ) complete en bloc tumor resection can be expected by total vertebrectomy of the third thoracic vertebra . prior to surgery , we performed embolization of the tumor - feeding arteries to prevent possible uncontrollable bleeding during vertebretcomy . we performed surgery first because complete resection is expected by this technique without preoperative induction therapy . in addition , life - threatening or extremely unpleasant spinal cord invasion and/or meningitis may occur when chemoradiation therapy does not result in tumor shrinkage . accordingly , we conducted adjuvant chemotherapy and radiation after surgery . as summarized , a careful preoperative evaluation is essential to determine surgical indication , surgical approach , and sequence of multimodality therapy for t4 tumor with vertebral invasion . Output:	we present a case who had left upper lobectomy with total vertebrectomy after arterial embolization in preparation for intraoperative bleeding . a 35-year - old man complained of left back pain . chest ct revealed a tumor in s1 + 2 of the left lung , invading the third thoracic vertebra . as no nodal or distant metastasis was detected , we performed left upper lobectomy and lymph node dissection ( nd2a-2 ) after embolization of the vessels feeding the tumor in order to reduce intraoperative bleeding . in addition , the team of orthopedics performed en bloc resection of the third thoracic vertebra and parts of the left third and fourth ribs . histological examination of the tumor revealed pleomorphic carcinoma ( pt4n0m0 , stage iiia ) .
PubmedSumm2069	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the mouse b lymphocyte cell lines wehi-231 expressing shrna construct control ( wehi - ct ) or a shrna construct directed against the transcription factor tfii - i ( wehi - tkii - i - kd ) were used to investigate the effect of tfii - i depletion on global gene expression and ctcf binding . to identify genes regulated by tfii - i , we extracted total rna from wehi - ct and wehi - tfii - i - kd from three independent samples . the quantity and the quality of the rna samples were assessed by a nanodrop spectrophotometer and agilent bioanalyser . data preprocessing was carried out with bioconductor package lumi , and we used log2 transformation followed by quantile normalization , . reproducibility between biological replicates was evaluated by calculating the correlation coefficient r ( see example of the scatter plot fig . clustering of the microarray was performed to ensure correct segregation between control and tfii - i knockdown samples ( fig . identification of differentially expressed genes between wehi - ct and wehi - tfii - i - kd was made using bioconductor package we identified 117 genes differentially regulated with a fold chance 2 and p - value 0.05 listed in table 1 . as a confirmation of the knockdown efficiency , we found gtf2i , the gene coding for tfii - i , being the gene the most down regulated in our data . to identify the ctcf binding sites that were affected by tfii - i depletion , we carried two independent chip - seq assays ctcf in wehi - ct and wehi - tfii - i - kd cells with ctcf antibody . briefly , cells were collected and crosslinked with 1% folmaldehyde in pbs for 10 min at room temperature . crosslinking reaction was stooped with glycerine 125 mm and cells were washed with pbs and stored at 80 c until assay was carried out . cells were lysed and dna sheered by sonication with cell lysis / chip buffer ( 0.25% np-40 , 0.25% trinton - x , 0.25% sodium deoxycholate , 0.1% sds , 50 mm tris ph 8.0 , 50 mm nacl , 5 mm edta ) for 15 s , 15 times . lysed cells were centrifuged for 15 min at 14,000 rpm at 4 c and supernantant was collected . 1 mg of protein was precleared for 2 h with protein g agarose beads ( 50% slurry blocked with salmon sperm ) at 4 c . immunoprecipitation was carried out by adding 2 g of antibody and 30 l of agarose g beads and nutated overnight at 4 c . after immunoprecipitation , beads were pelleted by centrifugation and were washed 4 times to remove unspecific binding using buffers with varying concentrations of salt . buffers 1 to 3 contained 0.1% sds , 1% triton - x , 2 mm edta , 20 mm tris ph 8.0 and 150 mm nacl , 300 mm nacl , 500 mm nacl respectively . buffer 4 contained 0.25 m licl , 1% np-40 , 1% sodium deoxycholate , 1 mm edta and 10 mm tris ph8.0 . two additional washes with te were done to remove any residual buffer from the beads . complexes bound to the beads were eluted with 500 l of elution buffer ( 1% sds , 1 mm edta , 50 mm tris ph 8.0 ) at 65 c for 25 min with occasional vortexing . crosslink reversal was achieved by adding 0.2 mm nacl at 65 c overnight . next proteins ( including dna bound factors and antibodies ) were degraded by a treatment with proteinase k , carried at 45 c for 1 h and a second incubation of 15 min at 65 c . pcr purification kit ( qiagen ) was used to retrieve the dna following manufactured instruction and store at 20 c . dna was sent to the iric ( institut de recherche en immunologie et cancrologie , montreal , canada ) sequencing facility where both the library construction and sequencing ( 100bases , paired - end , hiseq2000 , illumina ) were carried out ( table 2 ) . quality of the sequencing was assessed using fastqc software , an example is presented in fig . dna sequences obtained were trimmed to 45 bases , filtered for high quality scores ( > 30 ) , and duplicates were removed before being aligned to the mouse genome ( u.s . national center for biotechnology information ( ncbi ) build 37 , july 2007 , mm9 ) using the bwa algorithm . quality of the alignment was assessed using samstat and only the sequences with mapq score 30 were kept for further analysis ( fig . the model based analysis of chip - seq peak - finding algorithm was used to identify peaks in wehi - ct and wehi - tfii - i - kd conditions using the default settings and an example of peak model obtain with macs is presented in fig . overlap for ctcf binding sites between biological replicates was assessed using the intersect function of bedtools , the results are shown with venn diagram ( fig . homer was used to annotate ctcf peaks , determine their genomic distribution and generated the bedgraph files to visualize the results in ucsc genome browser ( homer.salk.edu/ ) . we used previously published ctcf chip - seq data available in the ucsc genome browser as controls for our dataset ( fig . the mouse b lymphocyte cell lines wehi-231 expressing shrna construct control ( wehi - ct ) or a shrna construct directed against the transcription factor tfii - i ( wehi - tkii - i - kd ) were used to investigate the effect of tfii - i depletion on global gene expression and ctcf binding . to identify genes regulated by tfii - i , we extracted total rna from wehi - ct and wehi - tfii - i - kd from three independent samples . the quantity and the quality of the rna samples were assessed by a nanodrop spectrophotometer and agilent bioanalyser . data preprocessing was carried out with bioconductor package lumi , and we used log2 transformation followed by quantile normalization , . reproducibility between biological replicates was evaluated by calculating the correlation coefficient r ( see example of the scatter plot fig . clustering of the microarray was performed to ensure correct segregation between control and tfii - i knockdown samples ( fig . identification of differentially expressed genes between wehi - ct and wehi - tfii - i - kd was made using bioconductor package we identified 117 genes differentially regulated with a fold chance 2 and p - value 0.05 listed in table 1 . as a confirmation of the knockdown efficiency , we found gtf2i , the gene coding for tfii - i , being the gene the most down regulated in our data . to identify the ctcf binding sites that were affected by tfii - i depletion , we carried two independent chip - seq assays ctcf in wehi - ct and wehi - tfii - i - kd cells with ctcf antibody . briefly , cells were collected and crosslinked with 1% folmaldehyde in pbs for 10 min at room temperature . crosslinking reaction was stooped with glycerine 125 mm and cells were washed with pbs and stored at 80 c until assay was carried out . cells were lysed and dna sheered by sonication with cell lysis / chip buffer ( 0.25% np-40 , 0.25% trinton - x , 0.25% sodium deoxycholate , 0.1% sds , 50 mm tris ph 8.0 , 50 mm nacl , 5 mm edta ) for 15 s , 15 times . lysed cells were centrifuged for 15 min at 14,000 rpm at 4 c and supernantant was collected . 1 mg of protein was precleared for 2 h with protein g agarose beads ( 50% slurry blocked with salmon sperm ) at 4 c . immunoprecipitation was carried out by adding 2 g of antibody and 30 l of agarose g beads and nutated overnight at 4 c . after immunoprecipitation , beads were pelleted by centrifugation and were washed 4 times to remove unspecific binding using buffers with varying concentrations of salt . buffers 1 to 3 contained 0.1% sds , 1% triton - x , 2 mm edta , 20 mm tris ph 8.0 and 150 mm nacl , 300 mm nacl , 500 mm nacl respectively . buffer 4 contained 0.25 m licl , 1% np-40 , 1% sodium deoxycholate , 1 mm edta and 10 mm tris ph8.0 . two additional washes with te were done to remove any residual buffer from the beads . complexes bound to the beads were eluted with 500 l of elution buffer ( 1% sds , 1 mm edta , 50 mm tris ph 8.0 ) at 65 c for 25 min with occasional vortexing . crosslink reversal was achieved by adding 0.2 mm nacl at 65 c overnight . next proteins ( including dna bound factors and antibodies ) were degraded by a treatment with proteinase k , carried at 45 c for 1 h and a second incubation of 15 min at 65 c . pcr purification kit ( qiagen ) was used to retrieve the dna following manufactured instruction and store at 20 c . dna was sent to the iric ( institut de recherche en immunologie et cancrologie , montreal , canada ) sequencing facility where both the library construction and sequencing ( 100bases , paired - end , hiseq2000 , illumina ) were carried out ( table 2 ) . quality of the sequencing was assessed using fastqc software , an example is presented in fig . 2a ( http://www.bioinformatics.babraham.ac.uk/projects/fastqc/ ) . using fastx tool kit ( http://hannonlab.cshl.edu/fastx_toolkit/ ) , dna sequences obtained were trimmed to 45 bases , filtered for high quality scores ( > 30 ) , and duplicates were removed before being aligned to the mouse genome ( u.s . national center for biotechnology information ( ncbi ) build 37 , july 2007 , mm9 ) using the bwa algorithm . quality of the alignment was assessed using samstat and only the sequences with mapq score 30 were kept for further analysis ( fig . the model based analysis of chip - seq peak - finding algorithm was used to identify peaks in wehi - ct and wehi - tfii - i - kd conditions using the default settings and an example of peak model obtain with macs is presented in fig . overlap for ctcf binding sites between biological replicates was assessed using the intersect function of bedtools , the results are shown with venn diagram ( fig . homer was used to annotate ctcf peaks , determine their genomic distribution and generated the bedgraph files to visualize the results in ucsc genome browser ( homer.salk.edu/ ) . we used previously published ctcf chip - seq data available in the ucsc genome browser as controls for our dataset ( fig . here , we described a dataset containing gene expression profiling using illumina beadchips ( microarray ) and chip - seq analysis of ctcf binding in mouse b cell lymphocyte cell lines expressing a shrna construct against tfii - i , a general transcription factor . these data were generated to analyze the influence of tfii - i on the genomic targeting of the epigenetic regulatory protein ctcf , and understand how these two factors co - regulate gene transcription . with this dataset , we were able to show that tfii - i is important for targeting ctcf to a cohort of promoter regions where they co - operate to activate transcription . this finding sheds new light on how ctcf targeting at specific genomic regions can occur . Output:	ctcf is a key regulator of nuclear chromatin structure , chromatin organization and gene regulation . the impact of ctcf on transcriptional output is quite varied , ranging from repression , to transcriptional pausing and transactivation . the multifunctional nature of ctcf is mediated , in part , through differential association with protein partners having unique properties . we identified the general transcription factor tfii - i as an interacting partner of ctcf . to gain an understanding of the function of tfii - i in regulating gene expression and ctcf binding genome wide , we conducted microarray experiments following tfii - i knockdown and chromatin immunoprecipitation of ctcf followed by next generation sequencing ( chip - seq ) from the same tfii - i depleted cells . here , we described the experimental design and the quality control and analysis that were performed on the dataset . the data is publicly available through the geo database with accession number gse60918 . the interpretation and description of these data are included in a manuscript in revision ( 1 ) .
PubmedSumm2070	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the most common disorders of consciousness include vegetative state ( vs ) and minimally conscious state ( mcs ) . patients in mcs show weak but definite self - awareness or environmental awareness , and some patients have spontaneous eye opening and sleep wake cycles . although the patient s cognitive behaviors are intermittent , they are reproducible or can last long enough to be distinguished from primitive reflexive activity 1,2 . although mcs patients remain unable to communicate , the behavioral evidence of self - awareness and environmental awareness is clear . a vs or an mcs may be prolonged and can continue until the patient s death 3 . currently , consciousness assessments mainly rely on clinical signs and various scales . the coma recovery scale - revised ( crs - r ) is an internationally recognized scale for assessing vs 4 . it evaluates auditory , visual , sensory , motor , and communicative responses and can sensitively reflect changes in patients conditions . therefore , the crs - r has become the primary method to diagnose mcs in clinical trials and in follow - up and evaluation . however , clinical scales are very subjective and do not accurately identify small changes in consciousness . electrophysiological examinations have certain advantages for the evaluation of patients with consciousness disorders because of their objectivity and high temporal resolution . many studies have used event - related potential ( erp ) to assess prognoses of patients with consciousness disorders . mismatch negativity ( mmn ) is a negative component appearing in the primary auditory and prefrontal cortices 5 around 100250 ms after any auditory change in a monotonous sequence of sounds ( i.e. an oddball paradigm aaab ) 6 . the low amplitude of mmn implies that many repetitions are needed to observe a response . the mmn is elicited by deviants generated by automatic change detection processes that occur when the current auditory input is different from the preceding one . deviants can be detected if patients are not aware of the auditory changes , provided the changes exceed the patient s discrimination threshold 7 . mmn is even evoked when the patients focus attention on aspects other than the auditory stimuli 8,9 . the p300 is a positive component generated when patients detect a rare and unexpected stimulus in a regular train of standard stimuli ( i.e. oddball paradigm ) 10 . mmn and p300 are two different brain responses elicited by similar stimuli ( deviant or novel ) , and their latencies after stimuli are different . for an auditory potential , p300 appears around 300 ms after stimulus presentation , but for a visual potential , it may appear after 500 or 600 ms . , a p300 wave might reflect either recognition of the target s intrinsic meaning or detection of its acoustic salience . many authors have investigated mmn and p300 in comatose patients , but with inconclusive results 1217 . one evidence - based medical analysis on the predictive values of n100 , mmn , and p300 ( auditory epr components ) found that mmn and p300 had greater predictive value in comatose and other low - response patients 18 . however , it is unclear whether auditory erp can be measured during recovery from coma and whether it can be used to measure sensitive changes in consciousness . moreover , most of these studies assessed patients in the acute stage ; patients in the recovery phase of coma were rarely studied . few clinical studies have observed how these erps change or examined the long - term predictive value of mmn and p300 in prognosis . , we performed a 15-week clinical follow - up on a 42-year - old man with intracerebral hemorrhage with persistent vs for 8 months . during admission wake cycle and flexor responses in the limbs to supraorbital pressure , but did not respond to sound and light stimulation . every 2 weeks , the patient was scored with the crs - r by two specialists . erp examinations were performed at 2 days and 2 , 6 , and 15 weeks after admission . the corresponding crs - r scores were 7/23 , 9/23 , 11/23 , and 11/23 , respectively . before the experiment , brainstem auditory evoked potential testing was performed to exclude hearing impairment . the experiment was approved by the ethics committee of the faculty of medicine of the hospital . written informed consent was obtained from a member of the patient s family and from control participants . auditory stimuli were delivered binaurally through inserted earphones at an intensity of 90 db hl using the presentation software ( e - prime software , electrical geodesics inc . , eugene , orlando , usa ) . the oddball stimulation paradigm with pure tones lasted 75 ms and included frequent tones ( 1000 hz ) and salient tones ( 1500 hz ) . the two tones selected for the acoustic test were determined as 20 and 80% of the 400 hz bandwidth range . we elaborated four sequences of 80 stimulations comprising four first names , with one of them always being the subject s own name ( son ) . the three other first names ( ofns ) were selected from chinese character tables as a series of first names with similar high frequencies . for each sequence , each name was presented 10 times randomly , thus making a complete series of 80 equiprobable first names ( 25% probability for each name ) , with an interstimulus interval between 1300 and 1400 ms . after each recording session , the patient or their family was asked whether any of the other names had particular emotional importance ( e.g. they corresponded to names of close relatives or friends ) . all first names were recorded by the same neutral female voice ( fp ) , and were digitized and replayed binaurally at a 90 db sound pressure level maximal intensity . the e - prime software was used to present the stimuli , and electroencephalography ( eeg ) activity was continuously recorded and analyzed with netstation 4.3 . the data obtained from 128 scalp electrodes mounted on an elastic cap ( electrical geodesics inc . , riverfront research park , eugene , orlando , usa ) were bandpass filtered at 330 hz , notched at 50 hz , and digitized at a sampling rate of 1024 hz . skin / electrode impedance was maintained below 5 k. all scalp signals were referenced online to both mastoids , but were later offline rereferenced to the average of all scalp electrodes . trials were averaged to erps separately for each condition and each patient relative to a 100 ms prestimulus baseline . single epochs with an amplitude of at least 60 v or those containing eye movements and epochs with an amplitude of at least 200 v because of eye blinking or electromyographic artifacts were excluded from averaging . we used standardized low - resolution electromagnetic tomography ( sloreta ) 19 to provide estimates of the cortical sources of the evoked potentials . the loreta solution was included in the egi analysis package ( geosource , electrical geodesics inc . , eugene , orlando , usa ) . loreta was used to compute 3d linear solutions ( loreta solutions ) for the eeg inverse problem within a three - shell spherical head model , including the scalp , skull , and brain compartments . a voxel was labeled as gray matter if its probability of being gray matter exceeded 33% , exceeded the probability of being white matter , and exceeded the probability of being cerebrospinal fluid . in the current implementation , auditory stimuli were delivered binaurally through inserted earphones at an intensity of 90 db hl using the presentation software ( e - prime software , electrical geodesics inc . , eugene , orlando , usa ) . the oddball stimulation paradigm with pure tones lasted 75 ms and included frequent tones ( 1000 hz ) and salient tones ( 1500 hz ) . the two tones selected for the acoustic test were determined as 20 and 80% of the 400 hz bandwidth range . we elaborated four sequences of 80 stimulations comprising four first names , with one of them always being the subject s own name ( son ) . the three other first names ( ofns ) were selected from chinese character tables as a series of first names with similar high frequencies . for each sequence , each name was presented 10 times randomly , thus making a complete series of 80 equiprobable first names ( 25% probability for each name ) , with an interstimulus interval between 1300 and 1400 ms . after each recording session , the patient or their family was asked whether any of the other names had particular emotional importance ( e.g. they corresponded to names of close relatives or friends ) . all first names were recorded by the same neutral female voice ( fp ) , and were digitized and replayed binaurally at a 90 db sound pressure level maximal intensity . the e - prime software was used to present the stimuli , and electroencephalography ( eeg ) activity was continuously recorded and analyzed with netstation 4.3 . the data obtained from 128 scalp electrodes mounted on an elastic cap ( electrical geodesics inc . , riverfront research park , eugene , orlando , usa ) were bandpass filtered at 330 hz , notched at 50 hz , and digitized at a sampling rate of 1024 hz . skin / electrode impedance was maintained below 5 k. all scalp signals were referenced online to both mastoids , but were later offline rereferenced to the average of all scalp electrodes . trials were averaged to erps separately for each condition and each patient relative to a 100 ms prestimulus baseline . single epochs with an amplitude of at least 60 v or those containing eye movements and epochs with an amplitude of at least 200 v because of eye blinking or electromyographic artifacts were excluded from averaging . we used standardized low - resolution electromagnetic tomography ( sloreta ) 19 to provide estimates of the cortical sources of the evoked potentials . the loreta solution was included in the egi analysis package ( geosource , electrical geodesics inc . , eugene , orlando , usa ) . loreta was used to compute 3d linear solutions ( loreta solutions ) for the eeg inverse problem within a three - shell spherical head model , including the scalp , skull , and brain compartments . a voxel was labeled as gray matter if its probability of being gray matter exceeded 33% , exceeded the probability of being white matter , and exceeded the probability of being cerebrospinal fluid . in the current implementation , a spatial resolution of 7 mm is used , producing 2394 voxels . figures 1 and 2 show the mmn and p300 in the normal control participants . the salient tone - induced mmn ( red line , negative wave between 100 and 200 ms ) location was 3 , 52 , 6 ; intensity : 0.01099 na ; brodmann area : 10 ; gyri : medial frontal gyrus in the frontal lobe . the frequent tone ( blue line ) was the same , except that the intensity was 0.011403 na . the son - induced p300 ( red line , positive wave around 300 ms ) location was 3 , 45 , 13 ; intensity : 0.037858 na ; brodmann area : 11 ; gyrus : medial frontal gyrus in the frontal lobe . the ofns ( blue line , positive wave around 300 ms ) elicited a response at 3 , 52 , 6 ; intensity : 0.019716 na ; brodmann area : 10 ; gyrus : medial frontal gyrus in the frontal lobe . eeg , electroencephalography ; erp , event - related potential ; mmn , mismatch negativity . erps to pure tone in the patient during each of the four assessments ( a d ) . erp was first performed 2 days after admission when the patient was in a vs ; the crs - r score was 7/23 . ( b ) erp was again performed 2 weeks after admission ; the patient was in an mcs with a crs - r score of 9/23 . ( c ) erp was again performed 6 weeks after admission ; the patient was in an mcs with a crs - r score of 11/23 . ( d ) erp was again performed 15 weeks after admission ; the patient was in an mcs with a crs - r score of 11/23 . crs - r , coma recovery scale - revised ; erp , event - related potential ; mcs , minimally conscious state ; mmn , mismatch negativity ; vs , vegetative state . the patient s erps to names during each of the four assessments ( a d ) . ( a ) p300 in the first erp assessment 2 days after admission ; the patient was in a vs and had a crs - r score of 7/23 . ( b ) erp was performed 2 weeks after admission ; the patient was in an mcs and had a crs - r score of 9/23 . ( c ) erp was performed 6 weeks after admission ; the patient was in an mcs and had a crs - r score of 11/23 . erp was performed 15 weeks after admission ; the patient was in an mcs and had a crs - r score of 11/23 . crs - r , coma recovery scale - revised ; erp , event - related potential ; mcs , minimally conscious state ; vs , vegetative state . latencies and amplitudes of individually detected waves the mmn amplitudes evoked by salient stimuli were higher than those evoked by frequent stimuli , whereas the latencies of the former were shorter than those of the latter . when the crs - r score indicated vs , the latencies of p300 evoked by sons were longer than those evoked by ofns . however , when the crs - r score indicated an mcs , the latencies of p300 evoked by sons were shorter than those evoked by ofns . for the control participant , mmn amplitudes and latencies evoked by salient and frequent stimuli were roughly the same . the son - evoked p300 amplitude was higher than that evoked by ofn , and the son - evoked p300 latency was shorter than that evoked by ofn . in addition , while studying erp , we carried out topographic eeg and source analysis . the results showed that mmn was generated in the frontal and temporal lobes and that p300 was generated in the frontal , temporal , and parietal lobes . the occurrences of mmn and p300 are indicative of a good prognosis in patients with consciousness disorders 14,2123 . along with increasing recovery of consciousness in these patients , there was a gradual increase in their ability to distinguish auditory stimuli . a sudden increase in mmn amplitude suggested that the patients could interact with their environment . 14 found that mmn could still be recorded in some patients in permanent vs or mcs , indicating some level of awareness in these patients and the possibility of further recovery . however , they did not carry out a follow - up study or describe the outcomes of these patients . 23 carried out an auditory erp study on 22 patients with disorders of consciousness caused by severe traumatic brain injury ( tbi ) and claimed that the occurrence of p300 in these patients indicated better patient outcome . 24 carried out an auditory erp study in 34 patients in a 23-month vs following tbi . the auditory stimuli were 1000 hz pure tones and sons , and they found that the conscious states of the patients in whom p300 was recorded were improved at the 1-year follow - up . 25 carried out an auditory erp study in 50 comatose patients ( including patients with tbi , stroke , and hypoxic - ischemic encephalopathy ) and found that the emergence of mmn could not predict whether patients would regain consciousness . conversely , their results indicated that p300 emergence played a predictive role in determining whether the patients would regain consciousness . taken together , the experimental results implied that the presence of p300 could predict whether patients would regain full consciousness . in the present study , an auditory erp study was carried out on one patient in a persistent vs ( crs - r7/23 ) using pure tone and name stimuli , while mmn and p300 were simultaneously recorded . the patient was evaluated by crs - r at 2 and 6 weeks after his first erp examination . interestingly , the patient s crs - r scores increased ( crs - r 9/23 and 11/23 ) , and intermittent verbal communication appeared . after 15 weeks , the patient s crs - r scores stabilized , and another erp examination was performed . according to the first assessment after he was admitted , the patient was in a vs because of an intracerebral hemorrhage . the disease course was up to 8 months ; thus , the patient should have been diagnosed as being in a persistent vs ( > 6 months ) . however , mmn and p300 were recorded simultaneously during erp examinations , suggesting that the patient might still recover . at 2 weeks after admission , the patient s consciousness state actually improved , indicating that mmn and p300 might occur earlier than improvements on clinical assessments of behavior ; however , this hypothesis needs to be tested in a large sample . however , the mmn and p300 amplitudes did not increase with the improved state of consciousness ( table 1 ) . in addition , the patient s consciousness state did not show significant recovery from 6 to 15 weeks after admission . although the erp examination showed mmn and p300 at all erp examinations , the consciousness state of the patient has not yet significantly improved at 15 months , which may indicate that mmn and p300 can be recorded in patients in permanent vs or mcs , indicating some level of awareness and possibility of further recovery . therefore , the predictive value of mmn and p300 for consciousness recovery needs to be explored in a large sample and with a long - term clinical follow - up . on the basis of our results , we can conclude that mmn was generated in the frontal and temporal lobes , whereas p300 was generated in the frontal , temporal , and parietal lobes . mmn and p300 might emerge earlier than previously believed and continue in persistent vs and mcs . the predictive value of these erps for subsequent recovery of consciousness should be studied in a large sample with long - term clinical follow - up . our group will continue this study and carry out a long - term clinical follow - up to find the predictive value of these erps . Output:	to observe changes in mismatch negativity ( mmn ) and p300 in a patient transitioning from a vegetative state ( vs ) to a minimally conscious state ( mcs ) . one patient with intracerebral hemorrhage and an 8-month disease course was evaluated as being in the vs using the coma recovery scale - revised . two weeks after the patient was admitted to the hospital , another evaluation was performed , and the patient was determined to be in an mcs . using the oddball paradigm , pure tone and name stimuli were presented to the patient to study event - related potentials ( erps ) . a 15-week clinical follow - up was carried out , and four erp examinations were performed at 2 days and 2 , 6 , and 15 weeks after admission . one healthy individual was assessed as a control participant . mmn and p300 were elicited in all four erp examinations . mmn and p300 may occur earlier than believed in patients in persistent vs and mcs ; their predictive values for prognosis need to be further confirmed by follow - up studies on a large clinical sample .
PubmedSumm2071	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: although priapism has been reported as a complication of malignant tumor metastasis , leukemia , sickle cell disease , embolism and spinal canal stenosis [ 1 , 2 , 3 , 4 , 5 ] , there are few reports of priapism as a complication of colorectal cancer in the absence of penile metastasis . here we report a patient with rectal cancer with no penile metastasis in whom chemotherapy resulted in an improvement of priapism . a 65-year - old man was admitted to our hospital for priapism with tenderness of the penis and dysuria . enhanced abdominal computed tomography demonstrated metastatic tumors in the liver , lung , sacrum ( fig . ultrasonography of the penis revealed no tumor and normal blood flow in the penis vessel . chemotherapy ( folfox4 ; levofolinate 25 mg , fluorouracil 250 mg , oxaliplatin 100 mg ) was administered , and although the primary rectal cancer showed no change ( fig . brain or splenic metastasis and hypercalcemia arising from bone metastasis have been reported as rare complications of colorectal cancer [ 6 , 7 , 8 ] . chemotherapy regimens for metastatic colorectal cancer include fluorouracil , leucovorin , either oxaliplatin or irinotecan , and the vegf inhibitor bevacizumab . priapism is a complex involuntary behavioral response that depends on the integration of vascular , endocrine and neurological mechanisms . stimuli for erection can be classified as psychogenic and reflexogenic . when visceral nerves at the sacrum are stimulated electrically , this results in persistent penile erection . nervi erigentes. a second , thoracolumbar erection centre at the level of t12l1 plays a role in psychogenically mediated erections via sympathetic efferent fibers . disturbances in penile blood flow due to metastasis in the penis have been reported to cause priapism . in this case , the cause of priapism might have been micrometastasis in the penis , metastatic tumors that affected the spinal cord , or continuous stimulation of nervi erigentes by a metastatic tumor . chemotherapy , which reduced the metastatic tumor in the sacrum , may also have reduced the level of nervi erigentes stimulation . the results from this case suggest that chemotherapy can be effective for treating rare complications of rectal cancer such as priapism . Output:	a 65-year - old man was admitted with penile tenderness and dysuria due to priapism . enhanced computed tomography revealed metastatic tumors in the liver , lung , sacrum and lymph nodes . advanced rectal cancer , detected by colonoscopy as a primary tumor , was treated with chemotherapy ( folfox4 ) . although the rectal cancer showed no change , five months of chemotherapy improveid the priapism , suggesting that chemotherapy can improve rare symptoms of rectal cancer .
PubmedSumm2072	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: during a 6-year period ( 20102015 ) , there were a total of 2,310 diagnoses of pulmonary adc in our institute . of 2,310 patients , 401 patients underwent a second biopsy or resection for recurrent or metastatic tumors . out of 401 patients , a total of six patients ( 1.5% ) with primary lung adc showed transformed sclc morphology in second biopsy . all patients were treated in the department of oncology , samsung medical center ( seoul , korea ) . clinical and follow - up data were obtained through a retrospective analysis of the medical records , including age , sex , smoking history , treatment , clinical course and follow - ups . all patients were followed until march 2016 with median follow - up period of 39.2 months . the study was approved by the institutional review board at samsung medical center ( 2014 - 14 - 08610 ) . dna was extracted from sections of formalin - fixed , paraffin - embedded ( ffpe ) tissue that was also used for histologic diagnosis . manual microdissection was performed if tumor cell percentages were less than 70% in available samples . genomic dna was extracted using qiagen dna ffpe tissue kit ( qiagen , hilden , germany ) according to the manufacturer s instructions . in cases of lung adc , routine testing for the egfr mutation was performed in the pathology laboratory using peptide nucleic acid mediated clamping polymerase chain reaction ( pcr ) mutation detection kit as previously described , and results were retrieved from electronic medical records . for one sclc sample , the egfr mutation was detected using targeted sequencing via illumina hiseq 2500 ( illumina inc . , san diego , ca , usa ) , which was performed for clinical trial enrollment . for the rest of sclc samples , the egfr mutation was newly evaluated using cobas test , a real - time pcr test as previously described . egfr mutation results were available for all samples except for one that had no residual tumor . in the current study , we used representative ffpe tissue sections for immunohistochemical staining ( ihc ) . ihc for cd56 and thyroid transcription factor 1 ( ttf-1 ) was performed for sclc or combined tumors . staining was performed on 3-m - thick sections from each case using a biotin - avidin - peroxidase method on a bond - max autostainer ( leica , wetzlar , germany ) after retrieval with t / e buffer ( cd56 ) or citrate buffer ( ttf-1 ) . we used primary antibodies to cd56 ( 1:200 , novocastra , newcastle upon tyne , uk ) and ttf-1 ( 1:100 , dako , glostrup , denmark ) . during a 6-year period ( 20102015 ) , there were a total of 2,310 diagnoses of pulmonary adc in our institute . of 2,310 patients , 401 patients underwent a second biopsy or resection for recurrent or metastatic tumors . out of 401 patients , a total of six patients ( 1.5% ) with primary lung adc showed transformed sclc morphology in second biopsy . all patients were treated in the department of oncology , samsung medical center ( seoul , korea ) . clinical and follow - up data were obtained through a retrospective analysis of the medical records , including age , sex , smoking history , treatment , clinical course and follow - ups . all patients were followed until march 2016 with median follow - up period of 39.2 months . the study was approved by the institutional review board at samsung medical center ( 2014 - 14 - 08610 ) . dna was extracted from sections of formalin - fixed , paraffin - embedded ( ffpe ) tissue that was also used for histologic diagnosis . manual microdissection was performed if tumor cell percentages were less than 70% in available samples . genomic dna was extracted using qiagen dna ffpe tissue kit ( qiagen , hilden , germany ) according to the manufacturer s instructions . in cases of lung adc , routine testing for the egfr mutation was performed in the pathology laboratory using peptide nucleic acid mediated clamping polymerase chain reaction ( pcr ) mutation detection kit as previously described , and results were retrieved from electronic medical records . for one sclc sample , the egfr mutation was detected using targeted sequencing via illumina hiseq 2500 ( illumina inc . , san diego , ca , usa ) , which was performed for clinical trial enrollment . for the rest of sclc samples , the egfr mutation was newly evaluated using cobas test , a real - time pcr test as previously described . egfr mutation results were available for all samples except for one that had no residual tumor . in the current study , we used representative ffpe tissue sections for immunohistochemical staining ( ihc ) . ihc for cd56 and thyroid transcription factor 1 ( ttf-1 ) was performed for sclc or combined tumors . staining was performed on 3-m - thick sections from each case using a biotin - avidin - peroxidase method on a bond - max autostainer ( leica , wetzlar , germany ) after retrieval with t / e buffer ( cd56 ) or citrate buffer ( ttf-1 ) . we used primary antibodies to cd56 ( 1:200 , novocastra , newcastle upon tyne , uk ) and ttf-1 ( 1:100 , dako , glostrup , denmark ) . sample information and pathologic features are summarized in table 1 . of the initial samples with diagnosis of adc , the histology of the six adcs was acinar ( n=4 ) , mixed acinar and papillary ( n=1 ) , and mixed acinar and solid ( n=1 ) . of samples that showed transformation to sclc upon second biopsy , four showed pure sclc morphology and two showed combined adc and sclc morphology . in two cases , adc components demonstrated acinar morphology ( fig . ttf-1 was expressed in four of six cases and cd56 was expressed in all five available cases . the clinical information and egfr mutation status of the six patients are summarized in table 2 . initial treatments included complete resection and adjuvant chemotherapy for case 1 ( ct2no ) , palliative chemotherapy for case 2 ( ct3n1m1 ) and case 4 ( ct1n0m1 ) , egfr tki for case 3 ( ct2n3m1 ) , incomplete resection and palliative chemotherapy for case 5 ( ct1n0m1 ) , and complete resection for case 6 ( ct1no ) . the sites of distant metastasis were as follows ; brain ( cases 2 and 4 ) , pleura ( case 3 , 4 , and 5 ) , bone ( case 4 ) , and liver ( case 4 ) . of the six patients with adc in initial biopsy , four ( cases 14 ) harbored an egfr mutation ( l858r mutation , n=1 ; exon 19 deletion , n=3 ) . all four , along with case 5 who had wild - type egfr but was enrolled in a clinical trial of gefetinib , were treated with egfr tkis . cases 1 and 3 were treated with irressa , and afatinib was also added for case 1 . case 2 was treated with afatinib only , and cases 4 and 5 were treated with gefetinib only . all five patients who underwent tki treatment were female non - smokers ( table 2 ) . despite tki treatment , all five patients showed disease progression , upon which a second biopsy was performed . the interval between initial biopsy and second biopsy ranged from 5 to 50 months with mean of 25.3 months . after confirmation of transformation of sclc on second biopsy , four patients received further treatment . the treatment option for two patients ( cases 1 and 3 ) was switched to etoposide and cisplatin , and one of them ( case 3 ) showed partial response . case 5 died due to disease progression and the other patients were alive in the short - term follow - up period . egfr mutation status was compared between initial and second samples , and all pairs showed the same egfr status . the original egfr mutation in cases 14 was retained in all transformed sclc samples , while cases 5 and 6 showed no egfr mutation in transformed sclc samples . sample information and pathologic features are summarized in table 1 . of the initial samples with diagnosis of adc , the histology of the six adcs was acinar ( n=4 ) , mixed acinar and papillary ( n=1 ) , and mixed acinar and solid ( n=1 ) . of samples that showed transformation to sclc upon second biopsy , four showed pure sclc morphology and two showed combined adc and sclc morphology . in two cases , adc components demonstrated acinar morphology ( fig . ttf-1 was expressed in four of six cases and cd56 was expressed in all five available cases . the clinical information and egfr mutation status of the six patients are summarized in table 2 . initial treatments included complete resection and adjuvant chemotherapy for case 1 ( ct2no ) , palliative chemotherapy for case 2 ( ct3n1m1 ) and case 4 ( ct1n0m1 ) , egfr tki for case 3 ( ct2n3m1 ) , incomplete resection and palliative chemotherapy for case 5 ( ct1n0m1 ) , and complete resection for case 6 ( ct1no ) . the sites of distant metastasis were as follows ; brain ( cases 2 and 4 ) , pleura ( case 3 , 4 , and 5 ) , bone ( case 4 ) , and liver ( case 4 ) . of the six patients with adc in initial biopsy , four ( cases 14 ) harbored an egfr mutation ( l858r mutation , n=1 ; exon 19 deletion , n=3 ) . all four , along with case 5 who had wild - type egfr but was enrolled in a clinical trial of gefetinib , were treated with egfr tkis . cases 1 and 3 were treated with irressa , and afatinib was also added for case 1 . case 2 was treated with afatinib only , and cases 4 and 5 were treated with gefetinib only . all five patients who underwent tki treatment were female non - smokers ( table 2 ) . despite tki treatment , all five patients showed disease progression , upon which a second biopsy was performed . the interval between initial biopsy and second biopsy ranged from 5 to 50 months with mean of 25.3 months . after confirmation of transformation of sclc on second biopsy , four patients received further treatment . the treatment option for two patients ( cases 1 and 3 ) was switched to etoposide and cisplatin , and one of them ( case 3 ) showed partial response . case 5 died due to disease progression and the other patients were alive in the short - term follow - up period . egfr mutation status was compared between initial and second samples , and all pairs showed the same egfr status . the original egfr mutation in cases 14 was retained in all transformed sclc samples , while cases 5 and 6 showed no egfr mutation in transformed sclc samples . transformation of nsclc to sclc was recently proposed as a mechanism of resistance to tki therapy . identification of histologic transformation may be an important factor in determining a patient s treatment plan due to the differences between nsclc and sclc . while most reports of transformation of adc to sclc were identified in egfr mutant patients related to tki treatment , it is not certain whether transformation is exclusively related to egfr mutation or egfr tki treatment . here , we report six cases of adc which showed histologic transformation to sclc over a 6-year period at a single institute . similar to previous reports , four cases in our series were adc with egfr activating mutations that underwent tki treatment and were subsequently found to have sclc transformation on second biopsy . however , we also identified two additional cases of sclc transformation that had no egfr mutation , and one of these cases underwent initial tki treatment . egfr tkis are now being used worldwide for first - line treatment in a subset of lung cancers bearing egfr - activating mutations , and they have demonstrated dramatic therapeutic efficacy . one of mechanism of resistance to egfr inhibitors is the histological transformation of adc to sclc . although the presence of combined adc and sclc histology at initial diagnosis is a possibility , genomic sequencing of egfr mutations shows that both the original tumor and transformed sclc at the time of resistance share the original egfr - activating mutation , thus supporting the conclusion that these were not independent tumors [ 6 - 11 ] . however , the small biopsy size represents only a portion of tumors , and sclc components may become dominant at the time of disease progression . of the six patients with initially diagnosed with adc in our report , two were diagnosed using surgically resected samples rather than needle biopsies . for these two cases , the possibility of combined histology at the initial biopsy can be excluded . in case 6 , it was difficult to distinguish between sclc transformation and second primary sclc considering the early stage of initial adc . in our series , this suggests that transformation to sclc is not unique to tumors bearing egfr mutations , nor does it exclusively result from tki treatment . transformation to sclc is also reported as a mechanism of acquired resistance to crizotinib in alk rearranged lung tumors . in addition , transformation to large cell neuroendocrine carcinoma was identified as an acquired resistance mechanism to egfr tkis and crizotinib . recent studies suggest that alveolar type ii cells can give rise to both adc and sclc , so egfr - mutant lung cancers derived from alveolar type ii cells may have the potential to transform into sclc during the disease progression . in sum , it seems that acquisition of neuroendocrine phenotype , which includes sclc transformation , can occur in the progression of disease in both egfr - mutant and egfr wild - type nsclcs . second biopsies are not routinely performed for lung cancer patients when patients showed resistance to tki treatment . therefore , the incidence of transformation to sclc in nsclcs can not be accurately calculated . an acquired tki resistance arising from the histological transformation to sclc has been reported to be as high as 3% . in our institute , the incidence of transformation to sclc identified in the second biopsy of total adcs was 1.5% . recently in our institute , there were two cases of sclc transformation in egfr mutant adc during treatment with azd9291 , an oral irreversible egfr tki with selectivity for activating egfr mutations and the t790 m resistance mutation . identification of small cell component is important as the treatment option can be switched to etoposide and cisplatin against sclc . in conclusion , we report six cases of lung cancer demonstrating transformation from adc to sclc . four cases were egfr - mutant tumors from female non - smokers who underwent tki treatment , and the egfr mutation was retained in the transformed sclc tumors . the other two adcs were egfr wild - type , and one of these patients received egfr tki treatment . the neuroendocrine phenotype can thus be acquired during adc disease progression independent of egfr tki treatment . Output:	background : epidermal growth factor receptor ( egfr ) tyrosine kinase inhibitors ( tkis ) are considered the first line treatment for a subset of egfr - mutated non - small cell lung cancer ( nsclc ) patients . although transformation to small cell lung cancer ( sclc ) is one of the known mechanisms of resistance to egfr tkis , it is not certain whether transformation to sclc is exclusively found as a mechanism of tki resistance in egfr - mutant tumors.methods:we identified six patients with primary lung adenocarcinoma that showed transformation to sclc on second biopsy ( n = 401 ) during a 6-year period . clinicopathologic information was analyzed and egfr mutation results were compared between initial and second biopsy samples.results:six patients showed transformation from adenocarcinoma to sclc , of which four were pure sclcs and two were combined adenocarcinoma and sclcs . clinically , four cases were egfr - mutant tumors from non - smoking females who underwent tki treatment , and the egfr mutation was retained in the transformed sclc tumors . the remaining two adenocarcinomas were egfr wild - type , and one of these patients received egfr tki treatment.conclusions:nsclc can acquire a neuroendocrine phenotype with or without egfr tki treatment .
PubmedSumm2073	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: small bowel volvulus is a rare but dangerous condition that can rapidly cause intestinal vascular compromise and lead to bowel ischemia and necrosis . we present a patient who developed a small bowel volvulus around an adhesion formed between an endotack from a recent laparoscopic hernia repair and the midportion of the patient 's ileum . this unusual complication has not , to our knowledge , been reported in the literature . a 66-year - old male with diabetes mellitus and a history of atrial fibrillation initially presented with longstanding , gradually enlarging bilateral inguinal hernias . both hernias reduced spontaneously . he was medically cleared for surgery and underwent elective , bilateral , total extraperitoneal laparoscopic hernia repair ( tep ) . a small peritoneal tear was made , allowing pneumoperitoneum and interfering with visibility in the preperitoneal space . the tear was closed in the following manner : both sides of the torn peritoneum were held up by smooth graspers so as to assure that no bowel was caught in the repair . both hernia defects were covered with mesh , and they were held in position with a spiral tack placed into cooper 's ligament . the patient 's early postoperative course was uneventful , and he was discharged home that same day . he was doing well 1 week later when seen at a routine postoperative office visit . he had had a normal bowel movement the previous day . when seen in the emergency room , he was found to have high - pitched bowel sounds and a soft , diffusely tender abdomen . a computed tomographic ( ct ) scan was obtained that showed thickened loops of small bowel and ascities . there was a high index of suspicion for mesenteric ischemia because of the patient 's history of atrial fibrillation . the study showed angulation of the ileocolic artery and compression of the superior mesenteric vessels consistent with small bowel volvulus . the patient was taken to the operating room for exploratory laparotomy . upon opening the abdomen , the terminal ileum and the ileocecal valve were necrotic , and there was hemorrhagic necrosis of the adjacent mesentery . the terminal ileum was found to be rotated clockwise around an adhesion extending from the midportion of the ileum to the spiral tack used in the peritoneal repair , which appeared to have eroded through the peritoneum . the volvulus was detorsed in a counterclockwise fashion . the gangrenous bowel , including the ileocecal valve , was resected and a side - to - side functional anastomosis between the small bowel and the cecum was created . the patient 's bowel function returned gradually , and he was discharged home on postoperative day 7 having normal bowel movements and tolerating a regular diet . most cases of acute obstruction after laparoscopic hernia repair are related to bowel herniation through a trocar site . adhesions are another widely recognized cause of intestinal obstruction but are generally found in patients who have undergone intraabdominal procedures and are not usually associated with total extraperitoneal laparoscopic hernia repair . in the literature , adhesions are most commonly discussed as a direct cause of mechanical obstruction . however , when small bowel volvulus occurs , it is frequently associated with an adhesion as its axis . cases of cecal and small bowel volvulus following intra - abdominal laparoscopic procedures have been reported with theories on the cause including adhesions and iatrogenic malpositioning of intestines during surgery . adhesions have been associated with foreign bodies as diverse as sutures , mesh , and umbilical piercing . given these observations , it is not surprising that a tack might be the nidus for adhesion formation and lead to a clinically significant complication . for a tack to serve as one end of an intraabdominal adhesion peritoneal violation may inadvertently occur during total extraperitoneal hernia , as occurred in our case . protrusion of the tack , by any means , could allow contact with the peritoneum and might lead to erosion into the peritoneal cavity . we found no cases of tack erosion into the peritoneum reported in the literature , though this may be because few surgeons would have sought to make this observation . these cases were associated with mesh tacked to cooper 's ligament for use in laparoscopic colposuspension for treatment of urinary incontinence . in these cases , as in our own , it is not clear whether significant perforation occurred at the time of operation or whether postoperative erosion occurred . a tack used to seal a peritoneal tear may have contact with the peritoneal contents by erosion or through a minute but persistent defect . the sharp point that allows spiral tacks to penetrate and hold thick tissues may increase the risk when the device is used in thin tissues . this is particularly treacherous when the sharp edge risks contact with bowel , blood vessels , or other delicate structures . the spiral turns , designed to grip and affix tissues , combined with the sharp point , which can penetrate , irritate , and inflame suggests that the tack , by its very design , is a likely point of adhesion formation . thus , we believe a tack is more appropriately used in parietal tissues and should be avoided in peritoneal repairs . repair of a peritoneal tear occurring during laparoscopic surgery is advocated primarily to improve visualization of the preperitoneal space that is obstructed by a developing pneumoperitonuem . repair also decreases adhesions to the abdominal wall and avoids internal herniation through the tear . endoscopic methods of repair include tacking , stapling , crimping , suturing , and suture loop ligation . one of the rare studies that attempts to evaluate and compare these techniques is done by lau et al . they conclude that stapling or tacking , pretied suture loop ligation , and endoscopic suturing of peritoneal tear are safe and do not differ in the rate of postoperative complications ; however , endoscopic suturing takes significantly longer than the other methods . our case is one instance that calls into question the safety of sharp devices for peritoneal repair . the ability to avoid peritoneal tears increases with the experience of the surgeon , and having an experienced surgeon assist while one is early in the learning curve is widely recommended . however , peritoneal tears remain a relatively frequent complication of this procedure . if visualization is obscured , repositioning the instruments , adding an additional port to facilitate retraction , or placement of a veress needle into the peritoneum to allow decompression are useful techniques . early identification and repair of a tear stops the progression of pneumoperitoneum and facilitates completion of the procedure . wavy or shaking appearance of the peritoneum may be the earliest sigh of a tear during tep . in light of the complication discussed here , we advocate the use of endoloops , which may require serial placement to control a large tear , or suturing , despite the increased time required , as safe and effective methods to repair peritoneal tears . as the dialogue continues , this rare complication is a reminder that every foreign object left in the body has the potential to cause a variety of complications . though use of a tack resulting in adhesion and volvulus is an unusual complication , it is a serious one . it is also avoidable by choosing devices that , by their design , pose a smaller risk of erosion and adhesion , for use in thin and delicate tissues , such as the peritoneum . all foreign bodies must be placed with care to ensure their stability and to minimize protrusion . Output:	background and objectives : we present a case of small bowel volvulus around an endotack applied during total extraperitoneal laparoscopic hernia repair ( tep ) . this complication prompts reconsideration of the role of tacks during extraperitoneal laparoscopic hernia repairs.methods:we undertook a chart review and provide a case presentation with review of the literature.result:a 66-year - old male with bilateral inguinal hernias underwent elective , bilateral , total extraperitoneal laparoscopic hernia repair . during dissection , a small peritoneal tear occurred . the tear was closed with a spiral tack . on postoperative day 22 , the patient developed an acute abdomen . exploratory laparotomy revealed a volvulus rotated around an adhesion to the spiral tack.discussion:volvulus can cause vascular compromise leading to bowel ischemia and necrosis . a tack resulting in adhesion and volvulus is an unusual , but serious , complication . repair of a peritoneal tear during preperitoneal hernia repair is advocated to improve visualization obstructed by a pneumoperitonuem and decrease adhesions to the abdominal wall.conclusion:the use of blunt endoloops or crimps may prove safer than tacks for repairing the peritoneum and placement in proximity to delicate or thin tissues . additionally , careful placement of foreign bodies to ensure their stability and to minimize protrusion may decrease the risk of erosion of the hardware .
PubmedSumm2074	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: triple - negative breast cancer ( tnbc ) accounts for approximately 15% of all breast tumors [ 1 , 2 , 3 ] . it is characterized by a lack of expression of the estrogen receptor , progesterone receptor , and human epidermal growth factor receptor 2 ( her2 ) [ 1 , 3 ] . due to the absence of expression of these receptors , targeted regimens using hormonal or anti - her2 therapy are ineffective , and treatment options are generally restricted to systemic chemotherapies . despite initial responses to standard cytotoxic therapies , relapse is common , and tnbc is associated with an aggressive clinical course . this is reflected by a high rate of visceral metastases , notably including the brain , rapid progression on systemic treatment , and poor survival times [ 1 , 2 , 3 ] . substantial overlap exists between brca1-positive breast cancer and tnbc , in terms of a number of shared clinical and pathological features ; in addition , both frequently exhibit high histologic grade tumors and aggressive courses . although not all patients with tnbc exhibit brca1 mutations , a considerable proportion of brca1 carriers do develop this condition . there is an urgent need for new treatment options in tnbc , particularly for those patients who are also brca1 positive . in the us , no targeted therapies are currently approved for women with tnbc , and cytotoxic chemotherapy remains the standard of care for such patients , including those with brca1 mutations . the angiogenesis inhibitor bevacizumab is one of a number of targeted therapies currently being evaluated in clinical trials . promising results with bevacizumab plus chemotherapy have been reported in patients with tnbc being treated in neoadjuvant , first- , and second - line settings [ 4 , 5 , 6 , 7 ] ; however , no benefit has yet been demonstrated in the adjuvant setting . recently , a novel regimen of bevacizumab combined with carboplatin , as well as nanoparticle albumin - bound paclitaxel particles ( nab - paclitaxel ) , has produced high response rates in phase ii trials of patients with tnbc in neoadjuvant and first - line settings . while the individual contribution of each of the experimental agents in these trials is difficult to determine , the studies suggest that carboplatin plus nab - paclitaxel may be an active cytotoxic combination in patients with tnbc . the case study reported here describes a patient with previously treated triple - negative brca1-positive breast cancer , who showed a strong response to nab - paclitaxel , first in combination with carboplatin and subsequently as monotherapy , after presenting to the author 's clinic with severe metastatic disease . a 51-year - old caucasian woman was admitted with significant abdominal pain in june 2012 . the patient had been diagnosed with cancer of the left breast at a private clinic in april 2008 , and the tumor was initially thought to be a small - cell carcinoma . following the initial diagnosis , the patient was treated with 4 cycles of carboplatin and etoposide ( vp-16 ) ; unfortunately , dosing information is not available for either drug . this regimen was completed by september 2008 and , at that time , the patient had not received any radiation or systemic therapy other than the initial treatment of carboplatin plus etoposide . the patient 's response to this regimen was excellent , with complete regression of the disease . following completion of the initial treatment , a review of the pathology findings established that the tumor was consistent with infiltrating ductal carcinoma , and that it was triple negative . routine follow - up appointments took place , and , in november 2009 , the patient was diagnosed with a recurrent infiltrating carcinoma of the left breast at the same clinic that had initially diagnosed and treated her . pathology tests performed at this time revealed a grade iii , 2.5-cm carcinoma , although neither of the 2 sentinel lymph nodes showed involvement . in addition , the patient 's brca1 status was tested , and she was identified as a mutation carrier . treatment for the carcinoma recurrence took place at loyola university , chicago , from march to july 2010 . the patient received neoadjuvant dose - dense doxorubicin at 60 mg / m plus cyclophosphamide at 600 mg / m once every 2 weeks for 4 cycles . in addition , pegfilgrastim was administered at a dose of 6 mg on the second day of each cycle to stimulate neutrophil production . as there was no evidence of metastasis , this regimen the patient continued to be actively monitored and , in march 2012 , she presented with expressive aphasia . upon further investigation , diffuse brain metastases were discovered , and treatment commenced with whole - brain radiation therapy ( 30 gy in 10 fractions ) at loyola university . this resulted in a significant improvement of the patient 's symptoms and regression of the brain metastases . follow - up continued at loyola university until june 2012 when , during a routine clinic visit , the patient experienced severe abdominal pain and was admitted to the author 's clinic . a ct scan of the abdomen and pelvis performed on the day of admission revealed a large mixed cystic and solid mass in the region of the porta hepatis , posterior to the uncinate process of the pancreas . this scan also revealed worrying signs of metastasis , namely necrotic adenopathy ; celiac , retroperitoneal , and left iliac chain adenopathy ; multiple tiny hypodensities in the liver , and right renal nodules of abnormal appearance . the following day , a separate ct scan of the chest found more than 20 diffuse , bilateral , pulmonary metastases . figure 1 shows abdominal and chest ct scans of the patient taken prior to treatment . urgent radiation therapy to the abdomen commenced that day , as the patient had reported a maximum score of 10 on a rating scale for her abdominal pain . in view of the patient 's history , the underlying cause was suspected to be metastatic breast cancer . a biopsy of the left lung mass was performed on the third day of admission , and a general pathologist identified non - small cell carcinoma . after discussing the pathology findings , it was agreed that the cancer was very poorly differentiated and high grade , and , although it was likely to be of lung origin , it was difficult to exclude other cancers , including malignancies of the breast or abdomen . the patient elected to continue treatment at the author 's clinic and , on june 12 , 2012 , she started a carboplatin plus nab - paclitaxel regimen . the dosing schedule was carboplatin 450 mg ( area under the curve = 5 ) once every 3 weeks , plus nab - paclitaxel at 100 mg / m once a week for 3 weeks in a 4-week cycle . despite filgrastim injections to stimulate neutrophil production , carboplatin was removed from the regimen after 4 cycles of treatment due to the persistent occurrence of neutropenia , which resulted in frequent dose delays ( see table 1 ) . however , from the end of august 2012 , the dose was reduced to 90 mg / m once a week for 3 weeks in a 4-week cycle , due to a decrease in body weight , body mass index , and body surface area . whole - body ct scans , including brain imaging , were scheduled to be performed on a regular basis , along with assessments of bone scans , laboratory results , and tumor markers . in this regard , a whole - body ct scan performed in october 2012 revealed that the patient had responded significantly to the therapy , as evidenced by a reduction of at least 50% in bulky tumor burden . the patient also reported decreased abdominal pain and shortness of breath . the positive response to nab - paclitaxel the number of metastases in the lung appeared to have reduced to 1 , and the large abdominal mass had decreased in size . figure 2 shows abdominal and chest ct scans taken after treatment with nab - paclitaxel . at follow - up in march 2013 , the patient was continuing to receive nab - paclitaxel and confirmed that the treatment had been well tolerated , with no treatment - related adverse events . this contrasted significantly with her original presentation to the author 's clinic , when she was bedridden with severe disease and related symptoms . a brain mri scan carried out on april 18 , 2013 , as part of routine tumor surveillance , revealed a recurrence of the brain metastases . however , whole - body ct scans carried out 5 days later confirmed that the patient 's response to nab - paclitaxel was still being maintained in all areas other than the cns . on april 24 , treatment was initiated with temozolomide 100 mg / m daily on days 17 and 1521 of each 28-day cycle , plus irinotecan administered at a dose of 125 mg / m once every 2 weeks . unfortunately , the patient 's condition deteriorated and she died on july 2 , 2013 . it is well established that tnbc is associated with a particularly aggressive clinical course [ 1 , 3 ] . therapeutic options are limited , and standard cytotoxic chemotherapy remains the mainstay of treatment , despite the disease progressing quickly in the majority of women [ 1 , 2 , 3 ] . median survival times of around 1 year have been reported along with a median duration of response to first - line therapy of just 12 weeks . there is an urgent need for the development of novel treatment strategies that will significantly increase survival times . nab - paclitaxel is an albumin - bound form of paclitaxel with a mean particle size of approximately 130 nm . it is able to target delivery of paclitaxel to tumors by interacting with albumin receptors involved in drug transport . nab - paclitaxel displays an improved therapeutic index compared with standard solvent - based paclitaxel , allowing the safe infusion of higher doses in a shorter time span with no requirement for premedication . nab - paclitaxel is currently approved for the treatment of breast cancer after either failure of combination chemotherapy for metastatic disease or relapse within 6 months of adjuvant chemotherapy . recent reports have shown the potential for nab - paclitaxel to be used in the treatment of tnbc . for example , a japanese case study reported a complete response in a patient with tnbc and lymph node metastases who received 8 cycles of adjuvant nab - paclitaxel therapy following surgery . however , the majority of ongoing research with nab - paclitaxel in tnbc is in combination with bevacizumab , which itself has provided modest improvements in efficacy versus chemotherapy alone in neoadjuvant , first- , and second - line settings [ 4 , 5 , 6 , 7 ] . two phase ii studies have recently reported promising activity with the combination of nab - paclitaxel , bevacizumab , and carboplatin in first - line and neoadjuvant treatment of patients with tnbc , and the combination was deemed to be well tolerated . another phase ii study examined the efficacy and safety of nab - paclitaxel administered in combination with bevacizumab and gemcitabine as first - line treatment for her2-negative , metastatic breast cancer . a subset of 13 patients was identified as having tnbc , and this cohort demonstrated a clinical benefit rate defined as complete response , partial response , or stable disease of 85% . a case study has been published of one of the tnbc participants in this trial , who achieved long - term complete remission of almost 2 years and a progression - free survival of just over 2.5 years . as a consequence of these findings , the phase ii / iii tnacity trial ( www.clinicaltrials.gov nct01881230 ) will compare nab - paclitaxel plus gemcitabine with nab - paclitaxel plus carboplatin , and also with gemcitabine plus carboplatin , with the aim of clarifying how to best use existing drugs in patients with tnbc . the case study described in this paper adds to previous trials and reports indicating the potential efficacy of nab - paclitaxel in tnbc , either as monotherapy in a later - line setting , or in combination with bevacizumab and chemotherapy agents at an earlier stage . to the author 's knowledge , this is the first account of a patient with metastatic , triple - negative , brca1-positive breast cancer achieving a strong response to treatment with nab - paclitaxel , which was sustained from the first routine whole - body ct scan in october 2012 until the recurrence of brain metastases in april 2013 . this case study also suggests that nab - paclitaxel is well tolerated , adding to the argument in favor of further investigations of this agent in metastatic tnbc , particularly in patients who are also brca1 positive . Output:	our case describes a 51-year - old female , diagnosed in april 2008 with a triple - negative , brca1-positive , infiltrating ductal carcinoma of the left breast . initial platinum - based therapy resulted in a complete regression until november 2009 , when a recurrence of the disease was detected . as no evidence of metastasis was found , a dose - dense regimen of doxorubicin plus cyclophosphamide was administered , followed by paclitaxel . the patient was actively monitored until march 2012 , when brain metastases were discovered and successfully treated with whole - brain radiation therapy . three months later , the patient experienced severe abdominal pain , and ct scans revealed extensive metastatic disease , including a large mass in the abdomen and more than 20 bilateral pulmonary metastases . treatment commenced with carboplatin plus nab - paclitaxel . however , carboplatin was stopped after 4 cycles due to persistent neutropenia , and nab - paclitaxel was continued as monotherapy . whole - body ct scans performed in october 2012 and march 2013 revealed a significant response to therapy , and the patient reported feeling well and being fully mobile . no treatment - related adverse events were observed . a routine brain mri scan carried out on april 18 , 2013 , revealed a recurrence of brain metastases ; however , ct scans confirmed that disease progression was not systemic , but confined to the central nervous system . despite the initiation of treatment with irinotecan plus temozolomide on april 24 , the patient died on july 2 , 2013 . the author believes that this case is the first report of a robust response to nab - paclitaxel monotherapy in triple - negative brca1-positive breast cancer , and that it supports further studies of nab - paclitaxel in this aggressive indication .
PubmedSumm2075	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: acromegaly is a syndrome seen in adults due to excessive production of pituitary growth hormone as a result of an adenoma or hyperplasia of the eosinophilic cells of the anterior pituitary gland . the main clinical features of acromegaly are skeletal overgrowth affecting the hands , feet , and skull along with spinal deformities like kyphosis . the interface between psychiatry and endocrinology is well documented and psychiatric problems in various endocrine disorders are well known . psychiatric issues in acromegaly have been addressed sparsely and no systematic studies exist for the same , though anecdotal case reports are present in literature . one case depicts pure depression with acromegaly , while another reports the presence of depression with pathological gambling . other psychiatric symptoms like schneiderian first - rank psychotic symptoms , persecutory delusions , visual and auditory hallucinations , apathy with lack of motivation and mood swings have all been documented in acromegaly . in all the cases , response to various medications used has been excellent bringing about a reduction of symptoms . one study on 12 patients with acromegaly failed to prove a relationship between psychopathology and growth hormone levels , while it did mention a sexual dichotomy with 11 patients being female . a 33-year - old lady from middle socioeconomic class presented with depressed mood , loss of appetite , lack of sleep at night , inability to concentrate on her day - to - day activities , and occasional passive suicidal themes in her thoughts . the patient had a similar episode three years ago and had responded well to sertraline . the patient 's father had a history of depression with psychotic features and during his hallucinatory episodes had committed suicide by jumping into a well . the facial features noted by us were prognathia of the mandible , enlargement of the supro - orbital ridges , enlarged facies , and macroglossia . on testing after endocrine consultation , the patient had a high fasting blood sugar of 185 mg% while post lunch blood sugar was 210mg% . the level of growth hormone was 7.2 ng / ml ( normal 0.044.5 ng / ml ) . a ct scan revealed an enlarged sella tursica and a thickened skull with pituitary enlargement . the patient was started on escitalopram 10 mg / day with zolpidem 10 mg at night for depressive symptoms . the patient was also called weekly for cognitive therapy sessions where suicidal and negative thoughts would be addressed . the patient started responding showing a 20% improvement in the first week itself with 80% symptom reduction at the end of four weeks . the patient was to undergo treatment with the endocrinologist for acromegaly but was noncompliant and did not follow - up after the fifth week . during the five weeks of clinical observation , improvement was 80% with no worsening of acromegaly symptoms . it was difficult to label the depression secondary to her acromegaly as the endocrine features did not bother the patient and were detected by us . a strong family history of depression with psychotic features and suicide of her father may have contributed to her depression . our limited knowledge and lack of data on the relationship between depression and acromegaly makes it very difficult to comment on any associations . Output:	depression is one of the commonest disorders encountered in general hospital psychiatry . acromegaly is a condition with excessive growth hormone secretion that may at times present with oversychopathology . we present the case of a 33-year - old lady with depression and acromegaly that successfully resolved after treatment with escitalopram and cognitive therapy .
PubmedSumm2076	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: diverse morphological tooth anomalies are found in various shapes , textures , and prevalence . they can be sporadic or syndromes , nutritional , environmental , genetic , and ethnic related . most of these tooth - born deformations ( e.g. , shovel - shaped incisors , tuberculum carabelli , variations in number and shape of cusps , dens invaginatus , taurodontism , palatogingival groove ) have been extensively investigated . labial - cervical - vertical groove ( lcvg ) , which starts on the cervical enamel and extends to the radicular surface , has also been described as a notch . it is assumed that this malformation is a developmental anomaly in which an infolding of the enamel organ and hertwig 's epithelial root sheath create a groove on the labial surface of permanent maxillary incisors . causative factors for this malfunction may be due to trauma , disease , and nutritional issues , or can be genetic or idiopathic . lcvg runs vertically from the crown surface to the root , starting at the enamel on the crown and extends apically , crossing the cementoenamel junction and resembling a short furrow . this furrow gradually grows deeper in the apical direction and may , occasionally , run throughout the root surface . anatomic factors , such as cervical enamel projections , enamel pearls , and radicular grooves are associated with advanced localized periodontal destruction . the presence of lcvg may exacerbate some clinical aberrations , such as esthetic deficiency of the gingival marginal contour , accumulation of plaque and , consequently , gingival pocket with bone loss as well as failure in endodontic and periodontal treatments . describe a 25-year - old female in which a periodontal lesion with vertical bone loss was directly related to the labial groove on a maxillary central incisor . this type of dental deformity has been scarcely investigated and reported in the dental literature . to the best of our knowledge and based on the available data , there are no reported cases of intrabony defect associated with lcvg treated with platelet rich fibrin ( prf ) . this article reports the effectiveness of prf for the treatment of intrabony defect associated with lcvg in the area of maxillary left central incisor . a 47-year - old male patient reported to the department of periodontology , with a chief complaint of pus discharge from maxillary left central incisors with dull intermittent pain . the patient was a non - smoker . on clinical examination , a localized gingival inflammation was present with soft edematous tissue with the accumulation of plaque and calculus with # 11 and # 21 . periodontal examination depicts a pocket depth of 11 mm on mesial aspect and 8 mm on midbuccal aspect of # 21 [ figure 1 ] with no mobility . fremitus was found to be negative precluding the possibility of trauma from occlusion and tooth was found to be vital electric and thermal pulp testing . on radiographic examination , a tear - shaped radiolucency was present with localized bone loss in # 21 [ figure 2a ] . on careful examination , clinical symptoms subsided , but the deep pocket was still present and hence a periodontal surgery was carried out to eradicate the underlying problem . the procedure was explained to the patient and the consent was obtained for the same . routine blood investigations were done . after assuring surgical asepsis , a pre - procedural rinse with 0.2% after administration of local anesthetic ( 2% lignocaine with 1:80000 adrenalin ) , the papilla preservation flap ( takei et al . ) was adopted since there was sufficient space between the maxillary central incisors . the exposed root surfaces were thoroughly scaled and root planed and bone defect carefully curetted . lcvg noted on # 21 [ figure 3a ] , terminated in the cervical third of root surface was sealed with glass ionomer cement ( fuji ii ; gc corporation , tokyo , japan ) [ figure 3b ] . pre operative pocket depth of 11 mm on mesial aspect and 8 mm on midbuccal aspect of # 21 ( a ) pre operative radiolucency with # 21 ; ( b ) post operative radiograph with # 21 at 6 months ( a ) lcvg noted on # 21 ; ( b ) lcvg was sealed with glass ionomer cement the prf was prepared following the protocol developed by choukroun et al . briefly , the procedure of prf preparation : 10 ml intravenous blood was collected by venepuncture at the antecubital fossa . this was transferred into 10 ml sterile tube without anticoagulant and immediately centrifuged at 3000 rpm for 10 min . fibrin clot formed in between the acellular plasma on top and the red blood cells at the bottom were separated using sterile tweezers and scissors [ figure 4 ] . amoxicillin 500 mg thrice for 7 days and analgesic aceclofenac 100 mg twice for 3 days . prf was placed in the intrabony defect with # 21 then , 1 week postoperatively , the dressing was removed and saline irrigation was done . the patient was monitored at regular intervals and was under maintenance therapy . at the end of 3 months and 6 months the clinical measurements were repeated and compared to the baseline values . on examination during subsequent follow - ups , treated area showed satisfactory healing without any post - operative complications . the pocket depth was 2 mm on mesial and 1 mm on midbuccal aspect at 6 months [ figure 6 ] . periodontal health was stable and bone regeneration was noticed in radiograph [ figure 2b ] . post operative pocket depth of 2 mm on mesial and 1 mm on midbuccal aspect of # 21 at 6 months periodontitis is a multifactorial infectious disease affecting primarily a subset of subjects and subset of sites . recent microbiological data have acknowledged that before disease progression can occur , a susceptible host and site are required , in addition to the presence of pathogenic bacteria . retrospective studies clearly suggest a strong association between anatomic aberrations ( lcvg ) and periodontal attachment loss . anatomic aberrations like root grooves can occur here , which form stagnant sites or ecological niches favoring the retention and growth of micro - organisms . grooves may facilitate plaque growth by providing surface areas sheltered from cleaning efforts as well as host defense mechanisms . later , bacterial selection and growth may be influenced by anaerobic conditions established inside the grooves . these grooves can be classified into mild , moderate , and complex based on its depth and extent . it has been pointed out that defect may vary from a simple , shallow developmental groove , to a complete lack of closure of calcified tissues , allowing for a direct soft tissue connection between the pulp and the periodontium . the presence of lcvg may exacerbate some clinical aberrations , such as esthetic deficiency of the gingival marginal contour , accumulation of plaque and , consequently , gingival pocket with bone loss , as well as failure in endodontic and periodontal treatments . describe a 25-year - old female in which a periodontal lesion with vertical bone loss was directly related to the labial groove on a maxillary central incisor . we have also observed an intrabony defect directly related to lcvg on # 21 . in the presence of periodontal disease , the therapeutical options for lcvg consist of grinding and flattening the affected area of the tooth or placement of a physical barrier between the tooth and soft tissue flap . the intra osseous defect , if present can be grafted with various bone graft materials . it is considered as a second - generation platelet concentrate and has been used in various surgical procedures in an attempt to enhance wound healing . it is prepared from the patient 's own blood thereby eliminating the possibility of disease transmission or foreign body reactions . blood is drawn into standard glass / silica coated blood collection tubes and centrifuged at a predetermined speed to ensure cell separation . this unique preparation technique allows prf to trap at least 95% of the platelets of the collected blood into a fibrin mesh . the fibrin mesh can then be easily manipulated into a membrane that allows it to be transferred to any surgical site . here , high concentrations of the collected platelets allow for the slow release of growth factors ( gfs ) from the platelet granules . these gfs include vascular endothelium growth factor ( vegf ) , platelet - derived growth factor ( pdgf ) , fibroblast growth factor ( fgf ) , epidermal growth factor ( egf ) , hepatocyte growth factor ( hgf ) , insulin - like growth factor ( igf ) , and transforming growth factor- ( tgf- ) . all of these play a role in replacing lost tissue , resurfacing of the wound , and restoring vascular integrity . compared to other platelet concentrates , prf releases these factors at a sustained rate over a longer period , thereby optimizing wound healing . recently , prf has also been shown to stimulate the growth of osteoblasts and periodontal ligament cells , both of which are significant for the regeneration of periodontal defects . currently , prf has been successfully tested in a number of procedures including maxillofacial surgery , periodontal surgery , and implantology . mazor et al . successfully used prf as the only grafting material in a series of sinus augmentation procedures . in a similar 6-year follow - up study , simonpeiri et al . were able to demonstrate that using prf as a sole grafting agent was a viable long - term option in sinus augmentation procedures . prf has also been used successfully to treat periodontal defects . in a series of clinical trials conducted by pradeep and sharma it was shown that prf could be used as a guided tissue regeneration ( gtr ) membrane to affect periodontal regeneration in three wall bony defects and degree ii furcation lesions . published several case reports showing the successful use of prf membranes in the management of both single and multiple gingival recession defects . and shah confirmed this observation and showed that prf could be used for root coverage procedures . in the presented case , lcvg was sealed with glass ionomer cement ( fuji ii ; gc corporation , tokyo , japan ) and associated intrabony defect was treated with prf . the lcvg is a potential problem area , which should be evaluated and treated accordingly . the groove can lead to combined endodontic periodontic lesions , which are difficult to deal with ; attention has been brought to its presence and clinical significance . Output:	a 47-year - old male patient reported to the department of periodontology , with a chief complaint of pus discharge from maxillary left central incisors with dull intermittent pain . on clinical examination , a localized gingival inflammation was present with soft edematous tissue with the accumulation of plaque and calculus with # 11 and # 21 . periodontal examination depicts a periodontal pocket depth of 11 mm on mesial aspect and 8 mm on midbuccal aspect of # 21 with no mobility . on radiographic examination , a tear - shaped radiolucency was present with localized bone loss in # 21 . on careful examination , labial - cervical - vertical groove ( lcvg ) was found on # 21 which was extending into gingival sulcus . this article reports the effectiveness of platelet rich fibrin for the treatment of intrabony defect associated with labial - cervical - vertical groove of # 21 .
PubmedSumm2077	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the primary function of the immune system is to protect the host from infectious microbes in its environment . this system has evolved over millions of years , in response of coexistence with microorganisms . basically , the system can be divided in two components , the innate and adaptive immunities . the innate also called natural immunity refers to a nonspecific response that involves the recruitment of diverse components of the immune system such as macrophages , neutrophils , natural killer cells ( nk cells ) , cytokines , several cellular receptors , complement components , cytokines , toll - like receptors ( tlrs ) , and antimicrobial peptides ( amps ) . this response is phylogenetically older in comparison to the adaptive immunity , which involves recognition of specific antigen , conferring both specificity and a memory effect . t cells recognize antigen in the form of peptide bound to major histocompatibility complex ( mhc ) molecules . b cells have immunoglobulin receptors that recognize the antigenic portions of determined molecules . in organ transplantation , the adaptive immunity is considered the main response exerted to the transplanted tissue , since the principal target of the immune response is the mhc molecules expressed on the surface of donor cells . however , we should not forget that the innate and adaptive immunities are divided only by educational purposes , since both are codependent . for example , t - cell activation leads to the production of cytokines and chemokines which in turn may recruit components of the innate immunity like nk cells or macrophages . furthermore , local tissue production of complement components seems to be essential for full t - cell activation , and some amps like defensins and cathelicidin have chemoattractant properties on t lymphocytes . because the immune system uses many different effector mechanisms to destroy the broad range of microbial cells and particles that it encounters , it is critical for the immune response to avoid unleashing these destructive mechanisms against its own tissues . this avoidance of destruction of self - tissues is referred to as self - tolerance . mechanisms to avoid reaction against self - antigens are expressed in many parts of both the innate and the adaptive immune responses . unfortunately , transplanted tissues from individuals of the same species ( allogenic ) or different species ( xenogeneic ) are recognized as nonself , causing graft rejection . the process by which the immune system recognizes pathogens , tumors , and transplantation antigens involves the same antigen recognition molecules . the rejection response to grafted tissue is caused by cell surface molecules that induce an antigenic stimulus . a wide variety of transplantation antigens have been described , including the mhc molecules , minor histocompatibility antigens , abo blood group antigens , and monocytes / endothelial cell antigens . the minor histocompatibility antigens are processed peptides derived from cellular antigens that are presented by mhc molecules but are not derived from the mhc . however , abo incompatibility can result in hyperacute rejection of primarily vascularized grafts , such as kidney and heart . as we mentioned before , the principal target of the transplantation immune response is the mhc molecules expressed on the surface of donor cells . according to their relative potencies in eliciting rejection , the major antigens in mammalian species are encoded by a closely linked series of genes called mhc . in humans , these genes reside in the short arm of chromosome 6 ( figure 1(a ) ) . organs transplanted between mhc identical individuals are readily accepted , whereas organs transplanted between mhc antigen - mismatched individuals are rejected in the absence of immunosuppressive therapy [ 8 , 9 ] . since the mhc was first defined in mice by gorer and snell [ 10 , 11 ] , the world health organization nomenclature committee has named hla ( human leukocyte antigen ) to the human mhc . the hla complex genes and their protein products have been divided into three classes ( i , ii , and iii ) on the basis of their tissue distribution , structure , and function [ 13 , 14 ] . mhc class i and ii genes encode codominantly expressed hla cell surface antigens , and class iii genes encode several components of the complement system ; all share important roles in immune function . class i mhc antigens are present on all nucleated cells and are composed of a 45-kd transmembrane heavy chain encoded by genes of the hla - a , hla - b , or hla - c loci on chromosome 6 ; the heavy chains are associated noncovalently with a 12-kd protein , 2-microglobulin , encoded by a gene on chromosome 15 ( figure 1(c ) ) . additional ( nonclassical ) class i molecules , like those encoded by the hla - e , -f , -g , -h loci , have been described and show limited variability and tissue distribution . the precise functions of these molecules are not yet clear , although they have been implied in presenting carbohydrate and peptide fragments to t cells and mother 's immunological tolerance of the fetus [ 1417 ] . mhc class ii antigens are expressed only on b lymphocytes , activated t lymphocytes , monocytes , macrophages , langerhans cells , dendritic cells , endothelium , and epithelial cells . class ii molecules are heterodimers composed of noncovalently associated and polypeptide chains chains encoded by genes of the hla - d region ( figure 1(b ) ) . there are 3 major class ii proteins designated , hla - dp , hla - dq , and hla - dr . class iii genes are located between the hla - b and hla - d loci and determine the structure of three components of the complement system : c2 , c4 , and factor b [ 13 , 19 ] . class i mhc molecules present cytoplasm - derived peptides , or intracellular parasites , principally viruses ; whereas mhc class ii molecules bind peptides derived from extracellular proteins . hla class i and ii molecules are recognized by cd8 and cd4 positive t cells , respectively [ 2022 ] . also , nk cells may recognize hla classical and nonclassical type i molecules [ 2325 ] . hla genes are almost always inherited together , thus the antigens of the entire hla region inherited from one parent collectively are called haplotype . because chromosome 6 is an autosome ( a chromosome with two pairs ) , all individuals have two hla haplotypes ( one for each chromosome ) . according to this , any sibling pair has a 25% chance of inheriting the same two parental haplotypes , a 50% chance of sharing one haplotype , and a 25% chance of having two completely different haplotypes . since the biologic function of the hla molecules is presenting endogenous and exogenous antigens , they manifest high structural polymorphism . until 2010 , mutations in microbial antigens might permit the microbe to avoid binding ( and , consequently , recognition ) by a few hla alleles , but no mutations will permit the microbe to avoid recognition broadly throughout the population ; assuring then , the continuity of species in the presence of pandemic infection . in transplantation immunology , the major impact in graft loss comes from the effects of hla - b and -dr antigens . hla - dr mismatch effect is the most important in the first 6 months after transplantation , the hla - b effect emerges in the first 2 years , and hla - a mismatches have a deleterious effect on long - term graft survival [ 2832 ] . the phenomenon by which the recipient immune system reacts with donor antigens that are considered to be non - self is named allorecognition . the main and strongest responses to alloantigens are mediated by host t cells , which recognize peptide antigens presented in the context of mhc , by antigen - presenting cells ( apcs ) . however , evidence that the innate alloimmunity has an important role in graft rejection has recently been proposed by land and coworkers [ 33 , 34 ] . they state in their injury hypothesis that initial allograft injury reflected by reactive oxygen species ( ros ) during reperfusion is associated with generation of damps ( meaning damage - associated molecular patterns ) such as heat shock proteins ( hsp ) and hyaluronan fragments ( fha ) among others , all of which are recognized by tlr4 and/or tlr2 . subsequent tlr4- and tlr2-triggered signaling pathways utilize adaptor proteins including myd88 ( myeloid differentiation marker 88 ) , which in turn initiate downstream signaling pathways that lead to activating the 3 master transcription factors nf-b ( nuclear factor - kappa b ) , ap-1 ( activator protein-1 ) , and irf-3 ( interferon regulatory factor 3 ) . nf-b seems mainly to be responsible for maturation of donor - derived and recipient - derived dendritic cells , which represents the bridge to development of an adaptive alloimmune response that results in rejection . certainly , further studies are needed to determine the extension and importance of this branch of the immune system in transplant rejection and/or tolerance . in adaptive allogenic immune response , the foreign or donor antigen presentation to t cells may occur by three ways ( figure 2 ) : ( 1 ) indirect recognition : donor 's hla molecules can be processed by apc ( antigen presenting cells ) from a receptor , then they are fractionated into peptides as well as other bacterial antigens and are presented according to the same route as the hla in the receptor . this type of mechanism has a dominant role in chronic rejection [ 3741 ] ; ( 2 ) direct recognition : the donor 's hla molecules can be recognized directly on the donor - presenting cells , without requiring antigen processing by receptor . in these circumstances , it could be said that the receptor identifies the foreign hla molecule as an own molecule with a foreign peptide . this mechanism determines a strong immune response in the acute rejection [ 37 , 38 , 4250 ] ; ( 3 ) a third mechanism could be mediated by immunoglobulin - like receptors of natural killer ( nk ) cells . in this mechanism , the activation of nk receptors promotes the inactivation of nk cells and cytotoxic t lymphocytes as well . these receptors recognize polymorphic sequences of hla - c , -b , or -a in the target cells . the absence of these sequences in the cell would make them sensitive to cytolysis and therefore the loss of tolerance [ 5156 ] . recently , it was shown that both nave and memory cd4 and cd8 t cells are frequently cross - reactive against allogeneic hla molecules and that this allorecognition exhibits exquisite peptide and hla specificity . such advances in the understanding of the immunogenetics of allorecognition have led some researches to suggest a new model for allorecognition whereby the majority of t cell alloresponses may occur via direct recognition ( cross - reactivity ) by thymically educated nave and memory t cells against allogeneic hla molecules presenting self - peptides . according to this model , thymically educated t cells are commonly and specifically allo - hla reactive and are activated by viral infection or vaccination to become alloreactive memory t cells which are a major barrier to successful tolerance . to support the transplant programs , several clinical laboratories perform various hla tests , including hla typing of the recipient and the donor , screening of hla antibodies in the recipient , and detection of antibodies in the recipient that are reactive with lymphocytes of a prospective donor ( cross - matching ) . historically , hla typing was conducted by serologic testing by using antiserum in complement - dependent cytotoxic assays . recently , more precise dna - based hla typing methods using molecular techniques , such as sequence - specific oligonucleotide probe hybridization , sequence - specific primer amplification , sequencing - based typing , and reference strand - based conformation analysis , have been developed and are frequently used . there is a clear relationship between the degree of hla matching and kidney graft survival in transplants from living - related donors . simultaneous analysis of 5,262 one haplotype - matched living - related allografts , and 973 hla identical allografts showed 10-year projected survival rates of 52% and 73% and graft half - lives of 11.9 and 23.6 years , respectively . conversely , the influence of hla matching on the survival of liver and thoracic organs is yet uncertain . to avoid hyperacute rejection , it is very important to identify recipient anti - hla antibodies to antigens expressed on donor with blood cells . the pioneer method to detect such antibodies , the complement - dependent cytotoxicity ( cdc ) , has been gradually replaced by more - sensitive solid - phase assays , such as the enzyme - linked immunosorbent assay and the bead - based technology ( i.e. , flow cytometry : flowpra and flow analyzer : luminex ) . however , the new techniques have been associated with decreased specificity , and some non - hla antigens with no clinical relevance have been able to give a positive crossmatch . these false - positive antibody results have as a consequence a decreased chance of the patient to receive an organ by way of exchange organizations , thus decreasing chances for the patient . thus , the experts recommend that the information these tests provide should complement that of the direct cdc assay . development of the field of organ and tissue transplantation has accelerated remarkably since the human major histocompatibility complex ( mhc ) was discovered in 1967 . however , has been elusive avoid the graft rejection . this is due to that the transplantation immunobiology is very complex , because of the involvement of several components such as antibodies , antigen presenting cells , helper and cytotoxic t cell subsets , immune cell , surface molecules , signaling mechanisms , and cytokines , which play a role in innate and adaptive immunities . Output:	the transplant of organs is one of the greatest therapeutic achievements of the twentieth century . in organ transplantation , the adaptive immunity is considered the main response exerted to the transplanted tissue , since the principal target of the immune response is the mhc ( major histocompatibility complex ) molecules expressed on the surface of donor cells . however , we should not forget that the innate and adaptive immunities are closely interrelated and should be viewed as complementary and cooperating . when a human transplant is performed , hla ( human leukocyte antigens ) molecules from a donor are recognized by the recipient 's immune system triggering an alloimmune response matching of donor and recipient for mhc antigens has been shown to have a significant positive effect on graft acceptance . this paper will present mhc , the innate and adaptive immunities , and clinical hla testing .
PubmedSumm2078	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: nonduplicate , imipenem - resistant isolates of 387 pseudomonas spp . and 267 acinetobacter spp . were collected from 2000 to 2001 from 28 hospitals in the korean nationwide surveillance of antimicrobial resistance group hospitals located in six cities or provinces . the identification of the species and the imipenem susceptibility were confirmed at the coordinating laboratory by using conventional tests ( 11 ) or atb 32 gn system ( biomerieux , marcy - letoile , france ) and by using the disk diffusion test ( 12 ) , respectively . mbl production was screened by using the hodge test and the imipenem - edta double disk synergy test ( 13 ) . the blaimp-1 and blavim-2 alleles were detected by polymerase chain reaction ( pcr ) , and three of the positive isolates were confirmed by sequencing , as described previously ( 4 ) . xbai - digested genomic dna of p. aeruginosa isolates was separated by pfge using the chef - dr - ii system ( bio - rad laboratories , hercules , ca ) ( 4 ) . the pattern was analyzed visually and by using uviband and map software ( uvitec ltd . , cambridge , uk ) . some of the pseudomonas and acinetobacter isolates collected were not fully resistant to imipenem but showed intermediate resistance when retested . among the isolates not susceptible to imipenem , 44 ( 11.4% ) of 387 pseudomonas spp . ( 42 p. aeruginosa and 2 p. putida ) and 38 ( 14.2% ) of 267 acinetobacter spp . were considered mbl producers on the basis of positive results by the hodge test and imipenem - edta double disk synergy test ( table 1 ) . mbl - producing pseudomonas spp . and acinetobacter spp . were detected in 11 ( 52.4% ) of 21 and 10 ( 41.7% ) of 24 hospitals that were located in four of five and five of six cities or provinces , respectively . we detected the blavim allele by pcr from all 42 isolates of mbl - producing p. aeruginosa and 2 isolates of p. putida . the blavim-2 and blaimp-1 alleles were detected in 27 ( 71.1% ) and 11 ( 28.9% ) of 38 acinetobacter isolates , respectively ( table 2 ) . nucleotide sequencing for three representative pcr - positive isolates confirmed the presence of the blavim-2 gene in one isolate each of p. aeruginosa and acinetobacter spp . , and the blaimp-1 gene in one isolate of acinetobacter spp . the mbl - producing strains were isolated mainly from intensive - care unit patients ( 31.7% ) and other inpatients ( 50.0% ) ; five ( 6.1% ) were from emergency service and other outpatients ( table 3 ) . overall , mbl - producing isolates were mainly obtained from specimens of sputum ( 50.0% ) and urine ( 29.3% ) . however , the proportion of mbl - producing isolates was relatively higher among urine isolates : 17.3% for pseudomonas spp . we obtained one mbl - producing acinetobacter isolate from each of the following specimen types : blood , spinal fluid , pleural fluid , and venous catheter tip ( table 4 ) . others included one acinetobacter isolate from specimens from blood , spinal fluid , pleural fluid , and a venous catheter tip . the pfge of the xbai - digested genomic dna of 39 isolates of p. aeruginosa showed 22 patterns ( data not shown ) . thirteen isolates ( 33.3% ) belonged to another identical pattern six from one hospital , two from each of two hospitals , and one from each of three hospitals , which were located in a city and two provinces . in this study , > 10% of all imipenem - nonsusceptible isolates of pseudomonas spp . and acinetobacter spp . were attributable to mbl production ( table 1 ) , and these mbl - producing isolates were detected in 62.5% of the participating hospitals . our finding indicates that mbl - producing p. aeruginosa is more prevalent in korea than in other countries ( 2 ) and that mbl - producing acinetobacter spp . the percentage of hospitals with mbl - producing isolates might have been higher if a larger number of imipenem - nonsusceptible isolates had been collected for this study . vim-2producing p. aeruginosa was isolated at almost the same time in europe ( 7 ) and korea ( 4 ) . however , imp-1producing isolates were rare until 2000 in korea . only one and three imp-1-positive p. aeruginosa and acinetobacter spp . , respectively , have been isolated at the reference laboratory ( 4 , unpub . data ) . in our study , 11 ( 28.9% ) of 38 mbl - positive isolates of acinetobacter spp . this increase suggests the possible introduction of imp - producing strains of acinetobacter spp . from japan , where 28 isolates of blaimp-1-positive acinetobacter baumannii were reported in a hospital as early as 1994 to 1996 ( 14 ) . rasmussen and bush ( 15 ) predicted that an increase of mbl - producing organisms was inevitable , given the more frequent use of carbapenems . imipenem has been used for only 9 years in korea , but the imipenem - resistance rate of p. aeruginosa has rapidly risen from 6% in 1996 to 19% in 2001 . a study by the korean nationwide surveillance of antimicrobial resistance group showed that the mean imipenem - resistance rates of p. aeruginosa in 1997 did not differ substantially depending on hospital size , ( i.e. , 17% in medium hospitals [ < 1,000 beds ] and 18% in large hospitals [ > 1,000 beds ] ) . the mean resistance rates to imipenem were not lower than those to ceftazidime in 2000 , i.e. , 21% versus 18% in large hospitals and 20% versus 19% in medium hospitals ( data not shown ) . the imipenem resistance rate of this organism isolated in korea was found to be much lower than that of p. aeruginosa , but its resistance rate rose from 4% in the first quarter to 20% in the third quarter of 2002 at the coordinating laboratory ( data not shown ) . in our study , mbl - producing pseudomonas spp . and acinetobacter spp . were isolated mainly from sputum and urine specimens , and most ( 81.7% ) isolates were from inpatients and intensive - care unit patients . therefore , proper treatment of respiratory secretions and urine from intensive - care unit patients is considered an important aspect of preventing the spread of mbl - producing organisms . the presence of p. aeruginosa isolates with identical pfge patterns among those collected not only from certain hospitals but also from different hospitals suggests that clonal spread is at least a part of the cause of intra- and inter - hospital dissemination of mbl - producing isolates . reported that five of seven patients infected with mbl - producing p. aeruginosa died , although the cause of death was difficult to establish with certainty ( 16 ) . clinical studies on the infection are rare because isolation of mbl - producing gram - negative bacilli increased only recently . we anticipate difficulties in treating patients infected with mbl - producing gram - negative bacilli , which can hydrolyze , in vitro , all available -lactams , except aztreonam for which clinical efficacy is unknown . our study indicates the urgent need for action to prevent further spread of mbl - producing organisms . previous experiences with penicillin - nonsusceptible pneumococci , methicillin - resistant staphylococcus aureus , vancomycin - resistant enterococcus faecium , and extended - spectrum -lactamase producing klebsiella pneumoniae indicate that once resistant bacteria can become widespread and can not be controlled ( 10 ) . although the national committee for clinical laboratory standards document ( 12 ) does not contain procedures for detection , simple procedures are available ( 13 ) . the prevalence of blavim-2 allele - positive p. aeruginosa and blaimp-1 allele - positive acinetobacter spp . is increasing possibly because of clonal and horizontal spread of the resistance determinant in korean hospitals . sputum and urine from inpatients and intensive - care unit patients were found to be the main sources of mbl - producing isolates . laboratories not only in korea but also in other countries with carbapenem - resistant organisms must be prepared to screen mbl - producing isolates to determine the clinical impact and prevent further spread of mbl - producing organisms . Output:	we determined the occurrence of acquired metallo--lactamase ( mbl)producing bacteria in korean hospitals . among the isolates nonsusceptible to imipenem that were collected from 28 hospitals from 2000 to 2001 , 44 ( 11.4% ) of 387 pseudomonas spp . and 38 ( 14.2% ) of 267 acinetobacter spp . infections produced mbl and had alleles of blavim-2 or blaimp-1 . mbl - producing isolates were detected in 60.7% of the hospitals .
PubmedSumm2079	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: restless legs syndrome ( rls ) is a common , distressing disease that affects a significant percentage of the adult population . in the us , 2% to 3% of the population experience clinically bothersome symptoms severe enough to warrant treatment.13 rls is characterized by discomfort of the legs during rest or inactivity and involves an urge to move the legs to relieve the symptoms . rls symptoms are usually present or worsen in the evening.4 because of the discomfort of the limbs during rest , rls can reduce sleep quality , daily function , and overall quality of life . several drugs are currently used to treat rls : dopaminergic agents such as levodopa ( in combination with carbidopa or benserazide ) , dopamine agonists ( such as ropinirole , and pramipexole ) , and nondopaminergic medications such as gabapentin , clonazepam and oxycodone.5,6 common side effects of these drugs are sedation , dizziness , fatigue , nausea , and vomiting . gabapentin is an anticonvulsant drug and is approved in the united states for the treatment of postherpetic neuralgia7 and partial seizures.710 gabapentin has also been shown effective in improving rls symptoms , reducing the frequency of periodic leg movements and improving sleep quality , suggesting a potential role in treating rls.1115 gabapentin is not approved for the indication of rls treatment , although it is often used off - label for this purpose.5,16,17 additional placebocontrolled , double - blind clinical trials have also suggested potential uses of gabapentin in the treatment of painful neuropathies in patients with diabetes mellitus18 and anxiety disorders.19 however , gabapentin has inherent pharmacokinetic deficiencies that limit its clinical effectiveness : variable bioavailability and short half - life . in healthy volunteers as well as epilepsy patients , clinical pharmacokinetic studies demonstrate limited absorption and high inter - patient variability.2023 bioavailability of orally administered gabapentin is dose - dependent . specifically , the oral bioavailability of gabapentin is approximately 60% , 47% , 34% , 33% , and 27% following 900 , 1200 , 2400 , 3600 , and 4800 mg / day given in 3 divided doses , respectively.24 importantly , the range of doses where bioavailability decreases coincides with doses reported to be useful to treat neuropathic pain.20 saturation of the gabapentin transport pathway may account for the observed dose - dependent decrease in bio - availability . absorption of gabapentin is mediated by a low - capacity solute transporter , probably an l - type amino acid transporter , which is located mainly in the upper small intestine.25,26 it has been suggested that the high inter - patient variability due to non - responders in clinical trials18,27 could be because of lower abundance of transporters in these patients,28 resulting in unpredictable plasma levels of gabapentin that may not reach the rapeutically meaningful levels . after oral administration and absorption , gabapentin is rapidly excreted via urine ; therefore gabapentin must be dosed 3 to 4 times a day to maintain therapeutic levels . the requirement of frequent administration can lead to noncompliance and missed doses which can reduce clinical effectiveness.29 enhancing the effectiveness of gabapentin has long been a goal , and pregabalin was developed in this direction . pregabalin is a structural analog of gabapentin , which does not bind to gamma - aminobutyric acid ( gaba ) receptors , and is approved as an add - on medication for epilepsy and neuropathic pain . in a small clinical trial , pregabalin was efficacious in treating primary rls as well as secondary rls , and further investigation with randomized , placebo - controlled trials was recommended.30 in an attempt to address the pharmacokinetic limitations of gabapentin , cundy and coworkers at xenoport inc . designed and synthesized a series of analogs of gabapentin and selected one candidate for further development based on its physico - chemical properties , enzymatic stability , transport , and in vivo pharmacokinetics . xp13512 is being further developed by xenoport inc / astellas pharma / glaxosmithkline . a new drug application ( nda ) has been filed with the food and drug administration ( fda ) and the current prescription drug user fee act ( pdufa ) goal date for its review is february 2010 . specifically , it is an acyloxyalkylcarbamate analog with an efficient enzymatic conversion to gabapentin in tissues . in addition , gabapentin enacarabil is designed to be recognized as a substrate for two high - capacity nutrient transporters with broad distribution through the human intestinal tract . one of the two transporters , monocarboxylate transporter type i ( mct-1 ) , regulates the absorption of small - chain fatty acids and may transport pharmaceutical agents across the intestine.31 the second transporter is a sodium - dependent multivitamin transporter ( smvt ) . the pharmacology was evaluated as immediate - release ( ir ) and extended release ( xr ) formulations.33,34 gabapentin enacarbil is rapidly absorbed and undergoes efficient enzymatic hydrolysis to yield gabapentin in vivo . in contrast to gabapentin , which uses lower capacity transport receptors located only in the upper intestinal area , gabapentin enacarbil uses alternative , higher - capacity transport receptors found across the whole length of the gastrointestinal tract . increased transport capacity allows the delivery of higher doses of gabapentin in good dose proportionality , avoiding uptake saturation at clinically relevant doses . no saturation of transport was observed in a dose escalation study up to 2800 mg using immediate release formulation ; the plasma concentration of gabapentin was observed to increase proportionally in accordance with an orally administered dose of gabapentin enacarbil.33 in contrast , gabapentin transport was saturated at 1400 mg oral dose levels . in the same study , similar observations were made with extended release tablet formulations of gabapentin enacarbil , with no sign of transport saturation up to 2100 mg oral dose levels . improved oral bioavailability for gabapentin enacarbil compared to gabapentin has been demonstrated in animal and human pharmacokinetic studies . cundy et al35 reported improved oral bioavailability , dose proportionality , and colonic absorption for the prodrug compared to gabapentin in rats and monkeys . compared with intracolonic gabapentin , intracolonic gabapentin enacarbil exhibited a 17-fold and 34-fold higher exposure in rats and monkeys , respectively . in addition , in monkeys , gabapentin enacarbil capsules resulted in 84.2% oral bioavailability compared with 25.4% bioavailability after a similar oral gabapentin dose . importantly , this improvement in bioavailability was also observed in a human study , specifically , it was reported that the oral bioavailability of gabapentin enacarbil compared to gabapentin was 74.5% vs 36.6%.33 based on the molecular weights and milligram - equivalent of gabapentin relative to the gabapentin enacarbil , a 365 mg dose of gabapentin as prodrug ( 700 mg ) achieved higher or comparable plasma concentrations of gabapentin compared to a 1200 to 1400 mg dose of free gabapentin in immediate release formulation.33 efficient oral absorption of gabapentin enacarbil and conversion to achieve higher and dose - dependent concentration of gabapentin in plasma will allow preparation of longer - acting extended release formulations . after intestinal absorption , gabapentin enacarbil is rapidly converted to gabapentin and the latter is the active form . although gabapentin is well established as an effective antiepileptic and antineuropathic pain drug , the underlying molecular mechanism of gabapentin is still not entirely clear.36 gabapentin is a structural analog of gaba ; however gabapentin does not interact significantly with any gaba subtype receptors,37,38 for instance , several studies reported no evidence that gabapentin binds substantially to any subtype of gabab receptors.37,39,40 even so , one study suggested that gabapentin acts as an agonist for a subset of gabab receptors,41 which may negatively regulate voltage - gated ca channels42 and activate inwardly rectifying k channels.43 gabapentin has shown a direct effect on the excitability of sensory neurons by blockade of ca and na channels or activation of k channels.44 lee and coworkers demonstrated that gabapentin upregulated renal outer medullary potassium channel , thus reducing neuronal excitability , suggestive of an important role in gabapentin s antiepileptic effect.45 d - serine , an agonist for n - methyl - d - aspartate ( nmda ) receptors , reverses some of gabapentin s actions.46 however , gabapentin does not directly interact with the glycine - nmda complex,37,47 and a rat study concluded that d - serine and gabapentin do not act at the same site.48 gabapentin also exhibits a profound synergistic anti - allodynic action with the ampa receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione ( cnqx).49 a recent study concluded that after peripheral nerve injury , gabapentin acts directly in the brainstem via a glutamate - dependent mechanism to stimulate descending inhibition to produce antihypersensitivity.50 suman - chauhan and coworkers identified a high - affinity binding site for gabapentin in rat brain homogenate.38 binding to this receptor was dependent on the system l transporter , indicating that gabapentin likely crosses the cell membrane and binding occurs intracellularly . there is no clear relationship between system l transporter binding and clinical efficacy of gabapentin.51 the binding site for gabapentin was subsequently identified as the 2 subunit of voltage - gated ca channel using pig cerebral cortex membranes.52 hendrich and coworkers provided a definitive proof that gabapentin acts primarily at an intracellular location.53 preclinical studies were conducted to establish the efficiency and conversion rate of gabapentin enacarbil to gabapentin in various tissues . gabapentin enacarbil was chemically stable at physiological ph range as indicated by > 95% recovery of the prodrug after incubation for 1 hour at 37c in buffers over the ph range of 2 to 8.28 however , the prodrug was rapidly converted to gabapentin by non - specific esterases in intestinal and liver tissues from rats , dogs , monkeys , and humans . rates of conversion to gabapentin in human intestinal and liver tissue preparations at 1 mg protein / ml were 196 and 146 pmol / min / mg protein . after 1 hour of incubation at 37c in tissue preparations of pancreatin , caco-2 cells , rat plasma , human plasma , rat liver , and human liver , 52% , 18% , 47% , 96% , 25% , and 4% of gabapentin enacarbil was recovered with release rates of gabapentin of 43% , 75% , 45% , 5% , 71% , and 81% , respectively.28 the conversion to gabapentin was quantitative in these studies and no formation of gabapentin lactam was observed . gabapentin enacarbil also did not show substrate or inhibitor properties for major p450 isoforms in human liver homogenates.28 therefore , gabapentin enacarbil is likely to have significantly low drug drug interaction following administration . to understand the mechanism of transport , in vitro transport studies across artificial lipid membranes were carried out . gabapentin enacarbil showed ph - dependent passive permeability across the membrane suggesting that the prodrug should have some ability to passively diffuse across cells.28 however , passive diffusion is likely a minor component because pka of gabapentin enacarbil is 5.0 . using in vitro cell culture assays , the transport of gabapentin enacarbil was shown to be dependent on mct-1 and smvt . to understand the biodistribution and oral bioavailability , pharmacokinetic studies were performed in rats and monkeys . a dose - proportional exposure to gabapentin was observed after oral administration of gabapentin enacarbil , while exposure to the intact prodrug was low . more than 95% of radio labeled [ c]-gabapentin enacarbil was excreted as gabapentin in urine in rats . in monkeys 84.2% of gabapentin enacarbil less than 1% of the intact prodrug was recovered in feces.53 in healthy human subjects , using supratherapeutic doses gabapentin enacarbil was converted rapidly to gabapentin after absorption . blood levels of gabapentin were proportional to gabapentin enacarbil dose over the range of 2400 to 6000 mg ( 12503125 mgequivalent gabapentin ) . blood concentrations of intact gabapentin enacarbil were low and transient ( 0.5% of the released gabapentin concentration at all doses).35 these studies indicate that gabapentin enacarbil is readily absorbed after oral delivery . in addition to one molecule of gabapentin , hydrolysis of gabapentin enacarbil also produces one molecule each of carbon dioxide , acetaldehyde , and isobutyric acid . both acetaldehyde and isobutyric acid are generally regarded as safe ( gras ) molecules by the fda . given that gabapentin enacarbil is converted into gabapentin , with a known safety profile , and also converted into gras compounds , it is likely that gabapentin enacarbil should exhibit a favorable safety profile . consistent with this hypothesis , no significant toxicities were observed with gabapentin enacarbil in monkeys following repeated oral administration of 2000 mg / kg / day doses . in addition no detectable accumulation was observed after 2 weeks.35 in a randomized - sequence , double - blind , placebo - controlled crossover study in healthy human subjects , gabapentin enacarbil was well tolerated up to supratherapeutic doses.54 in this study 600 mg extended - release tablets of gabapentin enacarbil were administered as a single oral dose of 2400 , 3600 , 4800 , or 6000 mg . in a 32 healthy volunteer study , the most commonly reported adverse events were dizziness and nausea ( 50% and 25% of subjects , respectively ) . two subjects experienced treatment - emergent adverse events rated as severe : psychomotor retardation , vertigo , and sedation ( 4800 mg dose ) and somnolence ( 6000 mg dose ) . these data support that gabapentin enacarbil is safe and well tolerated with minimal side effects . to study the bioavailability of gabapentin enacarbil from immediate release formulation , pharmacokinetic studies were conducted in healthy volunteers in phase i clinical trials.34,5557 single doses of ir gabapentin enacarbil ( 350 , 700 , 1400 , 2100 , and 2800 mg orally ; n = 40 ) or placebo ( n = 10 ) were evaluated . after 1 week , approximately equimolar doses of gabapentin ( 200 , 800 , 1200 , and 1400 mg ) were given to participants . gabapentin exposure with prodrug was dose - proportional ; bioavailability was 82.9% for the 350 mg dose and 79.7% for the 2800 mg dose . in contrast , with gabapentin parent the bioavailability was 65.2% and 26.5% with the lowest and the highest doses respectively . multiple doses of gabapentin enacarbil ( 350 , 400 , 800 , 1200 and 1400 mg orally ; twice daily , n = 29 ) , administered over 7 days , resulted in dose - proportional gabapentin . bioavailability from sustained release formulation of gabapentin enacarbil ( 1200 mg orally ) was evaluated either with ( n = 10 ) or without food ( n = 12 ) . gabapentin ( 600 mg orally ; n = 11 ) taken without food served as a reference . exposure to gabapentin from gabapentin enacarbil was higher in the with food group compared to the fasted group . bioavailability of gabapentin was 46.5% and 73.7% for the fasted and fed gabapentin enacarbil groups respectively , while the bioavailability was 37.7% in parent drug group.58 the effect of fat content in food on gabapentin enacarbil absorption was evaluated under fasted , low - fat ( 200300 kcal , 6% from fat ) , moderate - fat ( 500600 kcal , 30% from fat ) , and high - fat ( 1000 kcal , 50% from fat ) conditions . the bioavailability of gabapentin was 42% , 64% , 65% , and 76% in fasted , low- , medium- , and high - fat food conditions respectively.34 potential for drug interactions between gabapentin enacarbil , naproxen ( an mct-1 substrate ) and cimetidine ( an organic cation transporter 2 substrate ) was evaluated to assess the alterations in drug absorption when the prodrug is used in combination with these and other transporter substrates . in healthy volunteers , gabapentin enacarbil did not affect the bioavailability of naproxen or cimetidine . the auc values of gabapentin enacarbil , when co - administered in combination with naproxen or cimetidine , increased up to 12% and 24% , respectively . the results indicate that gabapentin enacarbil can be used together with mct-1 , organic cation transporter 2 , or other transporter substrates without the need to alter the dose regimes.59,60 because various phase i , ii and iii trials were recently summarized by merlino et al 2009,34 this review will focus on key phase ii and iii studies in conjunction with drug efficacy . readers are encouraged to refer the review by merlino et al for a summary of clinical trials prior to the publication of this article . efficacy of gabapentin enacarbil in relieving rls symptoms was evaluated over a two week period in a multicenter , randomized , double - blind , placebo - controlled , crossover , polysomnographic phase iia clinical trial in rls patients . patients were treated for two weeks with either gabapentin enacarbil ( 600 mg orally at 5 pm and 1200 mg orally 1 hour before bed ) or placebo , with a 1-week washout period between treatments . at the end of the treatment period , a significant improvement was observed in rls symptoms such as quality of sleep , number of awakenings per night and hours awake per night . compared to 14.7% patients reporting improvement of rls symptoms in placebo group , 79.5% of patients in the gabapentin enacarbil group reported either much improved or very much improved symptoms.61 efficacy and tolerability of 1800 mg dose of gabapentin enacarbil was evaluated in 38 nave subjects with rls , over 9 clinical sites.62 patients were treated with 1800 mg / day for 14 days , with a 7-day washout period between treatment periods . after 14 days , significantly reduced rls symptoms and improved sleep were observed in subjects with moderate - to - severe primary rls while dose levels were well tolerated . in a 12-week , multicenter , randomized , double - blind , placebo controlled phase iii clinical trial , efficacy of gabapentin enacarbil ( 1200 mg / day orally at 5 pm ) was evaluated in patients with primary rls . patients treated with gabapentin enacarbil showed an improved international rls ( irls ) total score ( 13.2 ) in contrast to placebo group ( 8.8 ) at 2 weeks . on the investigator - rated cgi ( clinical global impression ) , significantly more gabapentin enacarbil - treated patients ( 76.1% ) responded than placebo ( 38.9% ) . at the end of study more than 50% of gabapentin enacarbil - treated patients showed no sign of rls over the 24-hour assessment period compared with 18% placebo patients.63,64 long - term safety and efficacy of gabapentin enacarbil was evaluated in a second multicenter , placebo - controlled clinical trial involving 327 patients with primary moderate - to - severe rls symptoms.65 patients received gabapentin enacarbil ( 1200 mg / day orally ) at 5 pm for 24 weeks in a single - blind phase of trial . the initial 24 weeks phase was completed with 221 patients , out of which 194 ( 88% ) responded either much improved or very much improved cgi scores . these patients were further entered into a 12- week randomized , double - blind phase . patients were given gabapentin enacarbil for either 12 weeks ( 1200 mg / day po ) or for 2 weeks ( 600 mg / day po ) and then placebo for 10 weeks . for the extended trial , the primary end - point was number of patients who relapsed or had worsening symptoms . gabapentin enacarbil treated group showed a statistically significant lower proportion of relapses than placebo , 9% versus 23% respectively . a third phase iii trial evaluated the safety and efficacy of 600 and 1200 mg / day over 12 weeks.66 both dose groups of gabapentin enacarbil resulted in significantly improved irls and cgi scores than with placebo . in addition to rls , other indications are being investigated for gabapentin enacarbil.67 the additional indications include post - herpetic neuralgia , painful diabetic neuropathy , and migraine prophylaxis . recently , top - line results were presented from a phase ii clinical study of placebo , 1200 , 2400 or 3600 mg / day of gabapentin enacarbil dosed twice a day for treatment of post - herpetic neuralgia . all doses demonstrated statistically significant improvements over placebo on the change from baseline to the end of maintenance treatment in the 24- hour average pain intensity score . this 14-week , double - blind , placebo - controlled study enrolled 376 subjects experiencing pain for at least 3 months following healing of herpes zoster skin rash . gabapentin enacarbil was generally well tolerated at all doses , and the most common adverse events were dizziness and somnolence , and most were mild or moderate . rls is a sleep - related movement disorder which involves an undesirable sensation to move the legs , which improves with activity . long - term use of dopaminergic drugs may cause augmentation or other side effects such as leg edema , dizziness , and nausea . gabapentin is only mildly effective in relieving rls symptoms , perhaps because of poor absorption from the gastrointestinal tract . gabapentin enacarbil , a prodrug of gabapentin , is designed to enhance gastrointestinal absorption and hence enhance bioavailability of gabapentin . clinical trials have shown gabapentin enacarbil to be a safe and effective drug for rls . an application for approval of gabapentin enacarbil for treatment of rls is currently pending with fda for approval , and additional clinical indications are being pursued , including neuropathic pain . the addition of one more therapeutic option to the rls drug armamentarium would provide more options to physicians dealing with rls patients with refractory symptoms . Output:	restless legs syndrome ( rls ) is a sleep - related movement disorder commonly involving an unpleasant urge to move the limbs , typically the legs . dopaminergic agents represent the first - line therapy for rls ; however , long - term use of such drugs results in worsening symptoms due to augmentation or other adverse events . gabapentin , an analog of the inhibitory neurotransmitter gamma - aminobutyric acid ( gaba ) , is an anticonvulsant / analgesic agent . gabapentin is only mildly effective in relieving rls symptoms , perhaps a result of its poor absorption from the gastrointestinal ( gi ) tract . gabapentin enacarbil is a prodrug of gabapentin specifically designed to enhance absorption via the gi tract , and hence provide improved circulating levels of gabapentin on metabolism . clinical trials to date have demonstrated favorable safety and ( compared to traditional gabapentin ) improved pharmacokinetics and efficacy in treating rls symptoms . thus , gabapentin enacarbil may prove to be a useful drug in treating rls . an application of gabapentin enacarbil for treatment of rls is currently pending with fda for approval .
PubmedSumm2080	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: xeroderma pigmentosum ( xp ) is a rare autosomal recessive disorder characterized by cutaneous photosensitivity , pigmentary changes , photophobia and propensity to develop early malignancy in sun - exposed skin and ocular structures . the malignancies described commonly in patients with xp include squamous cell carcinoma ( scc ) , basal cell carcinoma ( bcc ) , and malignant melanoma ( mm ) . other types of cutaneous neoplasm including angiosarcoma and fibrosarcoma have also been reported . atypical fibroxanthoma ( afx ) is a pleomorphic spindle cell neoplasm of the dermis that occurs in actinically damaged skin of elderly individuals . however , cases also have been reported occurring in the limbs and trunks of younger persons . histologically , it mimics malignant fibrous histiocytoma ( mfh ) , but the tumor usually follows an indolent or locally aggressive course . only a small number of metastatic cases have been reported . afx has been regarded as the neoplasm of low - grade malignancy related to mfh . a 20-year - old man , farmer by occupation , presented with two tumors over his cheek . on examination , the patient showed pigmentation and depigmentation all over his body [ figure 1 ] which , according to him , were present since childhood . on further enquiry , he gave a history of six operations on his face over the last 4 years . on three occasions , the lesions were diagnosed as baso - squamous carcinoma , once as scc , and on two occasions he developed bcc . he had five siblings out of which two were affected by the same disease process and died of metastasizing scc . the other three , all younger to him , were phenotypically normal with no manifestation of the disease so far . during the present visit , two tumors were excised from his cheek . on gross examination , one of the tumors measured 3.5 2 2 cm and showed ulceration of the skin surface . sections showed a relatively well - circumscribed mass with tumor cells occupying papillary and reticular dermis . the tumor cells comprised polygonal- to spindle - shaped cells with bizarre and multinucleated tumor giant cells showing marked pleomorphism [ figure 3 ] . mitotic figures were present but not brisk . with the differential of a poorly differentiated epithelial neoplasm and a soft tissue tumor in mind , immunohistochemistry was performed and the cells showed strong positivity for vimentin [ figure 4 ] , and were negative for pancytokeratin , s-100 and hmb45 . the tumor cells also showed positivity for cd68 [ figure 5 ] , suggesting a histiocytic differentiation . taking into account the gross , histomorphologic , and immunohistochemical features , a diagnosis of afx was made . in the 8-month follow - up period , the patient did not develop any metastases . mottled pigmented and depigmented areas of facial skin along with scars of previous surgeries photomicrograph showing a well - circumscribed dermal neoplasm with intact epidermis ( hematoxylin and eosin , 40 ) photomicrograph showing highly pleomorphic spindle - shaped cells with bizarre giant cells ( h and e , 400 ) photomicrograph showing tumor cells expressing vimentin ( immunohistochemistry for vimentin , 400 ) photomicrograph showing tumor cells expressing cd68 ( immunohistochemistry for cd68 , 400 ) xp is an autosomal recessive disorder characterized by defective deoxyribonucleic acid ( dna ) repair . the skin cells from patients with xp have an impaired ability to repair ultraviolet ( uv)-induced dna damage . defective dna repair is also reflected in dermal fibroblasts , lymphocytes of peripheral blood , and conjunctival cells . xp being an autosomal recessive disease , the heterozygous parents are normal and both the sexes are equally affected . in the present case , out of five siblings of the patient , two were affected by the same disease process and succumbed to repeated development of scc . malignant skin neoplasms have been reported in 45 - 60% of xp patients in different studies . robbins has shown 10 - 20 times higher incidence of internal neoplasm in xp patients compared to normal controls . . only five cases of fibrosarcomas , three angiomas , two afx , two mfh , and one angiosarcoma have been demonstrated so far.[131012 ] the histogenesis of afx is controversial . however , electron microscopic and immunohistochemical studies indicate that as for mfh , the progenitor cell is an undifferentiated mesenchymal cell , capable of showing histiocytic , fibroblastic , and myofibroblastic differentiation . the concept that afx is a form of pleomorphic mfh has received further support from a study in which no significant difference between afx and mfh was demonstrated in their apoptotic behavior , proliferation indices , p53 protein expression and presence of the bcl2 product . despite marked cytological atypia , dilek et al . have found few mitotic figures in afx . however , other studies have shown numerous mitotic figures including abnormal forms . despite its apparently malignant histologic features , the tumor usually follows an indolent or locally aggressive course . due to the small number of metastatic cases reported , afx has been regarded as the neoplasm of low - grade malignancy related to mfh from which it is indistinguishable histologically . according to this view , the more favorable prognosis of afx is related to its superficial location and small size . the data regarding the outcome of afx are mostly available from the cases not associated with xp . the outcome of the cases of afx in patients with xp has not been discussed in literature . the differential diagnosis in the present case , occurring in a patient of xp , included a poorly differentiated epithelial neoplasm . immunohistochemistry for cytokeratin , s100 , vimentin , and cd68 helped confirmation of the diagnosis . dilek et al . have recommended cytokeratin , s100 , hmb45 , and histiocytic markers in difficult cases . cases of afx are usually associated with severe actinic damage and few have arisen in areas treated by radiation . deficient repair of uv - induced dna damage is directly involved in the development of cancer in xp . Output:	xeroderma pigmentosum ( xp ) is a rare autosomal recessive disorder related to defective deoxyribonucleic acid ( dna ) repair . various cutaneous manifestations related to ultraviolet ( uv ) damage characterize the clinical course . primary malignant cutaneous neoplasms like squamous cell carcinoma , basal cell carcinoma and malignant melanoma have been reported . atypical fibroxanthoma is a rare dermal neoplasm occurring in uv - damaged skin . we report an unusual case of atypical fibroxanthoma in a 20-year - old male with xp .
PubmedSumm2081	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: nitrogenase is a complex metalloenzyme that catalyzes the nucleotide - dependent reduction of atmospheric n2 to bioavailable nh3 . the best - studied mo - dependent nitrogenase is a binary system consisting of the fe protein and the fe mo ( mofe ) protein . the fe protein is an 2 homodimer containing one [ fe4s4 ] cluster at the subunit interface and one mgatp binding site within each subunit , whereas the mofe protein is an 22 heterotetramer containing one p - cluster ( [ fe8s7 ] ) at each /-subunit interface and one fe mo cofactor ( femoco ) ( [ mofe7s9x - homocitrate ] , where x is c , n , or o ) within each subunit . during catalysis , the fe protein serves as an obligate electron donor for the mofe protein and , in an atp - dependent process , mediates the transfer of electrons from its [ fe4s4 ] cluster , through the p - cluster , to the femoco of the mofe protein , where substrate reduction eventually occurs [ 1 , 4 ] . biosynthesis of femoco has remained a central topic in the field of bioinorganic chemistry because of its complexity and intricacy . this process is presumably initiated by the mobilization of fe and s by nifu ( encoded by nifu ) and nifs ( encoded by nifs ) for the assembly of small fe / s fragments , which are subsequently used for the generation of an fe / s core on nifb ( encoded by nifb ) . such an fe / s core , which likely contains all fe and s required for the formation of the mature cofactor , is then transferred to nifen ( encoded by nife and nifn ) , where it is further matured before being delivered to its binding site in the mofe protein . the hypothesis that nifen is a scaffold protein for femoco assembly is largely based on the significant sequence homology between nifen and mofe protein . comparison between the primary sequences of these two proteins has led to the proposal that nifen contains a p - cluster site that houses a p - cluster homolog and a femoco site that hosts the conversion of femoco precursor to a mature cofactor . although the p - cluster homolog was identified as an [ fe4s4]-type cluster earlier , the femoco precursor was identified only recently [ 8 , 9 ] following the biochemical / spectroscopic characterization of two forms of nifen from azotobacter vinelandii : one , designated nifb nifen , contains an [ fe4s4 ] cluster at the ; the other , designated nifen , contains an [ fe4s4 ] cluster at the p - cluster site and a precursor form of femoco at the femoco site ( fig . 1 ) . subtractive x - ray absorption spectroscopy ( xas)/extended x - ray absorption fine structure ( exafs ) analysis revealed that the precursor on nifen is a mo / homocitrate - free cluster that closely resembles the fe / s core of femoco . furthermore , combined biochemical and xas / exafs investigations showed that , in an atp- and reductant - dependent process , fe protein can insert mo and homocitrate into the nifen - bound precursor , leading to the formation of a third form of nifen ( designated nifen ) , which contains a fully complemented . 1the 22-heterotetrameric nifb nifen ( a ) , nifen ( b ) , nifen ( c ) , and mofe protein ( d ) . the permanent [ fe4s4 ] clusters of nifen at the -subunit interfaces are indicated by red cubes . compared with nifb nifen , nifen , and nifen contain additional clusters within the subunits ( i.e. , precursor and femoco , respectively ) , which are indicated by yellow ovals . the mofe protein contains one [ fe8s7 ] p - cluster at each -subunit interface ( gray ovals ) and one femoco within each subunit ( yellow oval ) the 22-heterotetrameric nifb nifen ( a ) , nifen ( b ) , nifen ( c ) , and mofe protein ( d ) . the permanent [ fe4s4 ] clusters of nifen at the -subunit interfaces are indicated by red cubes . compared with nifb nifen , nifen , and nifen contain additional clusters within the subunits ( i.e. , precursor and femoco , respectively ) , which are indicated by yellow ovals . the mofe protein contains one [ fe8s7 ] p - cluster at each -subunit interface ( gray ovals ) and one femoco within each subunit ( yellow oval ) epr spectroscopy has yielded information regarding the spin states of the different clusters of the three nifen species , which complements the structural information generated by xas / exafs analysis . however , neither technique has revealed information necessary for the complete characterization of the electronic structures of the clusters . to elucidate the electronic structures of the nifen - associated clusters , we performed a variable - temperature , variable - field magnetic circular dichroism ( mcd ) investigation of the three forms of nifen in the dithionite - reduced state . the results reported herein provide the electronic information of the nifen - associated clusters that is necessary for a better understanding of the maturation process of femoco . unless noted otherwise , all chemicals and reagents were obtained from fisher scientific or sigma aldrich . wild - type ( avop ) and variant ( dj1041 and ym9a ) strains of a. vinelandii were used in this study . dj1041 expresses a his - tagged nifen ( designated nifen ) in the nifhdkty - deletion background , and ym9a expresses a his - tagged nifen ( designated nifb nifen ) in the nifhdkty / nifb - deletion background [ 7 , 8 ] . all a. vinelandii strains were grown in 180-l batches in a 200-l new brunswick fermentor ( new brunswick scientific , edison , nj , usa ) in burke s minimal medium supplemented with 2 mm ammonium acetate . the growth rate was measured by cell density at 436 nm using a spectronic 20 genesys ( spectronic instruments , westbury , ny , usa ) . after the consumption of ammonia , the cells were derepressed for 3 h , followed by harvesting using a flow - through centrifugal harvester ( cepa , lahr , germany ) . the cell paste was washed with 50 mm tris(hydroxymethyl)aminomethane ( tris)hcl ( ph 8.0 ) . published methods were used for the purification of wild - type a. vinelandii fe protein and his - tagged nifen . three forms of nifen were used in the comparative mcd studies : ( 1 ) nifb nifen , which contains one [ fe4s4 ] cluster at each p - cluster site of nifen ; ( 2 ) nifen , which contains , in addition to the [ fe4s4 ] clusters at the p - cluster sites , a femoco precursor at the femoco site of nifen ; and ( 3 ) nifen , which contains the [ fe4s4 ] clusters at the p - cluster sites and a fully complemented femoco at the femoco site of nifen . nifen was obtained by incubating nifen with fe protein , mgatp , dithionite , sodium molybdate ( na2moo4 ) , and homocitrate , and reisolating nifen following such treatment [ 10 , 11 ] . all mcd samples were prepared in an ar - filled drybox ( vacuum atmospheres , hawthorne , ca , usa ) at an oxygen level of less than 4 ppm . dithionite - reduced protein samples were in 25 mm tris hcl ( ph 8.0 ) , 10% glycerol , and 2 mm dithionite ( na2s2o4 ) . samples were subsequently concentrated in a centricon-50 concentrator ( amicon ) in anaerobic centrifuge tubes outside the drybox as described earlier . after concentration , these protein samples [ 50100 mg ml in 25 mm tris hcl ( ph 8.0 ) and 50% glycerol ] were transferred to mcd sample cuvettes under anaerobic conditions and frozen in liquid n2 . mcd sample cells were constructed from optical - quality spectrosil quartz ( 1702,200 nm , 1-mm path length , model bs-1-q-1 , starna , model suv r-1001 or fuv ; spectrocell , oreland , pa , usa ) . each cuvette was cut into the appropriate dimensions to fit the sample holder ( 2.0 cm 12.5 mm ) , resulting in a sample volume of approximately 160 l . all samples contained 50% glycerol to ensure the formation of an optical glass upon freezing and were kept on dry ice in transit . mcd spectra were recorded with a circular dichroism ( cd ) spectropolarimeter ( model j-710 ; jasco , easton , md , usa ) interfaced with a superconducting magnet ( model spectromag 400 - 7 t ; oxford instruments , oxford , uk ) as previously described . sample temperatures were monitored with a thin film resistance temperature sensor ( model cx1050-cu-1 - 4l ; lakeshore , westerville , oh , usa ) positioned directly ( 1 mm ) above the sample cuvette . the linearity of the magnetic field was monitored with a calibrated hall generator ( model hgca-3020 ; lakeshore , westerville , oh , usa ) placed directly outside the superconducting magnet . mcd spectra were recorded at a rate of 50 nm min from 800 to 350 nm at a resolution of 210 nm . since optical glasses formed at low temperature often generate a strain - induced background cd spectrum , the cd spectrum was recorded in a zero magnetic field to determine whether the background signal was excessive ( if it was excessive , the sample was replaced with a new sample ) . to further eliminate interference by this signal , the corrected mcd spectrum was obtained for each sample by first recording the spectrum with the magnetic field in the normal direction and subtracting from it the spectrum with the field in the reversed direction . all spectral intensities were quantified per heterodimer and corrected for path length , sample concentration , and depolarization effects . the extent of depolarization was determined by placing a standard sample of nickel tartarate between the magnet cryostat and the detector . the cd spectrum was then recorded before and after light had passed through the frozen protein sample in the magnet . the quantitative results of these analyses are specified in the text for each protein studied . magnetization curves were recorded at a set wavelength and temperature while the magnetic field was linearly varied from 0 to 6 t at a rate of 0.45 t min with a resolution of 2 s. mcd data were analyzed using a fit / simulation program created by neese and solomon . the program allows the calculation of best - fit saturation magnetization curves using experimental data as a basis set and is valid for any spin state , half - integer or integer , at any specified temperature . experimental data were analyzed by fitting the spin hamiltonian parameters and the effective transition moment products . the effective transition moment products represent the planes of polarization that reflect the anisotropy of the g factors . since the initial slope of the magnetization curve is dependent on the g factors , the transition polarizations relate the transition dipole to the g - factor axes of a powder or randomly oriented sample . for s > spin systems , the spin parameters , including the g factor ( g ) , the axial zero - field splitting ( d ) and the rhombic distortion of the electronic environment ( e / d ) , are determined on the basis of the hamiltonian1\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$ \hat{h } = \beta bg\hat{s } + d\left [ { \hat{s}_{z}^{2 } - \ , \left ( { 1/3 } \right)\hat{s}\left ( { \hat{s } + 1 } \right ) + \left ( { e / d } \right)\left ( { \hat{s}_{x}^{2 } -\hat{s}_{y}^{2 } } \right ) } \right ] $ $ \end{document}which is the expression for the energy of the zeeman interaction and the correction to the energy of the individual spin states arising from spin orbit coupling . at low temperatures ( approximately 1.6 k ) , the lowest energy level is predominantly populated and dictates the behavior of the magnetization curve . as the temperature is raised , the spectral parameters of excited states become increasingly important in the profile of the magnetization curve . best - fit simulations of the experimental data were initially performed at the lowest temperature to enable the determination of the spectral parameters . subsequent simulations of high - temperature data facilitated the determination of the axial zero - field splitting , d. wild - type ( avop ) and variant ( dj1041 and ym9a ) strains of a. vinelandii were used in this study . dj1041 expresses a his - tagged nifen ( designated nifen ) in the nifhdkty - deletion background , and ym9a expresses a his - tagged nifen ( designated nifb nifen ) in the nifhdkty / nifb - deletion background [ 7 , 8 ] . all a. vinelandii strains were grown in 180-l batches in a 200-l new brunswick fermentor ( new brunswick scientific , edison , nj , usa ) in burke s minimal medium supplemented with 2 mm ammonium acetate . the growth rate was measured by cell density at 436 nm using a spectronic 20 genesys ( spectronic instruments , westbury , ny , usa ) . after the consumption of ammonia , the cells were derepressed for 3 h , followed by harvesting using a flow - through centrifugal harvester ( cepa , lahr , germany ) . the cell paste was washed with 50 mm tris(hydroxymethyl)aminomethane ( tris)hcl ( ph 8.0 ) . published methods were used for the purification of wild - type a. vinelandii fe protein and his - tagged nifen . three forms of nifen were used in the comparative mcd studies : ( 1 ) nifb nifen , which contains one [ fe4s4 ] cluster at each p - cluster site of nifen ; ( 2 ) nifen , which contains , in addition to the [ fe4s4 ] clusters at the p - cluster sites , a femoco precursor at the femoco site of nifen ; and ( 3 ) nifen , which contains the [ fe4s4 ] clusters at the p - cluster sites and a fully complemented femoco at the femoco site of nifen . nifen was obtained by incubating nifen with fe protein , mgatp , dithionite , sodium molybdate ( na2moo4 ) , and homocitrate , and reisolating nifen following such treatment [ 10 , 11 ] . all mcd samples were prepared in an ar - filled drybox ( vacuum atmospheres , hawthorne , ca , usa ) at an oxygen level of less than 4 ppm . hcl ( ph 8.0 ) , 10% glycerol , and 2 mm dithionite ( na2s2o4 ) . samples were subsequently concentrated in a centricon-50 concentrator ( amicon ) in anaerobic centrifuge tubes outside the drybox as described earlier . after concentration , these protein samples [ 50100 mg ml in 25 mm tris hcl ( ph 8.0 ) and 50% glycerol ] were transferred to mcd sample cuvettes under anaerobic conditions and frozen in liquid n2 . mcd sample cells were constructed from optical - quality spectrosil quartz ( 1702,200 nm , 1-mm path length , model bs-1-q-1 , starna , model suv r-1001 or fuv ; spectrocell , oreland , pa , usa ) . each cuvette was cut into the appropriate dimensions to fit the sample holder ( 2.0 cm 12.5 mm ) , resulting in a sample volume of approximately 160 l . all samples contained 50% glycerol to ensure the formation of an optical glass upon freezing and were kept on dry ice in transit . mcd spectra were recorded with a circular dichroism ( cd ) spectropolarimeter ( model j-710 ; jasco , easton , md , usa ) interfaced with a superconducting magnet ( model spectromag 400 - 7 t ; oxford instruments , oxford , uk ) as previously described . sample temperatures were monitored with a thin film resistance temperature sensor ( model cx1050-cu-1 - 4l ; lakeshore , westerville , oh , usa ) positioned directly ( 1 mm ) above the sample cuvette . the linearity of the magnetic field was monitored with a calibrated hall generator ( model hgca-3020 ; lakeshore , westerville , oh , usa ) placed directly outside the superconducting magnet . mcd spectra were recorded at a rate of 50 nm min from 800 to 350 nm at a resolution of 210 nm . since optical glasses formed at low temperature often generate a strain - induced background cd spectrum , the cd spectrum was recorded in a zero magnetic field to determine whether the background signal was excessive ( if it was excessive , the sample was replaced with a new sample ) . to further eliminate interference by this signal , the corrected mcd spectrum was obtained for each sample by first recording the spectrum with the magnetic field in the normal direction and subtracting from it the spectrum with the field in the reversed direction . all spectral intensities were quantified per heterodimer and corrected for path length , sample concentration , and depolarization effects . the extent of depolarization was determined by placing a standard sample of nickel tartarate between the magnet cryostat and the detector . the cd spectrum was then recorded before and after light had passed through the frozen protein sample in the magnet . the quantitative results of these analyses are specified in the text for each protein studied . magnetization curves were recorded at a set wavelength and temperature while the magnetic field was linearly varied from 0 to 6 t at a rate of 0.45 t min with a resolution of 2 s. mcd data were analyzed using a fit / simulation program created by neese and solomon . the program allows the calculation of best - fit saturation magnetization curves using experimental data as a basis set and is valid for any spin state , half - integer or integer , at any specified temperature . experimental data were analyzed by fitting the spin hamiltonian parameters and the effective transition moment products . the effective transition moment products represent the planes of polarization that reflect the anisotropy of the g factors . since the initial slope of the magnetization curve is dependent on the g factors , the transition polarizations relate the transition dipole to the g - factor axes of a powder or randomly oriented sample . for s > spin systems , the spin parameters , including the g factor ( g ) , the axial zero - field splitting ( d ) and the rhombic distortion of the electronic environment ( e / d ) , are determined on the basis of the hamiltonian1\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$ \hat{h } = \beta bg\hat{s } + d\left [ { \hat{s}_{z}^{2 } - \ , \left ( { 1/3 } \right)\hat{s}\left ( { \hat{s } + 1 } \right ) + \left ( { e / d } \right)\left ( { \hat{s}_{x}^{2 } -\hat{s}_{y}^{2 } } \right ) } \right ] $ $ \end{document}which is the expression for the energy of the zeeman interaction and the correction to the energy of the individual spin states arising from spin orbit coupling . at low temperatures ( approximately 1.6 k ) , the lowest energy level is predominantly populated and dictates the behavior of the magnetization curve . as the temperature is raised , the spectral parameters of excited states become increasingly important in the profile of the magnetization curve . best - fit simulations of the experimental data were initially performed at the lowest temperature to enable the determination of the spectral parameters . subsequent simulations of high - temperature data facilitated the determination of the axial zero - field splitting , d. the precursor - free nifb nifen has a metal content of eight fe per 22 tetramer . as shown in the past , the epr spectrum of this protein shows a single g 2 signal of an s = spin system at a concentration of approximately two spins per tetramer . high - resolution mcd spectra of the nifb nifen at high field ( 6 t ) and various temperatures ( 1.57 , 4.20 , and 10.5 k ) are shown in fig . 2 . these spectra exhibit two broad positive inflections at 520 and 764 nm , respectively , and a broad negative inflection at 660 nm with a shoulder at 610 nm . this pattern of inflections is characteristic of [ fe4s4 ] clusters , which exhibit broad positive inflections in the 520530- and 740780-nm regions and broad negative inflections in the 620- and 660-nm regions . the 520530-nm region is associated with the s fe charge - transfer region [ 18 , 19 ] , whereas the 740780-nm region has been attributed to a valence - delocalized intervalence band . the intensity at 520 nm ( 75 m cm per dimer ) is well within the average range of intensity ( 6090 m cm ) for a single [ fe4s4 ] cluster [ 21 , 22 ] . all of the analyses are consistent with reduced nifb nifen containing two permanent [ fe4s4]-like clusters ( one cluster per dimer ) [ 7 , 8].fig . 2magnetic circular dichroism ( mcd ) spectra of nifb nifen at 6 t and 1.57 , 4.20 , and 10.5 k magnetic circular dichroism ( mcd ) spectra of nifb nifen at 6 t and 1.57 , 4.20 , and 10.5 k in addition to the classic broad mcd inflections of the [ fe4s4 ] clusters , four sharp positive inflections at 360 , 396 , 430 , and 458 nm can also be observed in the spectra of nifb nifen . 2 ) parallels that of the broad inflections , implying that all of them arise from the same cluster in the protein . unlike the broad long - wavelength features , the sharp short - wavelength inflections are not characteristic of all [ fe4s4 ] clusters and , therefore , represent an identifying fingerprint of the specific permanent [ fe4s4]-like clusters in nifb nifen . the [ fe4s4]-like clusters in nifb nifen were further characterized by examining the magnetization of this protein at various temperatures ( 1.57 , 4.20 , and 10.5 k ) ( fig . the epr spectrum of nifb nifen exhibits a dominant s = signal in the g 2 region . it is obvious that the experimental magnetization curves do not mimic the simulated curve for the pure s = system . first , the 1.57 k magnetization curve does not approach a horizontal ( i.e. , saturation ) limit at high magnetic field . this observation suggests the presence of a small b term ( i.e. , diamagnetic ) and/or high - spin contribution to the spectral intensity . a b - term contribution is typically temperature - independent with a near - linear increase in intensity upon an increase in field . nesting is the flaring of the magnetization curves as the temperature increases and often occurs in higher - spin states ( i.e. , s > ) , where low - lying , excited - levels ( with different magnetization parameters ) become populated as the temperature increases ( eq . 1 ) . nesting is not observed in the case of a pure s = system , where there are no low - lying excited states . both the mcd spectral pattern and the magnetization data are similar to those observed in another nifen species that is practically free of precursor .fig . 3magnetization curves of nifb nifen at 1.57 , 4.20 , and 10.5 k ( black ) recorded at 520 nm and an example of a possible simulation ( red ) of the magnetization curves assuming a mixture of an s = spin system ( 65% ) with g [ 2.07 , 1.91 , 1.84 ] and an s = 3/2 spin system ( 30% ) with e / d = 0.052 and d = 5.0 cm ( polarizations used in simulations mxy = 1.0 , mxz = 1.0 , myz = 1.0 ) and a 5% linear b term magnetization curves of nifb nifen at 1.57 , 4.20 , and 10.5 k ( black ) recorded at 520 nm and an example of a possible simulation ( red ) of the magnetization curves assuming a mixture of an s = spin system ( 65% ) with g [ 2.07 , 1.91 , 1.84 ] and an s = 3/2 spin system ( 30% ) with e / d = 0.052 and d = 5.0 cm ( polarizations used in simulations mxy = 1.0 , mxz = 1.0 , myz = 1.0 ) and a 5% linear b term the magnetization data of nifb nifen , therefore , suggest the presence of an additional high - spin state in this protein . the existence of mixed spin states is not unusual for the [ fe4s4 ] clusters . the most common high - spin states mixing with the s = state are s = 3/2 and s = 7/2 , with the s = 3/2 state being the more common one . because no signals of high - spin states can be observed in the epr spectrum of nifb nifen , no spectral parameters of any high - spin state ( e.g. , s , d , e / d , or the relative percentage of a high - spin state ) can be obtained . as such , it is extremely difficult to simulate the data and determine the exact origin of the nesting in fig . 3 or the percentage contribution of any high - spin state . only a range of acceptable parameters can be obtained . assuming an additional contribution from only an s = 3/2 state , we could construct acceptable simulations ( not shown ) with the range of parameters as follows : 1030% s = 3/2 , d = 4.5 2.5 cm , e / d = 0.10 0.10 , and 5% linear , temperature - independent b term , where the remaining 6585% comes from the s = state . another possible explanation for the nesting of the magnetization curves is the presence of a low - lying s = 3/2 spin state that becomes thermally populated at elevated temperatures . this explanation is consistent with the epr spectrum , where the s = signal has high concentration ( approximately two spins per protein ) and no high - spin signals are observed at low temperatures . when the mcd spectra of reduced nifb nifen , nifen , and nifen are compared , it is apparent that they differ from one another ( fig . 4 ) . this observation is not surprising , as nifen and nifen contain paramagnetic clusters ( i.e. , precursor and femoco , respectively ) [ 9 , 12 ] in addition to the permanent [ fe4s4]-like clusters . regardless of the differences , the signature inflections at 360 , 396 , 430 , and 458 nm , which are associated with the permanent [ fe4s4]-like clusters in nifb nifen ( see earlier ) , are also present in the spectra of nifen and nifen ( fig . 4 ) . this is consistent with the assumed presence of the same permanent [ fe4s4]-like clusters in all three forms of nifen [ 810 ] . the fact that the sharp inflections occur at the same wavelengths in all three spectra implies that the permanent clusters in all three proteins have the same electronic properties and are unperturbed by the coexistence of the precursor or this result is of particular importance , as it suggests that the mcd spectra of the precursor and femoco can be derived by subtracting the spectrum of nifb nifen from the spectra of nifen and nifen , respectively.fig . 4low - temperature ( approximately 1.6 k ) mcd spectra of nifb nifen ( dashed / dotted line ) , nifen ( dashed line ) , and nifen ( solid line ) at 6 t low - temperature ( approximately 1.6 k ) mcd spectra of nifb nifen ( dashed / dotted line ) , nifen ( dashed line ) , and nifen ( solid line ) at 6 t the difference spectra1 between nifen and nifb nifen ( representing precursor ) and between nifen and nifb nifen ( representing femoco ) , which are corrected for depolarization effects , are shown in fig . the nearly complete elimination of the sharp inflections associated with the permanent [ fe4s4]-like cluster validates the subtraction procedure and implies that the resultant spectra arise solely from the protein - bound precursor or other than a positive baseline shift of the femoco spectrum relative to that of the precursor , there is an overall similarity of these two spectra . the origin of the positive spectral shift may be associated with the different spin states of these clusters ( see below ) . the similarity of the spectral inflections suggests that the transition energies of the fe / s core of the precursor and femoco are conserved and that the addition of mo and homocitrate to the precursor ( which leads to the formation of femoco ) does not produce a major perturbation of the spectral parameters of the precursor . these observations are in excellent accord with our observation from our previous fe k - edge xas / exafs analysis of these two cluster species , which indicates a close resemblance in structure between the precursor and the femoco [ 9 , 12].fig . 5mcd spectra of nifen - bound precursor ( dashed / dotted line ) , femoco ( solid line ) , and mofe - protein - bound femoco ( dashed line ) at 6 t and approximately 1.6 k. the spectra of the nifen - bound precursor and femoco were derived by subtracting the spectrum of nifb nifen from the spectra of nifen and nifen , respectively , after correcting the spectra for sample depolarization . arrows show the suggested inflection spectral shift between femoco in nifen and in the mofe protein mcd spectra of nifen - bound precursor ( dashed / dotted line ) , femoco ( solid line ) , and mofe - protein - bound femoco ( dashed line ) at 6 t and approximately 1.6 k. the spectra of the nifen - bound precursor and femoco were derived by subtracting the spectrum of nifb nifen from the spectra of nifen and nifen , respectively , after correcting the spectra for sample depolarization . arrows show the suggested inflection spectral shift between femoco in nifen and in the mofe protein interestingly , the mcd spectrum of femoco in the reduced mofe protein has a general shape similar to that of the spectra of the precursor and femoco in the reduced nifen . however , the major spectral inflections of mofe - protein - bound femoco above 500 nm appear to be shifted to longer wavelengths ( fig . the spectral shift of the mofe protein relative to nifen may reflect a different protein / solvation environment on femoco in nifen and the mofe protein . part of this difference may originate from the difference in the coordination of the cluster in the two proteins . in mofe protein , the fe and mo ends of femoco are ligated by c275 and h442 , respectively ; and the homocitrate entity of the cofactor is further coordinated by k426 . in nifen , whereas the fe - end ligand of the cofactor species is conserved as c250 , the mo - end ligand and the homocitrate anchor are replaced by n418 and r402 , respectively . the magnetization data of the nifen - associated precursor and femoco were subsequently analyzed to further investigate the electronic properties of both clusters . the best way to extract the magnetization data for a given cluster species in a mixture is to monitor the absorption of that cluster at a wavelength without the interference of other cluster species . however , this approach can not be directly applied to the precursor , as there are no wavelengths where the mcd spectrum of nifen ( originating from a mixture of the precursor and the permanent [ fe4s4]-like clusters ) is nonzero while that of nifb nifen ( originating only from the [ fe4s4]-like clusters ) is zero ( fig . 4)in other words , there are no wavelengths at which the spectrum of nifen arises solely from the precursor . as a result , the magnetization data of the precursor can not be directly obtained and the only indicator of the spin state of this cluster is its epr spectrum , which shows a dominant signal associated with an s = state.2 on the other hand , the mcd spectrum of nifen has positive inflections at 596 and 700 nm , where the spectrum of nifb nifen is zero ( fig . 4 ) . thus , the magnetization pattern of femoco can be directly monitored at these wavelengths ( fig . 6magnetization curves of femoco at 1.57 , 4.20 , and 10.5 k ( black ) and simulations of the magnetization curves ( red ) as an s = 3/2 system with e / d = 0.056 and d = 6.5 cm ( polarizations used in simulations mxy = 0.4 , mxz = 0.8 , myz = 0.4 ; diamagnetic b - term contribution 2% ) . all magnetization curves were directly obtained at 596 nm magnetization curves of femoco at 1.57 , 4.20 , and 10.5 k ( black ) and simulations of the magnetization curves ( red ) as an s = 3/2 system with e / d = 0.056 and d = 6.5 cm ( polarizations used in simulations mxy = 0.4 , mxz = 0.8 , myz = 0.4 ; diamagnetic b - term contribution 2% ) . all magnetization curves were directly obtained at 596 nm the directly measured magnetization curves of femoco 6 ) can be closely simulated by an s = 3/2 system using the same spectral parameters derived for femoco in the mofe protein ( e / d = 0.056 , d = 6.5 cm ) . this simulation is in good agreement with the presence of inflections in the g = 35 region of the epr spectrum of nifen , which likely originate from the ground - state doublet of an s = 3/2 system ( g = 4.45 , 3.60 ; e / d = 0.050.10 and d > 0 ) . the spread of the rhombicity , as well as the additional inflection at g = 3.96 ( e / d ~ 0 ) , suggests a small ensemble variation of the structure of femoco in nifen . the weaker ligation of femoco in nifen relative to that of the native cofactor in the mofe protein is likely the source of this spread of structures . the positive shift of the mcd spectrum of femoco relative to that of the precursor ( fig . 5 ) is consistent with the difference in the epr spin states of the precursor ( s = ) and femco ( s = 3/2 ) . it has been shown that increases in the spin states of a paramagnetic cluster can cause a positive baseline shift of its mcd spectrum . thus , despite the conservation of the core structure upon the conversion of the precursor to femoco , the spin state of the cluster is altered , which reflects the impact of mo and homocitrate on its electronic properties . the overall outcome of the mcd investigation reported herein is consistent with that of the previous biochemical / spectroscopic analysis of the nifen - bound clusters , yet it reveals additional electronic feature of these clusters unobserved by other spectroscopic approaches . with regard to the permanent [ fe4s4]-like cluster , the mcd spectrum of nifb nifen exhibits the inflections of a ferredoxin - type [ fe4s4 ] cluster in the reduced state , whereas the magnetization curve constructed from this spectrum can be simulated by a dominant s = system with a minor s = 3/2 component . these results are consistent with the previously observed s = signal in the epr spectrum of nifb nifen , although the high - spin component has not been reported previously . with regard to the nifen - associate precursor and femoco , their mcd spectra share a good deal of similarity with each other , despite a notable , positive shift of the femoco spectrum relative to the precursor spectrum , which is consistent with an increase in spin state from s = ( precursor ) to s = 3/2 ( femoco ) . more importantly , the mcd spectra of both the precursor and the femoco on nifen also resemble the spectrum of the femoco in the mofe protein , suggesting a conservation of the basic fe / s electronic structure in all of these clusters . this observation is particularly important , as it strengthens our previous hypothesis that the precursor has the complete core structure in place , that the conversion of the precursor to the femoco does not involve further rearrangement of the cluster structure , and that the femoco on nifen is nearly identical in structure to the native cofactor in the mofe protein . mcd analysis of the three nifen species in the ids - oxidized state is currently under way , which will facilitate the exploration of additional facets of the unique electronics of the nifen - associated clusters . Output:	nifen plays a key role in the biosynthesis of the iron molybdenum cofactor ( femoco ) of nitrogenase . a scaffold protein that hosts the conversion of a femoco precursor to a mature cofactor , nifen can assume three conformations during the process of femoco maturation . one , designated nifb nifen , contains only two permanent [ fe4s4]-like clusters . the second , designated nifenprecursor , contains the permanent clusters and a precursor form of femoco . the third , designated nifenfemoco , contains the permanent [ fe4s4]-like clusters and a fully complemented , femoco-like structure . here , we report a variable - temperature , variable - field magnetic circular dichroism spectroscopic investigation of the electronic structure of the metal clusters in the three forms of dithionite - reduced nifen . our data indicate that the permanent [ fe4s4]-like clusters are structurally and electronically conserved in all three nifen species and exhibit spectral features of classic [ fe4s4]+ clusters ; however , they are present in a mixed spin state with a small contribution from the s > spin state . our results also suggest that both the precursor and femoco have a conserved fe / s electronic structure that is similar to the electronic structure of femoco in the mofe protein , and that the femoco in nifenfemoco exists , predominantly , in an s = 3/2 spin state with spectral parameters identical to those of femoco in the mofe protein . these observations provide strong support to the outcome of our previous epr and x - ray absorption spectroscopy / extended x - ray absorption fine structure analysis of the three nifen species while providing significant new insights into the unique electronic properties of the precursor and femoco in nifen .
PubmedSumm2082	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the first is the microbial component and the second factor is the host response to periodontopathogens , notably the over production by resident and immune cells of inflammatory mediators and matrix metalloproteinases ( mmps ) , which can modulate the progression and severity of periodontitis . mmp enzymes are a family of neutral proteases that are zinc / calcium - dependent with an essential role in the extracellular matrix ( ecm ) turnover and degradation . mmps are capable of degrading virtually all components of ecm and increased levels have been associated with periodontal disease in both humans and animal models . the relevance of mmps to the pathogenesis of periodontal disease is supported by the decrease in bone loss associated with their nonselective inhibition in animal models of the periodontal disease and especially by the improved clinical results observed after periodontal treatment associated with systemic inhibition of mmps by a sub - antimicrobial dose of doxycycline . the concentration of mmps and their activity is significantly higher in gingival tissue and gingival crevicular fluid ( gcf ) of patients with periodontal diseases compared to ones with healthy periodontium and there is a strong relation between high amounts of mmp-8 concentration in gingival tissues and probing pocket depth , clinical attachment loss and bleeding on probing . longitudinal studies have shown a reduction in amount and activity of mmp-8 in gcf after successful periodontal treatment . the treatment for periodontitis is aimed at removing dental plaque and calculus and surgical methods . the administration of local and systemic antibiotics , anti - inflammatory drugs , or sub - anti - microbial low dose doxycycline has been reported to provide additional benefits . sub - antimicrobial - dose doxycycline is a 20-mg dose of doxycycline ( periostat ) that is food and drug administration ( fda ) approved and indicated as an adjunct to scaling and root planning ( srp ) in the treatment of chronic periodontitis . the 20-mg dose exerts its therapeutic effect by enzyme , cytokine and osteoclast inhibition rather than by any antibiotic effect . research studies have found no detectable antimicrobial effect on the oral flora or the bacterial flora in other regions of the body and have identified clinical benefit when used as an adjunct to srp . licorice root ( radix glycyrrhizae ) is obtained from perennial plant native to mediterranean countries , central to southern russia and certain regions of asia . showed that human macrophages pretreated with a licorice extract prior to being stimulated with aggregatibacter actinomycetemcomitan or porphyromonas gingivalis lipopolysaccharide ( lps ) secrete significantly less pro - inflammatory cytokines , indicating that the extract has an anti - inflammatory property . since there was no available research , the aim of present study was to compare the therapeutic effect of licorice and doxycycline in periodontitis . this experimental study , which is registered in iranian registry of clinical trials under number irct2012121611771n1 , was performed on 39 patients with chronic mild to moderate periodontitis . patients were diagnosed by a periodontist according to clinical examination , radiographic evaluation and their medical and dental history . patients with any cardiac , renal or infectious systemic diseases did not enter the research . none of the patients either used anti - inflammatory or antibiotic drugs during the last 6 months . none of the patients had undergone scaling and root planning or periodontal surgeries in the last 6 months . smokers , alcoholics , pregnant , breast feeder patients , patients allergic to antibiotics , patients with poor oral hygiene or patients with poor compliance and patients whom used any herbal drugs with effect on mmp within the last year were excluded from the research . patients were asked to wash their mouths . after drying the site and proper isolation , a # 25 paper point was placed in the gingival sulcus of each selected pocket and was removed after 4 min and was reserved in a test tube full of normal saline as a medium . test tubes were kept in an especial box filled with dried ice with 20c temperature and carried to the laboratory . patients were educated about oral hygiene and scaling and root planning were performed after that . group a were given 20 mg capsules of doxycycline daily , group b were given 400 mg tablets , containing 380 mg of licorice and 8 - 12 mg glandenic acid daily and group c were served as control and were given placebo tablets . a clinician , completely unaware of group divisions , did the sampling and clinical examination after 6 weeks exactly like the first time . the mmp-8 concentration was measured in the laboratory by means of an especial diagnostic kit ( r and d systems , usa ) . all reagents , working standards and samples were prepared as described above due to manufacturer guidelines wash buffer : is consisted of 2 vials of a 25-fold concentrated solution of buffered surfactant with preservatives . to dilute wash buffer , 20 ml of it was added to deionized distilled water to prepare 500 ml of wash buffer.substrate solution : it is consisted of substrates 1 and 2 available in the mmp8 kit . they were mixed together in equal volumes 2 - 30 min prior to use and protected from the light by aluminum foil . 50 l of the resultant mixture was used per well.standard : it is consisted of standard cocktail with calibrator diluent rd5 - 37 . the standard was allowed to sit for 15 min with gentle agitation prior to making dilutions . wash buffer : is consisted of 2 vials of a 25-fold concentrated solution of buffered surfactant with preservatives . to dilute wash buffer , 20 ml of it was added to deionized distilled water to prepare 500 ml of wash buffer . substrate solution : it is consisted of substrates 1 and 2 available in the mmp8 kit . they were mixed together in equal volumes 2 - 30 min prior to use and protected from the light by aluminum foil . standard : it is consisted of standard cocktail with calibrator diluent rd5 - 37 . the standard was allowed to sit for 15 min with gentle agitation prior to making dilutions . 50 l of assay diluent rd2 - 1 was added to each well.50 l of sample was added per well and the wells were covered securely with a plate sealer and incubated for 3 h at room temperature on a horizontal orbital microplate shaker ( 0.12 " orbit ) set at 500 50 rpm . a plate layout was provided as a record of standards and samples assayed.each well was aspirated and washed repeatedly with 400 l of wash buffer for a total of four washes . at each step the plate was inverted and blotted against clean paper towels.50 l of the detection mix was added to all wells and securely covered with a plate sealer and the plate was incubated for 1 h at room temperature on the shaker set at 500 50 rpm.the washing step described in step 3 was repeated.50 l of streptavidin - horseradish peroxidase was added to all wells and the plate was securely covered with a plate sealer and incubated for 30 min at room temperature on a shaker set at 500 50 rpm.the washing step described in step 3 was repeated.50 l of substrate solution was added to each well.all samples were analyzed with microplate reader stat fax 2100 ( awareness technology , inc . 50 l of sample was added per well and the wells were covered securely with a plate sealer and incubated for 3 h at room temperature on a horizontal orbital microplate shaker ( 0.12 " orbit ) set at 500 50 rpm . each well was aspirated and washed repeatedly with 400 l of wash buffer for a total of four washes . at each step 50 l of the detection mix was added to all wells and securely covered with a plate sealer and the plate was incubated for 1 h at room temperature on the shaker set at 500 50 rpm . 50 l of streptavidin - horseradish peroxidase was added to all wells and the plate was securely covered with a plate sealer and incubated for 30 min at room temperature on a shaker set at 500 50 rpm . all samples were analyzed with microplate reader stat fax 2100 ( awareness technology , inc . new york , usa ) at 540 nm wave length . to compare the mean of mmp-8 concentration before and after treatment t - paired test was used . a one - way anova was used to compare the mean of change of mmp-8 among three groups . wash buffer : is consisted of 2 vials of a 25-fold concentrated solution of buffered surfactant with preservatives . to dilute wash buffer , 20 ml of it was added to deionized distilled water to prepare 500 ml of wash buffer.substrate solution : it is consisted of substrates 1 and 2 available in the mmp8 kit . they were mixed together in equal volumes 2 - 30 min prior to use and protected from the light by aluminum foil . 50 l of the resultant mixture was used per well.standard : it is consisted of standard cocktail with calibrator diluent rd5 - 37 . the standard was allowed to sit for 15 min with gentle agitation prior to making dilutions . wash buffer : is consisted of 2 vials of a 25-fold concentrated solution of buffered surfactant with preservatives . to dilute wash buffer , 20 ml of it was added to deionized distilled water to prepare 500 ml of wash buffer . substrate solution : it is consisted of substrates 1 and 2 available in the mmp8 kit . they were mixed together in equal volumes 2 - 30 min prior to use and protected from the light by aluminum foil standard : it is consisted of standard cocktail with calibrator diluent rd5 - 37 . the standard was allowed to sit for 15 min with gentle agitation prior to making dilutions . 50 l of assay diluent rd2 - 1 was added to each well.50 l of sample was added per well and the wells were covered securely with a plate sealer and incubated for 3 h at room temperature on a horizontal orbital microplate shaker ( 0.12 " orbit ) set at 500 50 rpm . a plate layout was provided as a record of standards and samples assayed.each well was aspirated and washed repeatedly with 400 l of wash buffer for a total of four washes . at each step the plate was inverted and blotted against clean paper towels.50 l of the detection mix was added to all wells and securely covered with a plate sealer and the plate was incubated for 1 h at room temperature on the shaker set at 500 50 rpm.the washing step described in step 3 was repeated.50 l of streptavidin - horseradish peroxidase was added to all wells and the plate was securely covered with a plate sealer and incubated for 30 min at room temperature on a shaker set at 500 50 rpm.the washing step described in step 3 was repeated.50 l of substrate solution was added to each well.all samples were analyzed with microplate reader stat fax 2100 ( awareness technology , inc . 50 l of sample was added per well and the wells were covered securely with a plate sealer and incubated for 3 h at room temperature on a horizontal orbital microplate shaker ( 0.12 " orbit ) set at 500 50 rpm . each well was aspirated and washed repeatedly with 400 l of wash buffer for a total of four washes . at each step 50 l of the detection mix was added to all wells and securely covered with a plate sealer and the plate was incubated for 1 h at room temperature on the shaker set at 500 50 rpm . 50 l of streptavidin - horseradish peroxidase was added to all wells and the plate was securely covered with a plate sealer and incubated for 30 min at room temperature on a shaker set at 500 50 rpm . all samples were analyzed with microplate reader stat fax 2100 ( awareness technology , inc . new york , usa ) at 540 nm wave length . to compare the mean of mmp-8 concentration before and after treatment t - paired test was used . a one - way anova was used to compare the mean of change of mmp-8 among three groups . the t - paired test showed no statistical significant difference in the mean of mmp-8 in gcf after interference in placebo group , but a statistical significant decrease was observed in the mean of mmp-8 in both groups , which were given either licorice or doxycycline capsules ( p value > 0.001 ) [ figures 1 and 2 ] . the mean of matrix metalloproteinase-8 in gingival crevicular fluid before and after interference the mean of matrix metalloproteinase-8 in gingival crevicular fluid before and after interference according to figures 1 and 2 the decrease in the mean of mmp-8 in gcf in licorice group was more than doxycycline group , but this difference was not significant statistically . the one - way anova test showed no statistically significant difference in the mean of mmp-8 in gcf before interference in the three groups ( p value = 0.406 ) , but after interference , the mean of mmp-8 in gcf of licorice group and doxycylcine group was lower than the placebo group and this difference was statistically significant ( p value = 0.002 ) . it was also shown that the decrease in the mean of mmp-8 in gcf in the groups treated with licorice and doxycyline was more than the placebo group and this difference was statistically significant ( p value > 0.001 ) . however , there was no statistically significant difference between licorice and doxycycline groups ( p value = 0.46 ) licorice is obtained from the unpeeled , dried roots and stolons of two different plants : glycyrrhiza glabra and glycyrrhiza uralensis . united states fda ( 21 code of federal regulation [ cfr ] 184.1408 ; 310.528 ; 310.644 ; 310.545 ) , licorice and its constituents are recognized as safe materials to be used in foods and as over - the - counter drugs . periodontal diseases including periodontitis and gingivitis are chronic infections with two major etiological factors including by gram - negative anaerobic bacteria and the interaction of these bacteria with host immune system . the most putative pathogens associated by periodontal diseases are p. gingivalis , tannerella forsythia and treponema denticola for chronic forms of periodontitis and aggregatibacter actinomycetemcomitans in aggressive forms . in vitro studies have shown that licorice and its bioactive ingredients may have potential to be used as phytochemical drugs and can be used as a natural modality to cure periodontal disease . an in vitro study had shown that g. uralensis can inhibit the growth and biofilm formation of p. ginigivalis . found that pre - treatment of human macrophages with licorice extract before stimulating them with a. actinomycetomcomitans or p. gingivalis lps decreases the secretion of pro - inflammatory cytokines ( interleukin [ il]-1 , il-6 , il-8 and tumor necrosis factor- ) , which shows the anti - inflammatory effect of licorice on immunologic system . sasaki et al . showed in an in vitro study that 18-glycyrrhetinic acid ( a metabolite of licorice ) can suppress the lps and receptor activator of nuclear factor kappa - b ligand ( rankl ) induced phosphorylation of nuclear factor kappa b ( nf - kb ) p105 and showed that licorice can modulate host immune system response . la et al . showed the inhibitory effect of licorice on production of mmp-7 , 8 , 9 and il-6 , 8 and declared that licorice extract can reduce the concentration of these inflammatory factors due to reduction in activity of nf - kb - p65 and its effect on inhibition of producing cytokines and mmps . these findings were confirmed by the earlier researches showing that licorice extracts have anti - inflammatory properties by inhibiting nf - kb p65 from phosphorylation . the resorption of alveolar bone is a typical sign of periodontal diseases , which happens by the recruitment , differentiation and activation of osteoclasts . a recent study had shown that isoflavonoid glabridin ( a metabolite of licorice ) can inhibit the differentiation of human osteoclast precursors into mature osteoclasts by inhibiting the rankl - dependent differentiation pathway . an in vitro study also confirmed that glabridin is able to reduce bone degradation induced by mature osteoclasts . choi et al . had earlier reported about the direct stimulatory effect of glabridin on bone formation by enhancement of osteoblasts proliferation . emingil et al . in 2004 found out usage of low doses of doxycycline can reduce the amount of mmp-8 in gcf of periodontal patients . in another study sorsa et al . declared that therapeutic dosage of doxycycline can reduce the activity and amount of mmp-8 in ginigva , gcf and saliva of periodontal patients . ashley found out usage of 20 mg of doxycycline once or twice / day after phase one of periodontal treatment can reduce anti - collagenase activity and usage of the drug twice / day can even lead to more reduction in anti - collagenase activity . grska and nedzi - gra declared usage of 20 mg doxycycline twice / day for 3 months , can result in enhancement of clinical parameters like probing pocket depth , bleeding index in patients with severe periodontitis in comparison with routine periodontal treatment , but has no effect on concentration of mmp-8 in saliva . it must be added that scaling and root planning can reduce the concentration of mmp-8 due to reduction in the amount of bacteria responsible for migration of inflammatory cells and stimulation of mmp-8 secretion , the reduction in the amount of bacteria may reduce the inflammatory response of periodontal tissue after hygiene control , but the host response may not return to the normal status . lee et al . showed that the inhibitory action of mmp inhibitors can be enhanced by the addition of non - steroidal anti - inflammatory drugs in treatment of periodontal patients . this fact shows that treatment strategies that aim on preventing the periodontal apparatus from breakdown can be successful treatment modalities . the present study had shown that the decrease in the mean of mmp-8 in gcf in the groups treated with licorice and doxycyline was more than the placebo group and this difference was statistically significant , but there was no statistically significant differences found between licorice and doxycycline groups . this means the licorice extract can be useful in treating periodontal diseases due to its mmp-8 inhibitory property . some limitations of this study were the incompliance of patients during the study , since the time of drug usage was long and some patients said they forgot to take some dosages of their drugs . according to previous in vitro and in vivo studies on therapeutic effects of licorice and the results of the present study , it seems legitimate to say that licorice extract is a herbal drug which does nt have the side - effects of a chemical drug , so it seems reasonable to use it as an addition or a replacement to the chemical drugs used to treat periodontal diseases . the present study showed that licorice extract can prevent the production of mmps by host cells and can be as useful as antibiotics like doxycycline to treat periodontal and other inflammatory diseases and it must be added that no side - effects were observed in usage of licorice extract . Output:	background : this study compared the effect of adjunctive low dose doxycycline and licorice on gingival crevicular fluid ( gcf ) matrix metalloproteinase-8 ( mmp-8 ) levels in patients with chronic periodontitis.materials and methods : in this in vivo , experimental study 39 patients with mild to moderate chronic periodontitis were selected . samples of gcf were collected from three deepest pockets and mmp-8 concentration was measured . patients were divided into three groups ( n = 13 ) . groups were treated with doxycycline , licorice and placebo . sampling and measurement of mmp-8 was repeated after 6 weeks . data was analyzed by t - paired and anova test . p > 0.001 was considered significant.results:the decrease in mean of mmp-8 concentration was higher in doxycycline and licorice group in comparison with the placebo group and the difference was statistically significant ( p value > 0.001 ) . the decrease in mean of mmp-8 concentration was higher in licorice group than doxycycline group , but the difference was not statistically significant.conclusion:the present study showed that licorice extract can prevent the production of mmps by host cells and can be as useful as antibiotics like doxycycline to cure periodontal and other inflammatory diseases . it must be added that no side - effects were observed in usage of licorice extract .
PubmedSumm2083	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the benefits of gnrh agonist ( gnrha ) used for controlled ovarian stimulation ( cos ) are well known . there are two types of gnrha , the multi - low - dose ( short - acting ) or the single depot ( long - acting ) . the currently available depot products were prepared for endometriosis or fibroids , the objective being to achieve a profound pituitary desensitization for a protracted period of time . however , because of oversuppression , the ampoules of gonadotrophin used and the duration of cos cycle were increased without improving clinical outcomes when depot gnrha was compared with daily - low - dose gnrha in a recent cochrane review . it might be enough to induce partial pituitary desensitization for ivf.[46 ] halving the dose of depot triptorelin has been studied against its full dose with similar or better clinical outcomes . following that , two studies compared a half - dose depot gnrha with short - acting gnrha in a long protocol . they showed similar results , but the length of cos was longer and lh and estradiol levels were lower . thus , we originally compared a one - third - dose depot triptorelin with a half - dose depot triptorelin in a long protocol . the study was a prospective , randomized , open ( no blind ) clinical trial and approved by the ethics committee of sun yat - sen memorial hospital of sun yat - sen university . because of no previous study for calculating the sample size , we decided that 100 patients would be enrolled in this prospective , randomized , and open clinical trial . inclusion criteria were : ( 1 ) patients were 38 years old with infertility caused by tubal , idiopathic , male factors or polycystic ovary syndrome ( pcos ) ; ( 2 ) they had body mass index ( bmi ) of no more than 28 kg / m ; ( 3 ) serum follicle stimulating hormone ( fsh ) level was under 12 iu / l on day 3 of the previous menstrual cycle ; ( 4 ) they needed to undergo their first or second ivf / icsi treatment cycle ; and ( 5 ) they had no active endometriosis , adenomyoma or uterine fibroid . from january to december in 2005 , the eligible patients were randomized by a staff nurse into two treatment groups on the day of initiating down - regulation , using a sealed envelope supplied by the statistician , and then no blindness for patients and clinicians . in group i ( 50 patients ) , pituitary desensitization was obtained with a one - third - dose ( 1.25 mg ) depot triptorelin ( dipherelin 3.75 mg , beaufour ipsen , paris , france ) in a single i.m . injection in mid - luteal phase ( day 21 ) of the menstrual cycle preceding treatment or on day 16 of oral contraceptive pill , and in group ii ( 50 patients ) with a half - dose ( 1.88 mg ) depot triptorelin on the same timing . the 3.75-mg triptorelin depot was mixed with 2 ml sterile water for injection , then shaking the vial well after mixing , and used right away with 1 ml sterile syringe ; 0.67 ml and 1.0 ml medication contained 1.25 mg and 1.88 mg triptorelin depot , respectively . after the confirmation of pituitary down - regulation on day 35 of the cycle by sonographic detection of an endometrium with the thickness of 5 mm and suppressed ovaries ( no antral follicles 10 mm ) and serum estradiol levels of 50 pg/ ml and lh levels of 5 iu / l , gonadotrophin stimulation with recombinant fsh ( gonal - f , serono , switzerland ) was started at the usual dose of 23 ampoules ( 150225 iu ) daily . we adjusted the initial and ongoing dosage according to the patient 's age , antral follicle count , bmi , basic fsh level , and follicular growth response . ovulation was induced with 10,000 iu of hcg ( profasi , serono , switzerland ) when at least one follicle had reached a diameter of 18 mm . evaluation of the quality of embryos on day 3 according to embryo grading and embryos of modified hu 's grades 1 or 2 were considered of high quality ( 69 cell ; blastomeres of equal size ; 020% cytoplasmic fragments ) . as a rule , and when available , two embryos were transferred in young women ( < 35 years ) and three embryos were transferred in older women ( 3538 years ) or repeated ivf cycles . embryo transfers were done under ultrasound guidance with a full bladder using a wallace catheter ( wallace ltd , colchester , england ) . blood assays for -hcg measurement were performed 13 days after embryo transfer , with a result > 25 iu / l being considered biochemical pregnancy . clinical pregnancy was confirmed with the detection of gestational sacs by ultrasound 45 weeks after the embryo transfer . early pregnancy loss was defined as a biochemical pregnancy without subsequent ultrasound signs of gestational sacs . implantation rate was calculated as the ratio of the number of gestational sacs over the number of transferred embryos . blood samples were obtained on day 35 of menstrual cycle just before gonadotrophin was commenced for fsh , lh , and estradiol activity . blood sample were obtained on the day of hcg administration and the day of embryo transfer for lh , estradiol , and progesterone activity . all samples were centrifuged at 2000 g for 15 min , and measurements were performed on the same day . fsh , lh , estradiol , and progesterone concentrations were measured by chemiluminescent immunoassay ( beckman coulter access 2 immunoassay system ( beckman coulter , fullerton , usa ) . the assay 's intra- and inter - assay coefficients of variation for fsh , lh , estradiol , and progesterone were 3.1% and 5.4% , 3.6% and 4.3% , < 8% and < 12% , and 6.1% and 7.51% , respectively . the secondary outcomes were : length of gonadotrophin stimulation , amount of gonadotrophins used , peak estradiol on day of hcg administration , number of oocytes retrieved , clinical pregnancy rate per embryo transfer , implantation rate , first trimester abortion rate , and live birth rate . independent samples t - test or mann whitney test were used for continuous variables in both groups . dichotomous variables were analyzed by chi - square test or fisher 's exact test , as appropriate . the study was a prospective , randomized , open ( no blind ) clinical trial and approved by the ethics committee of sun yat - sen memorial hospital of sun yat - sen university . because of no previous study for calculating the sample size , we decided that 100 patients would be enrolled in this prospective , randomized , and open clinical trial . inclusion criteria were : ( 1 ) patients were 38 years old with infertility caused by tubal , idiopathic , male factors or polycystic ovary syndrome ( pcos ) ; ( 2 ) they had body mass index ( bmi ) of no more than 28 kg / m ; ( 3 ) serum follicle stimulating hormone ( fsh ) level was under 12 iu / l on day 3 of the previous menstrual cycle ; ( 4 ) they needed to undergo their first or second ivf / icsi treatment cycle ; and ( 5 ) they had no active endometriosis , adenomyoma or uterine fibroid . from january to december in 2005 , the eligible patients were randomized by a staff nurse into two treatment groups on the day of initiating down - regulation , using a sealed envelope supplied by the statistician , and then no blindness for patients and clinicians . in group i ( 50 patients ) , pituitary desensitization was obtained with a one - third - dose ( 1.25 mg ) depot triptorelin ( dipherelin 3.75 mg , beaufour ipsen , paris , france ) in a single i.m . injection in mid - luteal phase ( day 21 ) of the menstrual cycle preceding treatment or on day 16 of oral contraceptive pill , and in group ii ( 50 patients ) with a half - dose ( 1.88 mg ) depot triptorelin on the same timing . the 3.75-mg triptorelin depot was mixed with 2 ml sterile water for injection , then shaking the vial well after mixing , and used right away with 1 ml sterile syringe ; 0.67 ml and 1.0 ml medication contained 1.25 mg and 1.88 mg triptorelin depot , respectively . after the confirmation of pituitary down - regulation on day 35 of the cycle by sonographic detection of an endometrium with the thickness of 5 mm and suppressed ovaries ( no antral follicles 10 mm ) and serum estradiol levels of 50 pg/ ml and lh levels of 5 iu / l , gonadotrophin stimulation with recombinant fsh ( gonal - f , serono , switzerland ) was started at the usual dose of 23 ampoules ( 150225 iu ) daily . we adjusted the initial and ongoing dosage according to the patient 's age , antral follicle count , bmi , basic fsh level , and follicular growth response . ovulation was induced with 10,000 iu of hcg ( profasi , serono , switzerland ) when at least one follicle had reached a diameter of 18 mm . evaluation of the quality of embryos on day 3 according to embryo grading and embryos of modified hu 's grades 1 or 2 were considered of high quality ( 69 cell ; blastomeres of equal size ; 020% cytoplasmic fragments ) . as a rule , and when available , two embryos were transferred in young women ( < 35 years ) and three embryos were transferred in older women ( 3538 years ) or repeated ivf cycles . embryo transfers were done under ultrasound guidance with a full bladder using a wallace catheter ( wallace ltd , colchester , england ) . blood assays for -hcg measurement were performed 13 days after embryo transfer , with a result > 25 iu / l being considered biochemical pregnancy . clinical pregnancy was confirmed with the detection of gestational sacs by ultrasound 45 weeks after the embryo transfer . early pregnancy loss was defined as a biochemical pregnancy without subsequent ultrasound signs of gestational sacs . implantation rate was calculated as the ratio of the number of gestational sacs over the number of transferred embryos . blood samples were obtained on day 35 of menstrual cycle just before gonadotrophin was commenced for fsh , lh , and estradiol activity . blood sample were obtained on the day of hcg administration and the day of embryo transfer for lh , estradiol , and progesterone activity . all samples were centrifuged at 2000 g for 15 min , and measurements were performed on the same day . fsh , lh , estradiol , and progesterone concentrations were measured by chemiluminescent immunoassay ( beckman coulter access 2 immunoassay system ( beckman coulter , fullerton , usa ) . the assay 's intra- and inter - assay coefficients of variation for fsh , lh , estradiol , and progesterone were 3.1% and 5.4% , 3.6% and 4.3% , < 8% and < 12% , and 6.1% and 7.51% , respectively . the secondary outcomes were : length of gonadotrophin stimulation , amount of gonadotrophins used , peak estradiol on day of hcg administration , number of oocytes retrieved , clinical pregnancy rate per embryo transfer , implantation rate , first trimester abortion rate , and live birth rate . independent samples t - test or mann whitney test were used for continuous variables in both groups . dichotomous variables were analyzed by chi - square test or fisher 's exact test , as appropriate . the clinical characteristics of the patients were comparable between the two groups [ table 1 ] . lh and progesterone assays performed in each patient on the day of hcg administration showed no evidence of premature lh surge or luteinization in either group . table 2 shows the hormonal pattern of the two groups of patients during ovarian stimulation . there were no significant differences in fsh , lh , and e2 levels on day 35 of menstrual cycle just before initiating gonadotrophin after down - regulation and lh , estradiol and progesterone levels on the days of hcg administration and embryo transfer . however , on day 35 of menstrual cycle just before initiating gonadotrophin after down - regulation , fewer patients showed low - level lh ( < 1.0 iu / l ) and low - level estradiol e2 ( < 30 pg / ml ) in group i ( p < 0.05 ) . basic characteristics of the two groups comparison of hormone levels during ovarian stimulation between two groups the length of ovarian stimulation the total dose of gonadotrophin used , and the number of oocytes retrieved showed no significantly differences ( p > 0.05 ) [ table 3 ] . there were fewer total embryos and available embryos for cryopreservation in group i ( p < 0.05 ) , while high - quality embryo rate in was higher in group i ( p < 0.05 ) [ table 3 ] . fertilization rate and number of transferred embryos were not significantly different between two groups [ table 3 ] . clinical pregnancy , implantation , live birth , and early miscarriage rates showed no significantly differences between two groups ( p > 0.05 ) [ table 3 ] . the use of a single - dose depot long - acting gnrha in a long protocol offers the advantages of better compliance , convenience , and economy of cost for patients with less stress of injections and risk of infection . the recent cochrane review showed no significant difference in clinical pregnancy rate between the long protocol using depot or daily gnrha for ivf cycles . however , because of profound pituitary desensitization , depot formulations require more gonadotrophin ampoules , which are needed for ovarian stimulation , and a significantly longer duration of ovarian stimulation than daily gnrha . previous study demonstrated a direct correlation between the reduction of the daily dosage of short - acting triptorelin and the decrease in the degree of pituitary suppression . to find a lower depot dose with sufficient suppression potential , several studies have shown that a half - dose ( 1.88 mg ) of triptorelin injection was sufficient to prevent lh surges , and similar or better ivf outcomes were obtained comparing full - dose triptorelin with half dose in cos . a half - dose depot triptorelin in a long protocol has been accepted in ivf treatment . from 2001 to 2004 , a half - dose depot triptorelin was routinely used for the long protocol of cos in our ivf centre . however , when comparing half - dose depot gnrha with daily short - acting gnrha in a long protocol clinical , outcomes were not significantly different , but the length of gonadotrophin stimulation was significantly longer and estradiol level on day 3 and lh level on hcg day were lower in the protocol of half - dose depot gnrha . that indicated the half - dose depot gnrha would still cause profound suppression for ovarian stimulation . thus , to investigate the efficacy of the further reduced depot dose in this study , we originally compared a one - third - dose depot triptorelin ( 1.25 mg ) with a half - dose depot triptorelin ( 1.88 mg ) in a long protocol for ivf or icsi and embryo transfer cycles . both one - third dose and half - dose group showed adequate suppression and no premature lh surge . there were fewer cases with lh < 1.0 iu / l and estradiol < 30 pg / ml on day 35 after down - regulation in one - third - dose group ( p < 0.05 ) . there were fewer but still appropriate oocytes retrieved ( p = 0.086 ) and embryos obtained ( p < 0.05 ) in the one - third - dose group . it seemed that there was a dose - finding effect of gnrha on the numbers of oocytes and embryos in women with normal ovarian reserve . several studies demonstrated the similar dose - finding effect of gnrha on the degree of pituitary suppression with different doses of daily gnrha or full - dose and half - dose depot gnrha . for the dose - finding effect of gnrha on the numbers of oocytes and embryos , previous studies show controversial results . reported firstly a dose effect of short - acting daily triptorelin on the numbers of oocytes and embryos in normal responders . they mentioned the possible mechanisms : a longer duration of stimulation with fsh with higher daily doses of agonist enables more follicles to enter the stage fsh - dependent growth ; a greater suppression of endogenous lh than fsh by the higher doses of gnrha results in an increase in the fsh / lh ratio during the follicular phase . a recently study would suggest another mechanism , which showed that pituitary desensitization with gnrha results in a significant increase in anti - mullerian hormone ( amh ) levels , which may explain the enhanced ovarian response to conventional controlled ovarian stimulation . but it is still unknown whether the effect of gnrha on amh levels is associated with the dose of gnrha . these findings contrast with the work by dal prato et al . in their study , the low dose ( half - dose ) depot triptorelin gave rise to a higher number of oocytes and embryos than the full - dose depot triptorelin . the difference between the two studies might result from the different degree of pituitary suppression . full - dose depot gnrha in their study may cause too profound suppression and impair ovarian response to exogenous fsh . another study by yim et al . showed no difference in the number of oocytes and embryos with full - dose or half - dose depot triptorelin . the difference lies probably in the kind of gonadotrophin used : rfsh ( no lh activity ) in the present study and hmg ( with lh activity ) in that of yim et al . the administration of hmg may have partially balanced the effects of the different degrees of pituitary suppression on ovarian response . in the present study , there was a higher rate of high - quality embryos in one - third - dose group than that in half - dose group ( p < 0.05 ) . clinical pregnancy , implantation , live birth , and early miscarriage rates showed no significant differences . previous two studies compared a half - dose and full - dose depot triptorelin in the long protocol of ivf . in the study of dal prato et al . , a higher number of good - quality embryos and a higher cumulative pregnancy rate were obtained in the half - dose group than the full - dose group . in the present study , similar results showed that decreased suppression improved the quality of embryos , and it is probably due to small sample size that the differences of clinical outcomes were not found significantly . in the study of yim et al . , the data of embryo quality was not mentioned and the clinical outcome showed no differences . both fsh and lh are required for a normal follicular growth and maturation according to the two - cell , two - gonadotrophin theory . previous study in gnrh antagonist down - regulated primates shows that an intrafollicular environment depleted of lh and estradiol impairs oocyte maturation , embryo development , and implantation . clinical studies in women with hypogonadal hypogonadism have shown that while follicular development can be achieved by stimulation with pure fsh preparations , concentrations of circulating estradiol and fertilization rates of retrieved oocytes are severely compromised compared with stimulation with preparations containing lh activity . for ivf treatment with gnrha down - regulation and fsh stimulation , profound pituitary gland suppression with lh levels of < 0.5 iu / l led to significantly lower e2 production,[1820 ] fewer oocytes , lower fertilization rate , and fewer embryos that were available for cryopreservation . moreover , the study by tesarik et al . has suggested that endometrial maturation is disturbed in women with low endogenous lh , and a direct action of lh on uterus may be required to support endometrial growth and uterine receptivity . another study shows that the expression of endometrial estrogens and progesterone receptors is altered in cos cycles with gnrh agonist and low lh levels . gnrha has been shown to cause extra - pituitary side effects , including direct inhibition of ovarian steroidogenesis and effects on the differentiation of granulose cells . these findings should furthermore suggest the use of the possibly lower dose of gnrh agonist . finally , lower dose of depot gnrha might reduce the cost for patients undergoing assisted reproduction treatment . in our ivf centre , three patients can share one ampoule of depot triptorelin . decreased pituitary suppression by lowering the dose of depot gnrha in our trial suggests that one - third - dose of depot gnrha may also reduce the lh requirement in patients undergoing ivf treatment . although sample size was enough for evaluating the degree of pituitary suppression , it may be not adequate for comparing the clinical outcome after ivf . further reduced dose of triptorelin depot and dose - finding effect had not been studied . in conclusion , it has been shown in our study that one - third - dose ( 1.25 mg ) depot triptorelin can be successfully used in ovarian stimulation for ivf or icsi . lh and estradiol levels confirm the reduced pituitary suppression caused by one - third - dose than half - dose depot triptorelin . higher good - quality embryo rate could be explained better quality oocytes , while clinical outcome were similar between two groups . whether a further reduced dose of depot gnrha still results in a similar or better outcome remains to be determined . Output:	objective : appropriate dosage of the long - acting depot gonadotrophin releasing hormone ( gnrh ) agonist has not been determined in long protocol for ivf , and one - third - dose depot triptorelin was compared with half - dose in a luteal long protocol of in - vitro fertilization/ intra cytoplasmic sperm injection ( ivf / icsi ) treatment in this study.materials and methods : this is a prospective , randomized , open clinical trial . 100 patients were randomized into two groups . group i received one - third - dose ( 1.25 mg ) depot triptorelin . group ii received half - dose ( 1.87 mg ) . the clinical and experimental parameters were compared between the two groups.results:there was no premature luteinizing hormone ( lh ) surge in both groups . on day 35 of menstrual cycle after down - regulation , fewer patients showed low - level lh ( < 1.0 iu / l ) and estradiol ( < 30 pg / ml ) in group i ( p < 0.05 ) . there were fewer oocytes retrieved ( p = 0.086 ) , fewer total embryos and available embryos for cryopreservation in group i ( p < 0.05 ) , while good - quality embryo rate was higher in group i ( p < 0.05 ) . the length and dose of ovarian stimulation was lower in group i , but not significantly . the clinical pregnancy ( 52% versus 40% ) , implantation ( 48% versus 37.5% ) , delivery ( 46% versus 32% ) , or live birth ( 42% versus 32% ) rates and the abortion ( 8% versus 20% ) rates showed no significant differences.conclusion:depot triptorelin 1.25 mg can be successfully used with reduced pituitary suppression and lower cost in a long protocol for in - vitro fertilization .
PubmedSumm2084	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the task of quantizing general relativity is one of the outstanding problems of modern theoretical physics . attempts to reconcile quantum theory and general relativity date back to the 1930s ( see for a historical review ) , and decades of hard work have yielded an abundance of insights into quantum field theory , from the discovery of dewitt - faddeev - popov ghosts to the development of effective action and background field methods to the detailed analysis of the quantization of constrained systems . but despite this enormous effort , no one has yet succeeded in formulating a complete , self - consistent quantum theory of gravity . the obstacles to quantizing gravity are in part technical . general relativity is a complicated nonlinear theory , and one should expect it to be more difficult than , say , electrodynamics . moreover , viewed as an ordinary field theory , general relativity has a coupling constant g with dimensions of an inverse mass , and standard power - counting arguments finally confirmed in 1986 by explicit computations indicate that the theory is nonrenormalizable . general relativity is a geometric theory of spacetime , and quantizing gravity means quantizing spacetime itself . in a very basic sense for example : ordinary quantum field theory is local , but the fundamental ( diffeomorphism - invariant ) physical observables of quantum gravity are necessarily nonlocal.ordinary quantum field theory takes causality as a fundamental postulate , but in quantum gravity the spacetime geometry , and thus the light cones and the causal structure , are themselves subject to quantum fluctuations.time evolution in quantum field theory is determined by a hamiltonian operator , but for spatially closed universes , the natural candidate for a hamiltonian in quantum gravity is identically zero when acting on physical states.quantum mechanical probabilities must add up to unity at a fixed time , but in general relativity there is no preferred time - slicing on which to normalize probabilities . ordinary quantum field theory is local , but the fundamental ( diffeomorphism - invariant ) physical observables of quantum gravity are necessarily nonlocal . ordinary quantum field theory takes causality as a fundamental postulate , but in quantum gravity the spacetime geometry , and thus the light cones and the causal structure , are themselves subject to quantum fluctuations . time evolution in quantum field theory is determined by a hamiltonian operator , but for spatially closed universes , the natural candidate for a hamiltonian in quantum gravity is identically zero when acting on physical states . quantum mechanical probabilities must add up to unity at a fixed time , but in general relativity there is no preferred time - slicing on which to normalize probabilities . faced with such problems , it is natural to look for simpler models that share the important conceptual features of general relativity while avoiding some of the computational difficulties . general relativity in 2 + 1 dimensions two dimensions of space plus one of time is one such model . as a generally covariant theory of spacetime geometry , ( 2 + 1)-dimensional gravity has the same conceptual foundation as realistic ( 3 + 1)-dimensional general relativity , and many of the fundamental issues of quantum gravity carry over to the lower dimensional setting . at the same time , however , the ( 2 + 1)-dimensional model is vastly simpler , mathematically and physically , and one can actually write down viable candidates for a quantum theory . with a few exceptions , ( 2 + 1)-dimensional solutions are physically quite different from those in 3 + 1 dimensions , and the ( 2 + 1)-dimensional model is not very helpful for understanding the dynamics of realistic quantum gravity . in particular , the theory does not have a good newtonian limit [ 107 , 49 , 94 ] . but for understanding conceptual problems the nature of time , the construction of states and observables , the role of topology and topology change , the relationships among different approaches to quantization the model has proven highly instructive . work on ( 2 + 1)-dimensional gravity dates back to 1963 , when staruszkiewicz first described the behavior of static solutions with point sources . progress continued sporadically over the next twenty years , but the modern rebirth of the subject can be traced to the seminal work of deser , jackiw, t hooft , and witten in the mid-1980s [ 107 , 105 , 106 , 249 , 103 , 277 , 279 ] . over the past twenty years , ( 2 + 1)-dimensional gravity has become an active field of research , drawing insights from general relativity , differential geometry and topology , high energy particle theory , topological field theory , and string theory . as i will explain below , general relativity in 2 + 1 dimensions has no local dynamical degrees of freedom . classical solutions to the vacuum field equations are all locally diffeomorphic to spacetimes of constant curvature , that is , minkowski , de sitter , or anti - de sitter space . broadly speaking , three ways to introduce dynamics have been considered : point particles can be added , appearing as conical most of the earliest papers in the field [ 246 , 105 , 106 , 107 , 249 , 103 ] were investigations of the dynamics of such conical singularities.if a negative cosmological constant is present , black hole solutions can be found [ 41 , 40 ] . for such solutions , dynamics at either the horizon or the boundary at infinity can lead to local degrees of freedom [ 78 , 247 , 59 , 96 , 51 , 24 , 91 ] , although these are certainly not yet completely understood .one can consider nontrivial spatial or spacetime topologies [ 277 , 279 ] . solutions have moduli a finite number of parameters that distinguish among geometrically inequivalent constant curvature manifolds and these can become dynamical . in this paper , i will limit myself to the third case , ( 2 + 1)-dimensional vacuum this review is based in part on a series of lectures in and an earlier review , and much of the material can be found in more detail in a book . there is not yet a comprehensive review of gravitating point particles in 2 + 1 dimensions , although [ 65 , 197 , 195 , 37 , 36 , 199 , 63 , 183 ] will give an overview of some results . several good general reviews of the ( 2 + 1)-dimensional black hole exist [ 75 , 39 ] , although a great deal of the quantum mechanics is not yet understood . most of the earliest papers in the field [ 246 , 105 , 106 , 107 , 249 , 103 ] were investigations of the dynamics of such conical singularities . if a negative cosmological constant is present , black hole solutions can be found [ 41 , 40 ] . for such solutions , dynamics at either the horizon or the boundary at infinity can lead to local degrees of freedom [ 78 , 247 , 59 , 96 , 51 , 24 , 91 ] , although these are certainly not yet completely understood . solutions have moduli a finite number of parameters that distinguish among geometrically inequivalent constant curvature manifolds and these can become dynamical . although string theory is perhaps the most popular current approach to quantum gravity , i will have little to say about it here : while some interesting results exist in 2 + 1 dimensions , almost all of them are in the context of black holes ( see , for example , [ 157 , 170 , 187 , 188 , 189 ] ) . i will also have little to say about ( 2 + 1)-dimensional supergravity , although many of the results described below can be generalized fairly easily , and i will not address the coupling of matter except for a brief discussion in section 5 . throughout , i will use units 16g = 1 and = 1 unless otherwise noted . the first step towards quantizing ( 2 + 1)-dimensional general relativity is to understand the space of classical solutions . one of the principal advantages of working in 2 + 1 dimensions is that for simple enough topologies , this space can be characterized completely and explicitly . indeed , there are several such characterizations , each leading naturally to a different approach to the quantum theory ; by understanding the relationships among these approaches , one can gain important insights into the structure of quantum gravity . in any spacetime , the curvature tensor may be decomposed into a curvature scalar r , a ricci tensor r , and a remaining trace - free , conformally invariant piece , the weyl tensor \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${c_{\mu \nu \rho}}^\sigma$\end{document}. in 2 + 1 dimensions , however , the weyl tensor vanishes identically , and the full curvature tensor is determined algebraically by the remaining pieces : 1\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${r_{\mu \nu \rho \sigma } } = { g_{\mu \rho}}{r_{\nu \sigma } } + { g_{\nu \sigma}}{r_{\mu \rho } } - { g_{\nu \rho}}{r_{\mu \sigma } } - { g_{\mu \sigma}}{r_{\nu \rho } } - { 1 \over 2}({g_{\mu \rho}}{g_{\nu \sigma } } - { g_{\mu \sigma}}{g_{\nu \rho}})r.$$\end{document } this means that any solution of the field equations with a cosmological constant , 2\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${r_{\mu \nu } } = 2\lambda { g_{\mu \nu}},$$\end{document } has constant curvature : the spacetime is locally either flat ( = 0 ) , de sitter ( > 0 ) , or anti - de sitter ( < 0 ) . physically , a ( 2 + 1)-dimensional spacetime has no local degrees of freedom : there are no gravitational waves in the classical theory , and no propagating gravitons in the quantum theory . this absence of local degrees of freedom can be verified by a simple counting argument [ 49 , 94 ] . in n dimensions , the phase space of general relativity is parametrized by a spatial metric at constant time , which has n(n 1)/2 components , and its conjugate momentum , which adds another n(n 1)/2 components . but n of the einstein field equations are constraints rather than dynamical equations , and n more degrees of freedom can be eliminated by coordinate choices . we are thus left with n(n 1 ) 2n = n(n 3 ) physical degrees of freedom per spacetime point . in four dimensions , this gives the usual four phase space degrees of freedom , two gravitational wave polarizations and their conjugate momenta . it is instructive to examine this issue in the weak field approximation . in any dimension , the vacuum field equations in harmonic gauge for a nearly flat metric g = + h take the form 3\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\square{\bar h_{\mu \nu } } = { \mathcal o}({h^2}),\quad \quad { \partial _ \mu } { \bar h^{\mu \nu } } = 0,$$\end{document } where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\bar h_{\mu \nu } } = { h_{\mu \nu } } - { 1 \over 2}{\eta _ { \mu \nu}}{\eta ^{\rho \sigma}}{h_{\rho \sigma}}$\end{document } and indices are raised and lowered with the flat metric . the plane wave solutions of equation ( 3 ) are , to first order , 4\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\bar h_{\mu \nu } } = { \epsilon _ { \mu \nu}}{e^{ik \cdot x}}\quad \quad { \rm{with}}\,{k^2 } = 0\,{\rm{and}}\,{k^\mu}{\epsilon _ { \mu \nu } } = 0.$$\end{document } choosing a second null vector n with n k = 1 and a spacelike unit vector with k m = n m = 0 , we can construct a ( 2 + 1)-dimensional analog of the newman - penrose formalism ; the polarization tensor then becomes 5\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\epsilon _ { \mu \nu } } = a{k_\mu}{k_\nu } + b({k_\mu}{m_\nu } + { k_\nu}{m_\mu } ) + c{m_\mu}{m_\nu},$$\end{document } apparently giving three propagating polarizations . there is , however , a residual symmetry : a diffeomorphism generated by an infinitesimal vector field with = 0 preserves the harmonic gauge condition of equation ( 3 ) while giving a gauge transformation \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\delta { \bar h_{\mu \nu } } = { \partial _ \mu}{\xi _ \nu } + { \partial _ \nu}{\xi _ \mu } - { \eta _ { \mu \nu}}{\partial _ \rho}{\xi ^\rho}$\end{document}. writing 6\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\xi _ \mu } = ( \alpha { k_\mu } + \beta { n_\mu } + \gamma { m_\mu}){e^{ik \cdot x}},$$\end{document } it is easy to check that 7\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\delta { \epsilon _ { \mu \nu } } = 2i\alpha { k_\mu}{k_\nu } + i\gamma ( { k_\mu}{m_\nu } + { k_\nu}{m_\mu } ) + i\beta { m_\mu}{m_\nu}.$$\end{document } the excitations ( 5 ) are thus pure gauge , confirming the absence of propagating degrees of freedom . fortunately , while this feature makes the theory simple , it does not quite make it trivial . a flat spacetime , for instance , can always be described as a collection of patches , each isometric to minkowski space , that are glued together by isometries of the flat metric ; but the gluing is not unique , and may be dynamical . this picture leads to the description of ( 2 + 1)-dimensional gravity in terms of geometric structures . the global geometry of vacuum spacetimes in 2 + 1 dimensions is described mathematically by the theory of geometric structures [ 256 , 62 , 147 ] ( for examples of geometric structures , see ; for a slightly more detailed version of this section , see ) . for simplicity , if the spacetime manifold m is topologically trivial , then by equation ( 1 ) the vacuum field equations imply that ( m , g ) is simply a subset of ordinary minkowski space ( v , ) . if m is topologically nontrivial , it can still be covered by contractible coordinate patches ui , each isometric to v , with the standard minkowski metric on each patch . the geometry is then encoded entirely in the transition functions on the intersections ui uj , which determine how these patches are glued together . since the metrics in ui and uj are identical , these transition functions must be isometries of , that is , elements of the poincar group iso(2 , 1 ) . similarly , if 0 , a vacuum spacetime can be built by patching together pieces of de sitter or anti - de sitter space by appropriate isometries : so(3 , 1 ) for > 0 and so(2 , 2 ) or sl(2 , ) sl(2 , )/2 for < 0 . such a construction is an example of a geometric structure , in the flat case a lorentzian or ( iso(2 , 1),v ) structure . in general , a ( g , x ) manifold is one locally modeled on x , much as an ordinary n - dimensional manifold is modeled on . more precisely , let g be a lie group that acts analytically on some n - manifold x , the model space , and let m be another n - manifold . a ( g , x ) structure on m is then a set of coordinate patches ui for m with coordinates i : ui x taking their values in x and with transition functions \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${g_{ij } } = { \phi _ i } \circ { \phi _ j}^{- 1}\vert{u_i } \cap { u_j}$\end{document } in g. a fundamental ingredient in the description of a ( g , x ) structure is its holonomy group , which can be viewed as a measure of the failure of a single coordinate patch to extend around a closed curve . let m be a ( g , x ) manifold containing a closed path . as illustrated in figure 1 , we can cover with coordinate charts 8\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\phi _ i}:{u_i } \rightarrow x,\quad \quad i = 1 , \ldots , n,$$\end{document } with constant transition functions gj g between ui and ui+1 , i.e. , 9\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{{\phi _ i}\|{u_i } \cap { u_{i + 1 } } = { g_i}\circ { \phi _ { i + 1}}\|{u_i } \cap { u_{i + 1}},\ , } \\ { \;\;{\phi _ n}\|{u_n } \cap { u_1 } = { g_n}\circ { \phi _ 1}\|{u_n } \cap { u_1}. } \\ \end{array}$$\end{document } let us now try to analytically continue the coordinate 1 from the patch u1 to the whole of . we can begin with a coordinate transformation in u2 that replaces 2 by = 2 2 , thus extending 1 to u1 u2 . continuing this process along the curve , with \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\phi _ j}^\prime = { g_1 } \circ \cdots \circ { g_j}_{- 1 } \circ { \phi _ j}$\end{document } if the new coordinate function \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\phi _ n}^\prime = { g_1 } \circ \cdots \circ { g_n}_{- 1 } \circ { \phi _ n}$\end{document } agrees with 1 on un u1 , we will have covered with a single patch . otherwise , the holonomy h( ) = g1 gn measures the obstruction to such a covering . figure 1the curve is covered by coordinate patches ui , with transition functions gi g. the composition g1 the curve is covered by coordinate patches ui , with transition functions gi g. the composition it may be shown that the holonomy of a curve depends only on its homotopy class . in fact , the holonomy defines a homomorphism 10\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$h:{\pi _ 1}(m ) \rightarrow g.$$\end{document } h is not quite uniquely determined by the geometric structure , since we are free to act on the model space x by a fixed element h g , changing the transition functions gi without altering the ( g , x ) structure of m. such a transformation has the effect of conjugating h by h , and it may be shown that h is unique up to such conjugation . the space of holonomies is thus the quotient 11\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{{\mathcal m } = \hom ( { \pi _ 1}(m),g)/\sim , } \\ { { \rho _ 1}\sim{\rho _ 2}\quad \quad { \rm{if}}\,\,{\rho _ 2 } = h\cdot{\rho _ 1}\cdot{h^{- 1}},h \in g. } \\\end{array}$$\end{document } note that if we pass from m to its universal covering space \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\widetilde m$\end{document } , we will no longer have noncontractible closed paths , and 1 will be extendible to all of \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\widetilde m$\end{document}. the resulting map \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$d:\widetilde m \to x$\end{document } is called the developing map . at least in simple examples , d embodies the classical geometric picture of development as unrolling for instance , the unwrapping of a cylinder into an infinite strip . the holonomies of the geometric structure in ( 2 + 1)-dimensional gravity are examples of diffeomorphism - invariant observables , which , as we shall see below , are closely related to the wilson loop observables in the chern - simons formulation . it is important to understand to what extent they are complete that is , to what extent they determine the geometry . it is easy to see one thing that can go wrong : if we start with a flat three - manifold m and simply cut out a ball , we can obtain a new flat manifold without affecting the holonomy . this is a rather trivial change , though , and we would like to know whether it is the only problem . for the case of a vanishing cosmological constant , mess has investigated this question for spacetimes with topologies . he shows that the holonomy group determines a unique maximal spacetime m specifically , a domain of dependence of a spacelike surface . mess also demonstrates that the holonomy group h acts properly discontinuously on a region w v of minkowski space , and that m can be obtained as the quotient space w / h . this quotient construction can be a powerful tool for obtaining a description of m in reasonably standard coordinates , for instance in a time - slicing by surfaces of constant mean curvature . similar results hold for anti - de sitter structures . some instructive examples of the construction of spacetimes with < 0 from holonomies are given in . for de sitter structures , on the other hand , the holonomies do not uniquely determine the geometry . an explicit example of the resulting ambiguity has been given by ezawa for the case of a topology t ( see also section 4.5 of ) . a similar ambiguity occurs for ( 2 + 1)-dimensional gravity with point particles , where , as matschull has emphasized , it may imply a physical difference between the metric and chern - simons formulations of ( 2 + 1)-dimensional gravity . we close this section with a partial description of the space of solutions of the vacuum einstein field equations on a manifold , where is a compact genus g two - manifold , that is , a surface with g handles . the fundamental group of such a spacetime , 1(m ) 1( ) , is generated by g pairs of closed curves with the single relation 12\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${a_1}{b_1}{a_1}^{- 1}{b_1}^{- 1}{a_2}{b_2}{a_2}^{- 1}{b_2}^{- 1 } \ldots { a_g}{b_g}{a_g}^{- 1}{b_g}^{- 1 } = 1.$$\end{document } by equation ( 11 ) , the space of holonomies is the space of homomorphisms from 1( ) to g ( where g is iso(2 , 1 ) for = 0 , so(3 , 1 ) for > 0 , or so(2 , 2 ) for < 0 ) modulo overall conjugation . for g > 1 , this space of homomorphisms has dimension 12 g 12 : 1( ) has 2 g generators and one relation , and the identification by conjugation leaves 2 g 2 choices of elements of a six - dimensional group g1 . there are two subtleties that prevent the space ( 11 ) from being the exact moduli space of solutions of the vacuum field equations . first , as noted above , the holonomies do not always determine a unique geometric structure . in particular , for > 0 one may need an additional discrete variable to specify the geometry . second , not all homomorphisms from 1( ) to g give geometric structures that correspond to smooth manifolds . the space of homomorphisms ( 11 ) is not connected , and , in general , only one connected component gives our desired geometry . even once these caveats are taken into account , though , we still have a ( 12 g 12)-dimensional space of solutions that can , in principle , be described completely . the formalism of geometric structures provides an elegant description of vacuum spacetimes in 2 + 1 dimensions , but it is rather remote from the usual physicist s approach . in particular , the einstein - hilbert action is nowhere in sight , and even the metric makes only a limited appearance . fortunately , the description is closely related to the more familiar first - order chern - simons formalism [ 108 , 277 , 279 , 2 ] , which , in turn , can connect us back to the standard metric formalism . the first - order formalism takes as its fundamental variables an orthonormal frame ( triad or dreibein ) \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${e_\mu}^a$\end{document } , which determines a metric \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${g_{\mu \nu } } = { \eta _ { ab}}{e_\mu}^a{e_\nu}^b$\end{document } , and a spin connection \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\omega _ \mu}^{ab}$\end{document}. as in the palatini formalism , e and are treated as independent quantities . in terms of the one - forms 13\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${e^a } = { e_\mu}^ad{x^\mu},\quad \quad { \omega ^a } = { 1 \over 2}{\epsilon ^{abc}}{\omega _ { \mu bc}}d{x^\mu},$$\end{document } the first - order action takes the form 14\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$i = 2\int\nolimits_m { \left [ { { e^a } \wedge \left({d{\omega _ a } + { 1 \over 2}{\epsilon _ { abc}}{\omega ^b } \wedge { \omega ^c } } \right ) + { \lambda \over 6}{\epsilon _ { abc}}{e^a } \wedge { e^b } \wedge { e^c } } \right]},$$\end{document } with euler - lagrange equations 15\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${t_a } = d{e_a } + { \epsilon _ { abc}}{\omega ^b } \wedge { e^c } = 0,$$\end{document } 16\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${r_a } = d{\omega _ a } + { 1 \over 2}{\epsilon _ { abc}}{\omega ^b } \wedge { \omega ^c } = - { \lambda \over 2}{\epsilon _ { abc}}{e^b } \wedge { e^c}.$$\end{document } the first of these implies that the connection is torsion - free , and , if e is invertible , that has the standard expression in terms of the triad . given such a spin connection , equation ( 16 ) is then equivalent to the standard einstein field equations . the action ( 14 ) has two sets of invariances , the local lorentz transformations 17\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\delta { e^a } = { \epsilon ^{abc}}{e_b}{\tau _ c},\quad \quad \delta { \omega ^a } = d { \tau ^a } + { \epsilon ^{abc}}{\omega _ b}\tau_c,$$\end{document } and the local translations 18\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\delta { e^a } = d{\rho ^a } + { \epsilon ^{abc}}{\omega _ b}{\rho _ { c,}}\;\;\delta { \omega ^a } = - \lambda { \epsilon ^{abc}}{e_b}{\rho _ c}.$$\end{document } provided the triad e is invertible , the latter are equivalent to diffeomorphisms on a shell ; more precisely , the combination of transformations with parameters = e and = is equivalent to the diffeomorphism generated by the vector field . the invertibility condition for e is important ; if it is dropped , the first - order formalism is no longer quite equivalent to the metric formalism . as first noted by achcarro and townsend and further developed by witten [ 277 , 279 ] , the first - order action ( 14 ) is equivalent to that of a chern - simons theory . the relevant gauge group the group g of the geometric structure is then the poincar group iso(2 , 1 ) , with standard generators \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal j^ a}$\end{document } and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal p^a}$\end{document } and commutation relations 19\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$[{{\mathcal j}^a},{{\mathcal j}^b } ] = { \epsilon ^{abc}}{{\mathcal j}_c},\quad \quad[{{\mathcal j}^a},{{\mathcal p}^b } ] = { \epsilon ^{abc}}{{\mathcal p}_c}\quad \quad [ { { \mathcal p}^a},{{\mathcal p}^b } ] = 0.$$\end{document } the corresponding gauge potential is 20\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$a = { e^a}{{\mathcal p}_a } + { \omega ^a}{{\mathcal j}_a}.$$\end{document } if one defines a bilinear form ( or trace ) 21\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\rm{tr}}({{\mathcal j}^a}{{\mathcal p}^b } ) = { \eta ^{ab}},\quad{\rm{tr}}({{\mathcal j}^a}{{\mathcal j}^b } ) = { \rm{tr}}({{\mathcal p}^a}{{\mathcal p}^b } ) = 0,$$\end{document } it is straightforward to show that the action ( 14 ) can be written as 22\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${i_{{\rm{cs}}}}[a ] = { k \over { 4\pi}}\int\nolimits_m { { \rm{tr } } } \left({a \wedge da + { 2 \over 3}a \wedge a \wedge a } \right),$$\end{document } with k = 1/(4 g ) . equation ( 22 ) may be recognized as the standard chern - simons action for the group iso(2 , 1 ) . e/ together constitute an so(2 , 1 ) so(2 , 1 ) gauge potential , with a chern - simons action 23\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$i[{a^{(+)}},{a^{(- ) } } ] = { i_{{\rm{cs}}}}[{a^{(+ ) } } ] - { i_{{\rm{cs}}}}[{a^{(-)}}]$$\end{document } that is again equivalent to equation ( 14 ) , provided we set k = (4 g ) . if > 0 , the complex one - form \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${a^a } = { \omega ^a } + i\sqrt \lambda { e^a}$\end{document } may be viewed as an sl(2 , ) gauge potential , whose chern - simons action is again equivalent to the first - order gravitational action . for any value of , it is easily checked that the transformations ( 17 ) are just the gauge transformations of a. vacuum general relativity in 2 + 1 dimensions is thus equivalent again up to considerations of the invertibility of e to a gauge theory . we can now connect the first - order formalism to the earlier description of geometric structures . the field equations coming from the action ( 22 ) are simply 24\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$f[a ] = da + a \wedge a = 0,$$\end{document } implying that the field strength of the gauge potential a vanishes , i.e. , that a is a flat connection . such a connection is completely determined by its holonomies , that is , by the wilson loops 25\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${u_\gamma } = p\exp \left({- \int\nolimits_\gamma a } \right)$$\end{document } around closed noncontractible curves . this use of the term holonomy is somewhat different from that of section 2.2 , but the two are equivalent . indeed , any ( g , x ) structure on a manifold m determines a corresponding flat g bundle : we simply form the product g uj in each patch , giving the local structure of a g bundle , and use the transition functions gij of the geometric structure to glue the fibers on the overlaps . the holonomy group of this flat bundle can be shown to be isomorphic to the holonomy group of the geometric structure , and for ( 2 + 1)-dimensional gravity , the flat connection constructed from the geometric structure is that of the chern - simons theory . an explicit construction may be found in section 4.6 of ; see also [ 7 , 263 ] . the first - order action allows us an additional step that was unavailable in the geometric structure formalism we can compute the symplectic structure on the space of solutions . the basic poisson brackets follow immediately from the action : 26\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\{{e_i}^a(x),{\omega _ j}^b(x^{\prime})\ } = { 1 \over 2}{\eta ^{ab}}{\epsilon _ i}_j{\delta ^2}(x - x^{\prime}).$$\end{document } the resulting brackets among the holonomies have been evaluated by nelson , regge , and zertuche [ 210 , 211 ] for < 0 , for which the two sl(2 , ) factors in the gauge group g may be taken to be independent . the brackets are nonzero only for holonomies of curves that intersect , and can be written in terms of holonomies of rerouted curves ; symbolically , 27\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\left\{{\left . { } \right\rangle , \left\langle { } \right . } \right\ } = \pm { 1 \over { 4\ell}}\epsilon ( p)\left({\left . { } \right\rangle \left\langle { } \right . \right),$$\end{document } where (p ) is the oriented intersection number at the point p that the curves cross . the composition of loops implicit in the brackets ( 27 ) makes it difficult to find small closed subalgebras of the sort needed for quantization . however , nelson and regge have succeeded in constructing a small but complete ( actually overcomplete ) set of holonomies on a surface of arbitrary genus that form a closed algebra [ 213 , 212 ] , and loll has found a complete set of configuration space variables . by generalizing a discrete combinatorial approach to chern - simons theory due to fock and rosly and alekseev et al . [ 3 , 4 , 5 ] , several authors have further explored the quantum group structure of these brackets , which can be expressed in terms of the quantum double of the lorentz group [ 37 , 36 , 61 , 201 ] . it is also interesting that the symplectic structure obtained in this way is closely related to the symplectic structure on the abstract space of loops on first discovered by goldman [ 145 , 146 ] . we next turn to a more traditional approach to classical general relativity , the conventional metric formalism in the space / time splitting of arnowitt , deser , and misner . as moncrief and hosoya and nakao have shown , this metric formalism can also be used to give a full description of the solutions of the vacuum field equations , at least for spacetimes with the topology . we start with the adm decomposition of the spacetime metric g , 28\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$d{s^2 } = { n^2}d{t^2 } - { g_{ij}}(d{x^i } + { n^i}dt)(d{x^j } + { n^j}dt),$$\end{document } as illustrated in figure 2 . the action then takes the usual form2 29\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${i_{{\rm{grav } } } } = \int { { d^3}x\sqrt { { - ^{(3)}}g } } { ( ^{(3)}}r - 2\lambda ) = \int { dt } \int\nolimits_\sigma { { d^2}x } ( { \pi ^{ij}}{\dot g_{ij } } - { n^i}{{\mathcal h}_i } - n{\mathcal h}),$$\end{document } with canonical momentum \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\rm{\pi}}^{ij } } = \sqrt g ( { k^{ij } } - { g^{ij}}k)$\end{document } and the momentum and hamiltonian constraints 30\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${{\mathcal h}_i } = - 2{\nabla _ j}{\pi ^j}i,\quad{\mathcal h } = { 1 \over { \sqrt g}}{g_{ij}}{g_{kl}}({\pi ^{ik}}{\pi ^{jl } } - { \pi ^{ij}}{\pi ^{kl } } ) - \sqrt g ( r - 2\lambda).$$\end{document } figure 2the adm decomposition is based on the lorentzian version of the pythagoras theorem . the adm decomposition is based on the lorentzian version of the pythagoras theorem . to solve the constraints , we can choose the york time - slicing , in which the mean ( extrinsic ) curvature is used as a time coordinate , \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$ - k = { g_{ij}}{\pi ^{ij}}/\sqrt g = t$\end{document}. andersson et al . have shown that this is a good global coordinate choice for classical solutions of the vacuum field equations . we next select a useful parametrization of the spatial metric and momentum . up to a diffeomorphism , any two - metric on can be written in the form [ 1 , 121 ] 31\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${g_{ij } } = { e^{2\lambda}}{\bar{g}_{ij}}({m_\alpha}),$$\end{document } where ij(m ) are a finite - dimensional family of metrics of constant curvature k ( k = 1 for the two - sphere , 0 for the torus , and 1 for spaces of genus g > 1 ) . these standard metrics are labeled by a set of moduli m that parametrize the riemann moduli space of . as in section 2.2 , such constant curvature metrics can be described in terms of a geometric structure for genus g > 1 an ( , psl(2 , ) ) structure with moduli parametrizing the homomorphisms ( 10 ) . we can count these just as in section 2.2 ; now , since psl(2 , ) is three - dimensional , we find that a constant negative curvature surface of genus g > 1 is described by 6 g 6 parameters . the corresponding decomposition of the conjugate momentum is described in : up to a diffeomorphism , the trace - free part of can be written as a holomorphic quadratic differential p , that is , a transverse traceless tensor with respect to the covariant derivative compatible with ij . the space of such quadratic differentials parametrizes the cotangent space of the moduli space , and the reduced phase space becomes , essentially , the cotangent bundle of the moduli space . with the decomposition of , the momentum constraints i = 0 become trivial , while the hamiltonian constraint becomes an elliptic differential equation that determines the scale factor in equation ( 31 ) as a function of ij and p , 32\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\bar \delta \lambda - \frac{1}{4}({t^2 } - 4\lambda ) { e^{2\lambda } } + \frac{1}{2}[{\bar g^ { - 1}}{\bar g_{ij}}({m_\alpha } ) { \bar g_{kl}}({m_\alpha } ) { p^{ik}}({p^\alpha } ) { p^{jl}}]{e^ { - 2\lambda } } - \frac{k}{2 } = 0,$$\end{document } where p are the momenta conjugate to the moduli , 33\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${p^\alpha } = \int\nolimits_\sigma { { d^2}x{p^{ij}}{\partial \over { \partial { m_\alpha}}}{{\bar g}_{ij}}.}$$\end{document } the theory of elliptic equations ensures that equation ( 32 ) determines a unique scale factor . the action ( 29 ) then simplifies to a reduced phase space action , involving only the physical degrees of freedom , 34\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${i_{{\rm{grav } } } } = \int { dt } \left({{p^\alpha}{{d{m_\alpha } } \over { dt } } - h(m , p , t ) } \right),$$\end{document } with a time - dependent hamiltonian 35\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$h = \int\nolimits_{{\sigma _ t } } { { d^2}x } \sqrt { \bar{g } } { e^{2\lambda ( m , p , t)}}.$$\end{document } the classical poisson brackets can be read off directly from equation ( 34 ) : 36\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\ { { m_\alpha } , { p^\beta } \ } = \delta _ \alpha ^\beta , \quad \quad \ { { m_\alpha } , { m_\beta } \ } = \ { { p^\alpha } , { p^\beta } \ } = 0.$$\end{document } three - dimensional gravity again reduces to a finite - dimensional system , albeit one with a complicated time - dependent hamiltonian . the physical phase space is parametrized by ( m,p ) , which may be viewed as coordinates for the cotangent bundle of the moduli space of . for a surface of genus g > 1 , this gives us 12 g 12 degrees of freedom , matching the results of section 2.2 . if = 0 , this correspondence can be made more explicit : for g = iso(2 , 1 ) and m , the space ( 11 ) of geometric structures is itself a cotangent bundle , whose base space is the space of hyperbolic structures on . this follows from the fact that the group iso(2 , 1 ) is the cotangent bundle of so(2 , 1 ) . concretely , in the first - order formalism of section 2.3 , the curvature equation ( 16 ) with = 0 implies that is a flat so(2 , 1 ) connection ; and if (s ) is a curve in the space of such flat connections , the tangent vector e = d(s)/ds satisfies the torsion equation ( 17 ) . for 0 , i know of no such direct correspondence , and the general relationship between the adm and first - order solutions seems less transparent . discrete approximations to general relativity have existed for decades . in 2 + 1 dimensions , this follows from the peculiar nature of the field equations in three dimensions : as discussed above , any vacuum solution can be patched together from finite pieces of constant curvature spacetime , and the dynamics occurs only in the patching . the standard discrete approach to classical general relativity is regge calculus , initially developed for ( 3 + 1)-dimensional gravity but extendible to arbitrary dimensions . classical regge calculus in 2 + 1 dimensions was investigated by rocek and williams , who showed that it gave exact results for point particle scattering . the first discrete formulation designed explicitly for 2 + 1 dimensions was developed by t hooft et al . this approach has been used mainly to understand point particle dynamics , but recent progress has allowed a general description of topologically nontrivial compact spaces . t hooft s hamiltonian lattice model is based on the metric formalism , and starts with a piecewise flat cauchy surface tessellated by flat polygons , each carrying an associated frame . the einstein field equations with = 0 then imply that edges of polygons move at constant velocities and that edge lengths may change , subject to a set of consistency conditions . one obtains a dynamical description parametrized by a set of lengths and rapidities , which turn out to be canonically conjugate . complications occur when an edge shrinks to zero length or collides with a vertex , but these are completely understood . the resulting structure can be simulated on a computer , providing a powerful method for visualizing classical evolution in 2 + 1 dimensions . a related first - order hamiltonian lattice model has been studied by waelbroeck et al . [ 266 , 267 , 268 , 270 ] . this model is a discretized version of the first - order formalism of section 2.3 , with triads assigned to faces of a two - dimensional lattice and lorentz transformations assigned to edges . in particular , for a spacetime m , one can choose a lattice that is simply a 4g - sided polygon with edges identified ; the resulting spacetime can be visualized as a polygonal tube cut out of minkowski spacetime , with corners lying on straight worldlines and edges identified pairwise . this reproduces the quotient space picture discussed by mess in the context of geometric structures . with a different gauge choice , waelbroeck s model is classically equivalent to t hooft s , but the two models are related by a nonlocal change of variables , and may not be equivalent quantum mechanically . much of the recent work on lattice formulations of ( 2 + 1)-dimensional gravity have centered on spin foams and on random triangulations , both inherently quantum mechanical . it is worth noting here , though , that recent work on diffeomorphisms in spin foam models may permit a classical description quite similar to that of waelbroeck . up to now , i have avoided discussing an important discrete symmetry of general relativity on topologically nontrivial spacetimes . the description of a solution of the field equations in terms of holonomies ( sections 2.2 and 2.3 ) or moduli ( section 2.4 ) is invariant under infinitesimal diffeomorphisms , and hence under small diffeomorphisms , those that can be smoothly deformed to the identity . but if the spacetime manifold is topologically nontrivial , its group of diffeomorphisms may not be connected : m may admit the group of such large diffeomorphisms ( modulo small diffeomorphisms ) , \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\mathcal d(m)$\end{document } , is called the mapping class group of m ; for the torus t , it is also known as the modular group . the archetype of a large diffeomorphism is a dehn twist of a torus , which may be described as the operation of cutting t along a circumference to obtain a cylinder , twisting one end by 2 , and regluing . similar transformations exist for any closed surface , and in fact the dehn twists around generators of 1( ) generate \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\mathcal d(\sigma)$\end{document } [ 57 , 56 ] . it is easy to see that the mapping class group of a spacetime m acts on 1(m ) , and therefore on the holonomies of section 2.2 . as diffeomorphisms , elements of the mapping class group also acts on the constant curvature metrics ij , and hence on the moduli of section 2.4 . classically , geometries that differ by actions of \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal d}(m)$\end{document } are exactly equivalent , so the true space of vacuum solutions for a spacetime with the topology is really \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\mathcal m/\mathcal d(m)$\end{document } , where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal m}$\end{document } is the moduli space ( 11 ) . quantum mechanically , it is not clear whether one should impose mapping class group invariance on states or whether one should merely treat \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal d}(m)$\end{document } as a symmetry under which states may transform nontrivially ( see , for instance , ) . in 2 + 1 dimensions , though , there seems to be a strong argument in favor of treating the mapping class group as a genuine invariance , as follows . using the chern - simons formalism , one can compute the quantum amplitude for the scattering of a point particle off another particle , a black hole , or a handle . in each case , it is only when one imposes invariance under the mapping class group that one recovers the correct classical limit . it may still be that simple enough representations of \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal d}(m)$\end{document } lead to sensible physical results , but it is at least clear that the mapping class group can not be ignored . the simplest nontrivial vacuum cosmology occurs for a spacetime with the topology t , where t is the two - dimensional torus . this case is in some ways exceptional for example , the standard metric ij of equation ( 31 ) is flat rather than hyperbolic but it is also simple enough that a great deal can be done explicitly . later in this review , the torus universe will be a canonical test of quantization ; here we review classical aspects . the problem of finding the classical solutions , as well as an approach to the quantization , was , i believe , first discussed by martinec . i refer the reader to [ 76 , 81 , 87 , 86 ] for further details . a similarly detailed analysis may be possible when the spatial topology is that of a klein bottle ( see , for instance , ) but so far , this and other nonorientable examples have been studied in much less detail . for simplicity , the group g of section 2.2 , or , equivalently , the gauge group in the chern - simons formalism of section 2.3 , is then so(2 , 2 ) . the fundamental group 1( t ) has two generators , [ 1 ] and [ 2 ] , satisfying a single relation similar to equation ( 12 ) : 37\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$[{\gamma _ 1 } ] \cdot [ { \gamma _ 2 } ] = [ { \gamma _ 2 } ] \cdot [ { \gamma _ 1}].$$\end{document } the holonomy group ( 11 ) is therefore generated by two commuting so(2 , 2 ) matrices , unique up to overall conjugation . it is a bit more convenient to describe the holonomies as elements of the covering group sl(2 , ) sl(2 , ) . let [a ] denote the two sl(2 , ) holonomies corresponding to the curve a . an sl(2 , ) matrix s is called hyperbolic , elliptic , or parabolic according to whether \|tr s\| is greater than , equal to , or less than 2 , and the space of holonomies correspondingly splits into nine sectors . it may be shown that only the hyperbolic - hyperbolic sector corresponds to a spacetime in which the t slices are spacelike [ 117 , 119 , 182 , 209 ] . by suitable overall conjugation , the two generators of the holonomy group in this sector can then be taken to be 38\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\rho ^ \pm } [ { \gamma _ 1 } ] = \left ( { \begin{array}{{20}c } { { e^{r_1^ \pm /2 } } } & 0\\ 0 & { { e^ { - r_1^ \pm /2}}}\\ \end{array } } \right),\quad { \rho ^ \pm } [ { \gamma _ 2 } ] = \left ( { \begin{array}{{20}c } { { e^{r_2^ \pm /2 } } } & 0\\ 0 & { { e^ { - r_2^ \pm /2}}}\\ \end{array } } \right),$$\end{document } where the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_a^ \pm$\end{document } are four arbitrary parameters . note that this gives the right counting : the riemann moduli space of the torus is two dimensional , so from section 2.4 we expect a four - dimensional space of solutions . to obtain the corresponding geometry , we can use the quotient space construction of section 2.2 . note first that three - dimensional anti - de sitter space can be represented as the submanifold of flat ( with coordinates ( x1 , x2 , t1 , t2 ) and metric \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$d{s^2 } = dx_1 ^ 2 + dx_2 ^ 2 - dt_1 ^ 2 - dt_2 ^ 2$\end{document } ) defined by the condition that 39\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\det \vert { \bf{x}}\vert = 1,\quad { \bf{x } } = { 1 \over \ell } \left({\begin{array}{{20}c } { { x_1 } + { t_1 } } & { { x_2 } + { t_2 } } \\ { - { x_2 } + { t_2 } } & { { x_1 } - { t_1 } } \\ \end{array } } \right).$$\end{document } this gives an isometry between ads3 and the group manifold of sl(2 , ) . the quotient of ads3 by the holonomy group ( 38 ) may now be obtained by allowing the [a ] to act on x by left multiplication and the [a ] to act by right multiplication . it is straightforward to show that the resulting induced metric is 40\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$ \begin{array}{{20}c}{d{s^2 } = d{t^2 } - { { { \ell ^2 } } \over 4}\left [ { { { ( r_1^ + ) } ^2 } + { { ( r_1^ -)}^2 } + 2r_1^ + r_1^ - \cos { { 2 t } \over \ell } } \right]d{x^2 } } \\ { \quad \quad \quad \;\ ; - { { { \ell ^2 } } \over 4}\left [ { r_1^ + r_2^ + + r_2^ - r_2^ - + ( r_1^ + r_2^ - + r_1^ - r_2^ + ) \cos { { 2 t } \over \ell } } \right]dx\;dy } \\ { \quad \quad \quad \;\ ; - { { { \ell ^2 } } \over 4}\left [ { { { ( r_2^ + ) } ^2 } + { { ( r_2^ -)}^2 } + 2r_2^ + r_2^ - \cos { { 2 t } \over \ell } } \right]d{y^2 } , } \\ the triad may be read off directly from equation ( 40 ) , and it is easy to solve equation ( 15 ) for the spin connection . the resulting chern - simons connections a of equation ( 23 ) are flat , and their holonomies reproduce the holonomies ( 38 ) of the geometric structure we began with . to relate these expressions to the adm formalism of section 2.4 , we must first find the slices of constant extrinsic curvature t. for the metric ( 40 ) , the extrinsic curvature of a slice of constant t is \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$t = - { 2 \over \ell}\cot { { 2 t } \over \ell}$\end{document } , which is independent of x and y. a constant t slice is thus also a slice of constant york time . the standard flat metric on t , the genus one version of the standard metric ( 31 ) , is 41\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$d{\sigma ^2 } = { \tau _ 2}^{- 1}\vert dx + \tau \;dy{\vert ^2},$$\end{document } where = 1 + i2 is the modulus . comparing ( 40 ) , we see that a slice of constant t has a modulus 42\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\tau = \left({r_1^ - { e^{it/\ell } } + r_1^ + { e^{- it/\ell } } } \right)\left({r_2^ - { e^{it/\ell } } + r_2^ + { e^{- it/\ell } } } \right){.^{- 1}}$$\end{document } the conjugate momentum p = p + ip can be similarly computed from equation ( 33 ) , 43\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$p = - { { i\ell } \over { 2\sin { { 2 t } \over \ell}}}\left({r_2^ + { e^{it/\ell } } + r_2^ - { e^{- it/\ell } } } \right){,^2}$$\end{document } while the adm hamiltonian h of equation ( 35 ) becomes 44\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$h = - { { { \ell ^2 } } \over 4}\sin { { 2 t } \over \ell}\left({r_1^ - { r_2}^ + - r_1^ + { r_2}^ - } \right ) = { \left({{t^2 } + { 4 \over { { \ell ^2 } } } } \right)^{- 1/2}}{[{\tau _ 2}^2p\bar p ] ^{1/2}}.$$\end{document } in the limit of vanishing , these relations go over to those of . to quantize this system , we will need the classical poisson brackets , which can be obtained from equation ( 26 ) : 45\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\{{r_1}^ \pm , { r_2}^ \pm \ } = \mp { 1 \over \ell},\;\;\;\;\{{r_a}^ + , { r_b}^ - \ } = 0.$$\end{document } these , in turn , determine the brackets among the moduli and momenta and p , 46\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\{\tau , \bar{p } \ } = \{\bar \tau , p\ } = 2,\;\;\;\;\{\tau , p\ } = \{\bar{\tau } , \bar{p } \ } = 0,$$\end{document } a result consistent with equation ( 36 ) . it may be shown that the version of hamilton s equations of motion coming from these brackets reproduces the time dependence ( 42 ) of the moduli ; see [ 87 , 135 ] for details . the poisson brackets among the traces of the holonomies ( 38 ) are also easy to compute . if we let 47\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{r_1^ \pm = { 1 \over 2}{\rm{tr}}\;{\rho ^ \pm}[{\gamma _ 1 } ] = \cosh { { r_1^ \pm } \over 2 } , } \\{r_2^ \pm = { 1 \over 2}{\rm{tr}}\;{\rho ^ \pm}[{\gamma _ 2 } ] = \cosh { { r_2^ \pm } \over 2 } , } \\{r_{12}^ \pm = { 1 \over 2}{\rm{tr}}\;{\rho ^ \pm}[{\gamma _ 1 } \cdot { \gamma _ 2 } ] = \cosh { { ( r_1^ \pm + r_2^ \pm ) } \over 2 } , } \\\end{array}$$\end{document } it is not hard to check that 48\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\{r_1^ \pm , r_2^ \pm \ } = \mp { 1 \over { 4\ell}}(r_{12}^ \pm - r_1^ \pm r_2^ \pm)\;\;\;{\rm{and}}\;{\rm{cyclical}}\;{\rm{permutations,}}$$\end{document } reproducing the poisson algebra of nelson , regge , and zertuche . this group is generated by two dehn twists , which act on 1(t ) by 49\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{s:{\gamma _ 1 } \rightarrow \gamma _ 2^{- 1 } , } & { { \gamma _ 2 } \rightarrow { \gamma _ 1 } , } \\ { t:{\gamma _ 1 } \rightarrow { \gamma _ 1 } \cdot { \gamma _ 2 } , } & { { \gamma _ 2 } \rightarrow { \gamma _ 2}. } \\\end{array}$$\end{document } these transformations act on the parameters \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_a^ \pm$\end{document } and the adm moduli and momenta as 50\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{s : r_1^ \pm \rightarrow r_2^ \pm } & { r_2^ \pm \rightarrow - r_1^ \pm } & { \tau \rightarrow - { 1 \over \tau } } & { p \rightarrow { { \bar \tau}^2}p } \\ { t : r_1^ \pm \rightarrow r_1^ \pm + r_2^ \pm } & { r_2^ \pm \rightarrow r_2^ \pm } & { \tau \rightarrow \tau + 1 } & { p \rightarrow p. } \\ \end{array}$$\end{document } these transformations are consistent with the relationships between the adm and holonomy variables , and that they preserve all poisson brackets . for a torus universe with zero or positive cosmological constant , i refer the reader to for details . for the torus universe of the preceding section 2.7 the area of a slice of constant t is essentially the hamiltonian ( 44 ) ; it increases from 0 at t = 0 to a maximum at t = and then shrinks to zero at t = /2 at the big bang and big crunch the modulus ( 42 ) is purely real , 2 = 0 . this means that even apart from the crunch in volume , the geometry is singular : a real value of represents a torus that has collapsed to a line . for 0 , the final big crunch disappears , and the torus universe expands forever from an initial big bang . it is not hard to check that as time increases , the modulus ( 42 ) moves along a semicircle in the upper half of the complex plane , with a center on the real axis . such a curve is a geodesic in the natural weil - petersson ( or poincar ) metric on the torus moduli space [ 159 , 135 ] . because of the invariance under the mapping class group ( 50 ) , however , the true physical motion in the moduli space of the torus the space of physical configurations with the large diffeomorphisms modded out is much more complicated ; there are arbitrarily long geodesics , and the flow is , in fact , ergodic . for spacetimes with being a surface of genus g > 1 , no explicit metrics analogous to equation ( 40 ) are known , except for the special case of solutions with constant moduli . the problem is in part that no simple form such as equation ( 41 ) for the standard constant curvature metrics exists , and in part that the adm hamiltonian becomes a complicated , nonlocal function of the moduli . for the case of an asymptotically flat genus g space , some interesting progress has been made by krasnov ; i do not know whether these methods can be extended to the spatially closed case . one can write down the holonomies of the geometric structure for a higher genus surface , of course though even there , it is nontrivial to ensure that they represent spacetimes with spacelike genus g slices but to a physicist , these holonomies in themselves give fairly little insight into the dynamics . in principle , the adm and chern - simons approaches might be viewed as complimentary : as moncrief has pointed out , one could evaluate the holonomies in terms of adm variables in a nice time - slicing , set these equal to constants , and thereby solve the adm equations of motion . in practice , though , this approach seems intractable except for the genus one case . for = 0 , it may be possible to extract a useful physical picture from the geometrical results of , which relate holonomies to the structure of the initial singularity and the asymptotic future geometry , but the implications have not yet been explored in any depth . the singular behavior of the torus universe carries over to higher genus : spacetimes with < 0 expand from a big bang and recollapse in a big crunch , while those with 0 expand forever [ 200 , 20 ] . moreover , the degeneration of the spatial geometry at the initial singularity carries over to the higher genus case [ 200 , 55 ] . by introducing a global cosmological time and exploiting recent results in two- and three - dimensional geometry , benedetti and guadagnini have shown that when = 0 , a set of parameters describing the initial singularity and a second set describing the geometry in the asymptotic future together completely determine the spacetime . it seems likely that these two sets are canonically conjugate , and a better understanding of the symplectic structure could be useful for quantum gravity . the reader may well have decided that for an author reviewing quantum gravity , i have spent an inordinate amount of time on the classical theory . there is a good reason for this , though : each of the approaches described in the preceding sections leads very naturally to an approach to quantization , which is now with a few twists fairly straightforward . indeed , the main reason that 2 + 1 dimensions offer such an attractive setting for quantum gravity is that the classical solutions can be completely described by a finite set of parameters . such a description effectively reduces quantum gravity to quantum mechanics , allowing us to evade the complications of quantum field theory . this is not to imply that all approaches to quantum gravity simplify the wheeler - dewitt equation , for example , apparently does not but it allows us to explore at least a few approaches in depth . perhaps the simplest approach to quantum gravity in 2 + 1 dimensions [ 66 , 160 ] begins with the reduced phase space action ( 34 ) , which describes a finite - dimensional system of physical degrees of freedom , albeit one with a complicated , time - dependent hamiltonian . we know , at least in principle , how to quantize such a system : we simply replace the poisson brackets ( 36 ) with commutators , 51\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$[{\hat{m}_\alpha},{\hat{p}^\beta } ] = i\hbar \delta _ \alpha ^\beta,$$\end{document } represent the momenta as derivatives , p = i/m , and choose our wave functions to be square integrable functions (m , t ) that evolve according to the schrdinger equation 52\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$i\hbar { { \partial \psi ( { m_\alpha},t ) } \over { \partial t } } = \hat h \psi ( { m_\alpha},t),$$\end{document } where the hamiltonian is obtained from equation ( 35 ) in a suitable operator ordering . invariance under the mapping class group of section 2.6 can be incorporated by demanding that (m , t ) transform under a representation of \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal d}(m)$\end{document}. a similar requirement may help determine the operator ordering in the hamiltonian operator [ 68 , 70 ] , although some ambiguities will remain . for spatial surfaces of genus g > 1 , the complexity of the constraint ( 32 ) seems to make this approach to quantization impractical . a perturbative expression for may still exist , though , as discussed in [ 217 , 218 ] , and the gauss map has been proposed as a useful tool . for genus one , the classical hamiltonian ( 44 ) becomes , up to operator ordering ambiguities , 53\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\hat{h } = { \left({{t^2 } + { 4 \over { { \ell ^2 } } } } \right)^{- 1/2}}\delta _ 0^{1/2},\quad { \delta _ 0 } = - { \tau _ 2}^2\left({{{{\partial ^2 } } \over { \partial { \tau _ 1}^2 } } + { { { \partial ^2 } } \over { \partial { \tau _ 2}^2 } } } \right),$$\end{document } where 0 is the ordinary scalar laplacian for the constant negative curvature poincar metric on moduli space , and one chooses the positive square root in order to have a hamiltonian that is bounded below . this laplacian is invariant under the modular transformations ( 50 ) , and its invariant eigenfunctions , the weight zero maass forms , have been studied extensively by mathematicians ( see , for example , ) . the behavior of the corresponding wave functions has been explored by puzio , who argues that they are well - behaved and nonsingular at the boundaries of moduli space . such behavior is relevant to the question of how quantum gravity handles singularities : the degeneration of the torus geometry at the big bang , described in section 2.8 , corresponds to an approach to the boundary of moduli space , and puzio s results suggest that the classical singularity may be better - behaved in the quantum theory . a related form of quantization comes from reexpressing the moduli space for the torus as a quotient space sl(2 , )sl(2 , )/so(2 ) [ 193 , 273 ] . here , the symmetric space sl(2 , )/so(2 ) describes the transverse traceless deformations of the spatial metric , while sl(2 , ) is the modular group . as waldron has observed , this makes it possible to reinterpret the quantum mechanical problem as that of a fictitious free particle , with mass proportional \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\sqrt \lambda$\end{document } , moving in a quotient space of the ( flat ) three - dimensional milne universe . with a suitable choice of coordinates , though , the problem again reduces to that of understanding the hamiltonian ( 53 ) and the corresponding maass forms . while the choice ( 53 ) of operator ordering is not unique , the number of alternatives is smaller than one might expect . the key restriction is diffeomorphism invariance : the eigenfunctions of the hamiltonian should transform under a one - dimensional unitary representation of the mapping class group ( 50 ) . the representation theory of this group is well - understood [ 120 , 184 ] ; one finds that the possible hamiltonians are all of the form ( 53 ) , but with 0 replaced by 54\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\delta _ n } = - { \tau _ 2}^2\left({{{{\partial ^2 } } \over { \partial { \tau _ 1}^2 } } + { { { \partial ^2 } } \over { \partial { \tau _ 2}^2 } } } \right ) + 2 in{\tau _ 2}{\partial \over { \partial { \tau _ 1 } } } + n(n + 1),\quad 2 n\;\in \;{\mathbb z},$$\end{document } the maass laplacian acting on automorphic forms of weight n. it has been suggested in that the choice n = 1/2 is most natural from the point of view of chern - simons quantization . note that when written in terms of the momentum p , the operators n differ from each other by terms of order , as one would expect for operator ordering ambiguities . nevertheless , the choice of ordering may have dramatic effects on the physics , since the spectra of the various maass laplacians are quite different . this ordering ambiguity may be viewed as arising from the structure of the classical phase space . the torus moduli space is not a manifold , but rather has orbifold singularities , and quantization on an orbifold is generally not unique . since the space of solutions of the einstein equations in 3 + 1 dimensions has a similar orbifold structure , we might expect a similar ambiguity in realistic ( 3 + 1)-dimensional quantum gravity . it appears to be self - consistent , and like ordinary quantum mechanics , it is guaranteed to have the correct classical limit on the reduced phase space of section 2.4 . the principal drawback is that the method relies on a classical choice of time coordinate , which occurs as part of the gauge - fixing needed to solve the constraints . in particular , the analysis of section 2.4 required that we choose the york time - slicing from the start ; a different choice might lead to a different quantum theory , as it is known to do in quantum field theory . in other words , it is not clear that this approach to quantum gravity preserves general covariance . the problem may be rephrased as a statement about the kinds of questions we can ask in this quantum theory . the model naturally allows us to compute the transition amplitude between the spatial geometry of a time slice of constant mean curvature tr k = t1 and the geometry of a later slice of constant mean curvature tr k = t2 . indeed , such amplitudes are given explicitly in , where it is shown that they are peaked around the classical trajectory . but it is far less clear how to ask for transition amplitudes between other spatial slices , on which tr k is not constant . such questions would seem to require a different classical time - slicing , and thus a different and perhaps inequivalent quantum theory . we will eventually find a possible way out of this difficulty in section 3.4 . as a first step , we next turn to an alternative approach to quantization , one that starts from the first order formalism . as we saw in section 2.3 , ( 2 + 1)-dimensional general relativity in first order form the quantization of chern - simons theory is well understood [ 278 , 243 , 244 , 116 , 34 ] . for noncompact groups such as those that appear in gravity , much less is known , though there has been some promising work [ 280 , 155 , 43 , 150 ] . nevertheless , interesting progress can be made , especially in the simple case of a manifold with topology t. in contrast to the reduced phase space quantization of the preceding section 3.1 , our understanding of the quantum chern - simons gravity depends strongly on the sign of the cosmological constant . for < 0 , the relevant gauge group is so(2 , 2 ) or its cover sl(2 , ) sl(2 , ) . this is the most poorly understood case ; an explicit quantization of the algebra holonomies exists for genus one ( see below ) and genus two , but more general results do not yet exist . for > 0 , the relevant gauge group is so(3 , 1 ) or its cover sl(2 , ) , a complex gauge group whose chern - simons theory is somewhat better understood [ 280 , 155 , 43 ] . as noted in section 2.3 , the poisson brackets for this theory are related to the quantum double of the lorentz group , and buffenoir et al . have used this structure to write down an explicit quantization . as far as i know , the relationship between this work , which is based on a hamiltonian formalism and combinatorial quantization , and that of witten and hayashi [ 280 , 155 ] , which is based on geometric quantization , has not yet been explored . for = 0 , the relevant gauge group is iso(2 , 1 ) , the ( 2 + 1)-dimensional poincar group , or its universal cover . here there is again a connection to the quantum double of the lorentz group , which has been used in [ 37 , 36 , 201 ] to explore the quantum theory , although largely in the context of point particles . in this case , one has the nice feature that the phase space has a natural cotangent bundle structure , allowing us to immediately identify the holonomies of the spin connection as generalized positions , and their derivatives as generalized momenta . this provides a direct link to the loop variables of ashtekar , rovelli , and smolin [ 26 , 29 ] , 55\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${t^0}[\gamma ] = { 1 \over 2}{\rm{tr}}\;{\rho _ 0}[\gamma , x],\quad \;{t^1}[\gamma ] = \int\nolimits_\gamma { { \rm{tr } } } \;\{{\rho _ 0}[\gamma , x(s)]{e^a}(\gamma ( s)){{\mathcal j}_a}\},$$\end{document } where 56\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\rho _ 0}[\gamma , x ] = p\;\exp \left({\int\nolimits_\gamma { { \omega^a } } { { \mathcal j}_a } } \right)$$\end{document } is the sl(2 , ) holonomy of the spin connection and t [ ] can be expressed as a derivative of t[ ] along a path in the space of flat connections . note that the generator \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal j}$\end{document } may , in principle , be in any representation of sl(2 , ) , and that the trace in equation ( 55 ) may depend on the choice of representation . i will return to the resulting quantum theory , loop quantization , in section 3.5 . as in reduced phase space quantization , matters simplify considerably for the torus universe t. let us again focus on the case < 0 . a complete in fact , overcomplete set of observables is given by the traces ( 47 ) of the holonomies , and our goal is to quantize the algebra ( 48 ) . to do so , we proceed as follows : we replace the classical poisson brackets { , } by commutators [ , ] , with the rule [ x , y ] = xy yx = i{x , y}.on the right hand side of equation ( 48 ) , we replace the product by the symmetrized product , \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$xy \to { 1 \over 2}(xy + yx)$\end{document}. we replace the classical poisson brackets { , } by commutators [ , ] , with the rule [ x , y ] = xy yx = i{x , y}. on the right hand side of equation ( 48 ) , we replace the product by the symmetrized product , \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$xy \to { 1 \over 2}(xy + yx)$\end{document}. the resulting algebra is defined by the relations 57\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\hat{r}_1^ \pm \hat{r}_2^ \pm { e^{\pm i\theta } } - \hat{r}_2^ \pm \hat{r}_1^ \pm { e^{\mp i\theta } } = \pm 2i\;\sin \;\theta \;\hat{r}_{12}^ \pm \quad { \rm{and}}\;{\rm{cyclical}}\;{\rm{permutations}}$$\end{document } with tan = /(8 ) . the algebra ( 57 ) is not a lie algebra , but it is related to the lie algebra of the quantum group uq(sl(2 ) ) with q = exp 4i . classically , the observables \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_1^ \pm , r_2^ \pm$\end{document } , and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_{12}^ \pm$\end{document } are not independent ; in the quantum theory , the corresponding statement is that the quantities 58\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\hat{f}^ \pm } = 1 - { \tan ^2}\theta - { e^{\pm 2i\theta}}\left({{{(\hat{r}_1^ \pm)}^2 } + { { ( \hat{r}_{12}^ \pm)}^2 } } \right ) - { e^{\mp 2i\theta}}{(\hat{r}_2^ \pm)^2 } + 2{e^{\pm i\theta}}\cos \theta \hat{r}_1^ \pm \hat{r}_2^ \pm \hat{r}_{12}^ \pm$$\end{document } commute with the holonomies , and can be consistently set to zero . in terms of the parameters \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_a^ \pm$\end{document } of equation ( 38 ) , the algebra can be represented by [ 87 , 86 ] 59\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{\hat{r}_1^ \pm = \sec \;\theta \;\cosh { { \hat{r}_1^ \pm } \over 2 } , } & { \hat{r}_2^ \pm = \sec \;\theta \;\cosh { { \hat{r}_2^ \pm } \over 2 } , } & { \hat{r}_{12}^ \pm = \sec \;\theta \;\cosh { { ( \hat{r}_1^ \pm + \hat{r}_2^ \pm ) } \over 2 } , } \\ \end{array}$$\end{document } with 60\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$[\hat{r}_1^ \pm , \hat{r}_2^ \pm ] = \pm 8i\theta , \quad [ \hat{r}_a^ + , \hat{r}_b^ - ] = 0.$$\end{document } for small , these commutators differ from the naive quantization of the classical brackets ( 45 ) by terms of order . an alternative quantization , also differing by terms of order , works directly with the holonomy matrices ( 38 ) , imposing a quantum - group - like quantization condition 61\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\rho ^ + } [ { \gamma _ 1}]{\rho ^ + } [ { \gamma _ 2 } ] = q{\rho ^ + } [ { \gamma _ 2}]{\rho ^ + } [ { \gamma _ 1}],\quad \quad q = { e^{- i\hbar/4\ell}},$$\end{document } with a similar condition for . we must also implement the action of the modular group ( 50 ) on the operators \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\hat r_a^ \pm$\end{document}. one can find an action preserving the algebraic relations ( 57 ) , corresponding to a particular factor ordering of the classical modular group . the nelson - picken quantization ( 61 ) admits a similar modular group action . for a full quantum theory , of course , one needs not only an abstract operator algebra , but a hilbert space upon which the operators act . for the t universe , equation ( 60 ) suggests that a natural choice is to take wave functions to be square integrable functions of the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_2^ \pm$\end{document}. there is a potential difficulty here , however : the modular group does not act properly discontinuously on this configuration space . this means that the quotient of this space by the modular group is badly behaved ; in fact , there are no nonconstant modular invariant functions of the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_2^ \pm$\end{document } [ 182 , 143 , 221 ] . the technique of chern - simons quantization relies on special features of general relativity in 2 + 1 dimensions , and does not readily generalize to higher dimensions . it is , however , closely related to a much more general approach , covariant canonical quantization [ 31 , 28 , 98 , 175 , 272 ] , or quantization of the space of classical solutions . our starting point is the observation that the phase space of a well - behaved classical theory is isomorphic to the space of classical solutions . indeed , if \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal c}$\end{document } is an arbitrary cauchy surface , then a point in the phase space determines initial data on \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal c}$\end{document } , which can be evolved to give a unique solution , while , conversely , a classical solution restricted to \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal c}$\end{document } determines a point in the phase space . moreover , the space of solutions has natural symplectic structure [ 175 , 272 ] , which can be shown to be equivalent to the standard symplectic structure on phase space . for the case of ( 2 + 1)-dimensional gravity , this equivalence is demonstrated in section 6.1 of . for ( 2 + 1)-dimensional gravity , the space of classical solutions is the space of geometric strictures of section 2.2 . if we restrict our attention to spacetimes with the topology with s closed and 0 , the holonomies of a geometric structure determine a unique maximal domain of dependence , exactly the right setting for covariant canonical quantization . but as we saw in section 2.3 , the holonomies of a geometric structure are precisely the holonomies of the chern - simons formalism , and the symplectic structures are the same as well . thus in this setting , chern - simons quantization is covariant canonical quantization . if > 0 or point particles are present , the holonomies do not quite determine a unique geometric structure , and the chern - simons theory is not quite equivalent to general relativity . in that case , additional discrete variables might be necessary ; see , for example , for the case of a torus universe with > 0 . as we shall see in section 3.4 , the construction of dynamical observables and time - dependent states in covariant canonical quantum theory requires an explicit isomorphism between the phase space and the space of classical solutions . for the torus universe , such an isomorphism is known . for higher genus spaces , however and certainly for realistic ( 3 + 1)-dimensional gravity it is not . often , however , we can determine such an isomorphism perturbatively in the neighborhood of a known classical solution . this raises the interesting question , so far answered only in simple models , of whether classical perturbation theory can be used to define a perturbative covariant canonical quantum theory . when one attempts to interpret the quantum theories coming from the chern - simons formalism or covariant canonical quantization , one finds an immediate and rather profound difficulty . the gauge - invariant observables the traces of the holonomies are automatically nonlocal and time - independent , and one obtains a frozen time formalism , or what kucha has called quantum gravity without time . in one sense , this is a good thing : one knows from general arguments that the diffeomorphism - invariant observables in any quantum theory of gravity must have these features . on the other hand , it is not at all easy to see how to extract local geometry and dynamics from such a picture : if our only observables are nonlocal and time - independent , how can we recover a classical limit with local excitations that evolve in time ? a geometric structure determines a spacetime , and must contain within it all of the dynamics of that spacetime . on the other hand , the basic data that fix the geometric structure the transition functions , or , often , the holonomies have no obvious dynamics . in principle , the classical answer is simple : use , say , the holonomies to determine a spacetime geometry.select a favorite time-slicing.read off the spatial metric and its time derivatives from the spacetime metric of step 1 in this slicing . this procedure can be understood as a concrete realization of the isomorphism described in section 3.3 between the phase space and the space of classical solutions , with the cauchy surface \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal c}$\end{document } fixed by the choice of time - slicing . read off the spatial metric and its time derivatives from the spacetime metric of step 1 in this slicing . for the simple case of the torus universe , these steps can be transcribed almost directly to the quantum theories . equations ( 42 , 43 , 44 ) become definitions of operators , 62\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{{{\hat\tau}_t } = \left({\hat{r}_1^ - { e^{it/\ell } } + \hat{r}_1^ + { e^{- it/\ell } } } \right){{\left({\hat{r}_2^ - { e^{it/\ell } } + \hat{r}_2^ + { e^{- it/\ell } } } \right)}^{- 1 } } , } \\ { { { \hat{p}}_t } = - { { i\ell } \over { 2\sin { { 2 t } \over \ell}}}{{\left({\hat{r}_2^ + { e^{it/\ell } } + \hat{r}_2^ - { e^{- it/\ell } } } \right)}^2 } , } \\ { { { \hat{h}}_t } = { { { \ell ^2 } } \over 4}\sin { { 2 t } \over \ell}(\hat{r}_1^ - \hat{r}_2^ + - \hat{r}_1^ + \hat{r}_2^ - ) , } \\ \end{array}$$\end{document } where the operator ordering has been chosen to respect the modular transformations ( 50 ) . the parameter t is now merely a label for a one - parameter family of diffeomorphism - invariant observables . these observables obtain their physical significance from the classical limit : \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat \tau}_t}$\end{document } , for example , is the operator whose expectation value gives the mean value of the modulus on a time slice of constant mean curvature \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$t = - { 2 \over \ell}\cot { { 2 t } \over \ell}$\end{document}. such observables are examples of what rovelli has called evolving constants of motion [ 236 , 237 ] . from this point of view , we should think of chern - simons / covariant canonical quantization as a sort of heisenberg picture , with time - independent states and time-dependent operators . to obtain the corresponding schrodinger picture , we proceed as in ordinary quantum mechanics : we diagonalize \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat \tau}_t}$\end{document } , obtaining a transition matrix \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$k(\tau , \bar \tau ; t\vert r_2^ + , r_2^ - ) = \left\langle { \tau , \bar \tau ; t\vert r_2^ + , r_2^ - } \right\rangle$\end{document } that allows us to transform between representations [ 68 , 88 ] . the resulting time-dependent wave functions obey a schrdinger equation of the form ( 52 , 53 ) , but with the laplacian in replaced by the weight 1/2 maass laplacian 1/2 of equation ( 54 ) . in , it has been shown that these wave functions are peaked around the correct classical trajectories . ( different operator orderings in equation ( 62 ) give different weight laplacians . ) as a useful byproduct , this analysis allows us to solve the problem of the poorly - behaved action of the modular group discussed at the end of section 3.2 [ 88 , 89 ] . if we start with a reduced phase space wave function \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\tilde \psi ( \tau , \bar \tau ; t)$\end{document } and use the transition matrix k to determine a chern - simons wave function \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\psi ( r_2^ + , r_2^ -)$\end{document } , we find , indeed , that \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\psi ( r_2^ + , r_2^ -)$\end{document } is not modular invariant . instead , though , the entire hilbert space of chern - simons wave functions splits into fundamental regions , orthogonal subspaces that transform into each other under the action of the modular group . any one of these fundamental regions is equivalent to any other , and each is equivalent to the hilbert space arising from reduced phase space quantization . moreover , matrix elements of any modular invariant function vanish unless they are taken between states in the same fundamental region . modular invariance thus takes a slightly unexpected form , but can still be imposed by restricting the theory to a single fundamental region of the hilbert space . we can also begin to address the problem raised at the end of section 3.1 , the limited and slicing - dependent range of questions one can ask in reduced phase space quantization . the operators ( 62 ) introduced here on the covariant canonical hilbert space were obtained from a particular classical time - slicing , and answer questions about spatial geometry in that slicing . in principle , however , we can choose any other slicing , with a new time coordinate \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\bar t}$\end{document } , and determine the corresponding operators \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat \tau}_t},{{\hat p}_{\bar t}}$\end{document } , and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat h}_{\bar t}}$\end{document}. the operator ordering of such operators will , of course , be ambiguous , though one might hope that the action of the modular group might again restrict the choices . but such an ambiguity need not be seen as a problem with the theory ; rather , it is merely a statement that many different quantum operators can have the same classical limit , and that ultimately experiment must decide which operator we are really observing . there is , to be sure , a danger that the schrdinger pictures coming from different time - slicings may not be consistent . suppose , for example , that we choose two slicings that agree on an initial and a final slice 1 and 2 , but disagree in between . if we start with an initial wave function on 1 , we must check that the hamiltonians coming from the different slicings evolve us to the same final wave function on 2 . for field theories , even in flat spacetime , this will not always happen . for ( 2 + 1)-dimensional gravity , on the other hand , there is evidence that one can always find operator orderings of the hamiltonians that ensure consistent evolution . if this ultimately turns out not to be the case , however , it may simply mean that we should treat the covariant canonical picture as fundamental , and discard the schrodinger pictures of time - dependent states . we now resume the discussion of alternative approaches to quantum gravity in 2 + 1 dimensions . in 3 + 1 dimensions , one of the most attractive programs of quantization is loop quantum gravity , or quantum geometry = 0 , this approach is closely related to the first order formalism of section 2.3 , but takes as its fundamental observables the loop variables t[ ] and t[ ] of equation ( 55 ) . more precisely , loop quantum gravity starts with a hamiltonian formulation of the first order formalism , with constraints , written in analogy to the ( 3 + 1)-dimensional case , that take the form 63\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${d_i}{\tilde{e}^{ia } } = 0,\quad { \tilde{e}^i}_ar_{ij}^a = 0,\quad { \epsilon _ { abc}}{\tilde{e}^{ib}}{\tilde{e}^{jc}}r_{ij}^a = 0.$$\end{document } here , the indices i , j , k are spatial indices on a surface of constant time , \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\tilde e}^{ia } } = { \in ^{ij}}{e_j}^a$\end{document } , di is the so(2 , 1 ) gauge - covariant derivative for the connection , and the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$r_{ij}^a$\end{document } are the spatial components of the curvature two - form ( 16 ) . when the spatial metric \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${g_{ij } } = { e_i}^a{e_{ja}}$\end{document } is nondegenerate , it is straightforward to show that these constraints are equivalent to the standard constraints of first order gravity , and the classical theories are identical . when gij , is noninvertible , on the other hand , the constraints are not equivalent . instead , the constraints ( 63 ) yield a phase space with infinitely many degrees of freedom , arising from the geometries formed from an arbitrary collection of independent patches of ordinary spacetime separated by regions with degenerate metrics [ 44 , 45 ] . let us restrict ourselves to invertible spatial metrics , and attempt to quantize the algebra of loop variables \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^0}[{\rm{\gamma ] } } $ \end{document } and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^1}[{\rm{\gamma ] } } $ \end{document}. for the torus universe , it is not hard to show that such a quantization simply reproduces the theory we already obtained in the chern - simons formulation ( see , for example , section 7.2 of ) . there is another way to look at the operator algebra of the operators \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^0}[{\rm{\gamma ] } } $ \end{document } and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^1}[{\rm{\gamma ] } } $ \end{document } , however , which leads to a new approach , the loop representation . up to now , we have been thinking of the operators \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\hat t}$\end{document } as a set of functions of the triad and spin connection , indexed by loops . our wave functions are thus functionals of the configuration space variable , or , more precisely , functions on the moduli space of flat so(2 , 1 ) or sl(2 , ) connections on . but we could equally well view the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\hat t}$\end{document } operators as functions of loops or , in 2 + 1 dimensions , homotopy classes [ ] of loops indexed by e and . wave functions would then be functions of loops or sets of loops . this change of viewpoint is rather like the decision in ordinary quantum mechanics to view a wave function e as a function on momentum space , indexed by q , rather than a function on position space , indexed by p. the loop representation is complicated by the existence of mandelstam identities among holonomies of loops , but for the case of the torus universe , a complete , explicit description of the states is again possible [ 26 , 29 ] . the simplest construction begins with a vacuum state \|0 annihilated by every operator \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^1}[{\rm{\gamma ] } } $ \end{document } , and treats the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^0}[{\rm{\gamma ] } } $ \end{document } as creation operators . since any homotopy class [ ] of loops on the torus is completely characterized by a pair of winding numbers ( m , n ) , one can write these states as \|m , n. the action 64\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}c}{{{\widehat{t}}^0}[m , n]\vert p , q\rangle = \quad { 1 \over 2}(\vert m + p , n + q\rangle + \vert m - p , n - q\rangle ) } \\ { { { \widehat{t}}^1}[m , n]\vert p , q\rangle = \quad - { i \over 8}(mq - np)(\vert m + p , n + q\rangle - \vert m - p , n - q\rangle ) } \\ \end{array}$$\end{document } then gives a representation of the loop algebra . observe now that the loop variables t[ ] depend only on the configuration space variable . we can thus relate the loop representation to the chern - simons representation by simultaneously diagonalizing these operators , obtaining wave functions that are functions of the so(2 , 1 ) holonomies alone . for the torus universe , this loop transform can be obtained explicitly [ 26 , 29 , 192 ] , and written as a simple integral transform . the properties of this transform depend on the holonomies , that is , the eigenvalues of \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^0}[{\rm{\gamma ] } } $ \end{document}. for simplicity , let us take the generator \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\mathcal j}$\end{document } in equation ( 56 ) to be in the two - dimensional representation of sl(2 , ) . in the timelike sector , in which the traces of the two holonomies are both less than two , the loop transform is a simple fourier transformation , and chern - simons and loop quantization are equivalent . unfortunately , though , this is not the physically relevant sector : it does not correspond to a geometric structure with spacelike t slices . for a physically interesting geometry , one must go to the spacelike sector , in which the traces of the holonomies are both greater than two . in this sector , the transform is not very well - behaved : in fact , a dense set of chern - simons states transforms to zero . the loop representation thus appears to be rather drastically different from the chern - simons formulation . the problems in the physical sector can be traced back to the fact that sl(2 , ) is a noncompact group . one is to start with a different dense set of chern - simons states that transform faithfully , determine the inner product and the action of the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\hat t}$\end{document } operators on the resulting loop states , and then form the cauchy completion to define the hilbert space in the loop representation . this is a consistent procedure , but many of the resulting states in the cauchy completion are no longer functions of loops in any clear sense ; they correspond instead to extended loops , whose geometrical interpretation is not entirely clear . a second possibility is to change the integration measure in the loop transform to make various integrals converge better . such a choice introduces order corrections to the action of the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${{\hat t}^1}$\end{document } operators , and one must be careful that the algebra remains consistent . this is possible , but at some cost the inner products between loop states become considerably more complex , as does the action of the mapping class group and it is not obvious that there is a canonical choice of the new measure and algebra . a third possibility is suggested by recent work on spin networks for noncompact groups [ 129 , 130 ] . this new technology essentially allows one to consider holonomies ( 56 ) that lie in infinite - dimensional unitary representations of the lorentz group , with a finite inner product defined by appropriate gauge - fixing . the quantities t and t can be represented as hermitian operators on this space of holonomies ( or on a larger space of spin networks ) . at this writing , implications of this approach for the loop transform in 2 + 1 dimensions have not yet been investigated . finally , i should briefly mention the role of spin networks in ( 2 + 1)-dimensional quantum geometry . in the ( 3 + 1)-dimensional theory , loop states have been largely superseded by spin network states , states characterized by a graph with edges labeled by representations and vertices labeled by intertwiners . such states can be defined in 2 + 1 dimensions as well , and there has been some interesting recent work on their role as , however , the full constraints imply that such states have their support on flat connections , and only holonomies around noncontractible curves describe nontrivial physics . an interesting step toward projecting out the physical states has recently been taken in , in the context of euclidean quantum gravity ; the ultimate effect is to reduce spin network states to loop states of the sort we have considered above . a better understanding of the relationship to the gauge - fixing procedure of [ 129 , 131 ] would be valuable . a long - standing approach to quantum gravity in 3 + 1 dimensions has been to look for discrete approximations to the path integral [ 179 , 232 ] : quantized regge calculus , for example , or sums over random triangulations . the basic idea is that although the full sum over geometries may be impossible to evaluate , a sum over appropriately discretized geometries might give a good approximation , perhaps becoming extremely good near a phase transition at which lattice spacing can go to zero . when applied to 2 + 1 dimensions , such methods have the added feature of sometimes being exact : since geometries satisfying the constraints have constant or zero curvature , a discrete approximation may give a complete description . regge calculus in 2 + 1 dimensions begins with a triangulated three - manifold , consisting of a collection of flat simplices joined along one - dimensional edges . the curvature of such a manifold is concentrated entirely at the edges . for a simplicial three - manifold with riemannian signature , composed of simplices with edges of length le , regge s form of the einstein - hilbert action is 65\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${i_{{\rm{regge } } } } = 2\sum\limits_{{\rm{edges}}:\;e } { { \delta _ e}{\ell _ e}},$$\end{document } where e is the conical deficit angle at the edge labeled by the index e. a similar expression exists for lorentzian signature , although the definition of the deficit angle is a bit more complicated . the first hint that ( 2 + 1)-dimensional gravity might be exceptional came from the observation by ponzano and regge that the regge action in 2 + 1 dimensions can be re - expressed in terms of wigner - racah 6j - symbols . consider first a single tetrahedron with edge lengths \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\ell _ i } = { 1 \over 2}({j_i } + { 1 \over 2})$\end{document } , where the ji are integers or half - integers . ponzano and regge noticed , and roberts later proved rigorously , that in the limit of large j , 66\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\exp \left({\pi i\sum\limits_{i = 1}^6 { ji } } \right)\left\{{\begin{array}{{20}c}{{j_1}\;{j_2}\;{j_3 } } \\ { { j_4}\;{j_5}\;{j_6 } } \\ \end{array } } \right\ } \sim { 1 \over { \sqrt { 6\pi v}}}\left\{{\exp \left [ { i\left({{i_{{\rm{regge } } } } + { \pi \over 4 } } \right ) } \right ] + \exp \left [ { - i\left({{i_{{\rm{regge } } } } + { \pi \over 4 } } \right ) } \right ] } \right\},$$\end{document } where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\left\ { { \begin{array}{{20}{c } } { { j_1}\;{j_2}\;{j_3 } } \\ { { j_4}\;{j_5}\;{j_6 } } \end{array } } \right\}$\end{document } is a 6j - symbol , iregge is the regge action ( 65 ) for the tetrahedron , and v is its volume . for a manifold made of a collection of such tetrahedra , the full regge action will occur in a product of such 6j - symbols . this suggests that the ( 2 + 1)-dimensional path integral , which is essentially a sum over geometries of terms of the form exp(iiregge ) , might be expressible as a sum of such products . ponzano and regge s specific proposal , developed by hasslacher and perry and modified by ooguri to account for boundaries , was the following : consider a three - manifold m with boundary m , with a given triangulation of m . label interior edges of tetrahedra by integers or half - integers and exterior ( boundary ) edges by ji , and for a given tetrahedron t , let ji(t ) denote the spins that color its ( interior and exterior ) edges . then 67\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\begin{array}{{20}{c } } { { z_\delta } [ \ { { j_i}\ } ] = \mathop { \lim } \limits_{l \to \infty } \sum\limits_{{x_e}l } { \left ( { \prod\limits_{{\text{ext}}.{\text{edges}}:i } { { { ( - 1)}^{2{j_i}}}\sqrt { 2{j_i } + 1 } } } \right.\prod\limits_{{\text{int}}.v{\text{ertices } } } { \lambda { { ( l)}^ { - 1 } } } } } \\ { \left . { \quad \quad \quad \quad \times \prod\limits_{{\text{int}}.{\text{edges}}:\ell } { ( 2{x_\ell } + 1)\prod\limits_{{\text{tetra}}:t } { ( - 1)\sigma _ { i = 1}^6{j_i}(t)\left\ { { \begin{array}{{20}{c } } { { j_1}(t)\;{j_2}(t)\;{j_3}(t ) } \\ { { j_4}(t)\;{j_5}(t)\;{j_6}(t ) } \end{array } } \right\ } } } } \right ) , } \end{array}$$\end{document } where int and ext mean interior and exterior and 68\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\lambda ( l ) = { \sum\limits_{j \leq l } { ( 2j + 1 ) } ^2}$$\end{document } is a regularization factor that controls divergences in the sum over interior lengths . with this weighting , identities among 6j - symbols may be used to show that the amplitude is invariant under refinement that is , subdivision of a tetrahedron into four smaller tetrahedra suggesting that we are dealing with a this is , of course , what one would hope for , based on the classical characteristics of ( 2 + 1)-dimensional gravity . topological feature of the ponzano - regge model was made more precise by turaev and viro , who discovered an improved regularization , based on the technology of quantum groups . the spins j in equation ( 67 ) can be viewed as labeling representations of su(2 ) . if these are replaced by representations of the quantum group uq(sl(2 ) ) ( quantum su(2 ) ) , with \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$q = \exp \left({{{2\pi i } \over { k + 2 } } } \right)$\end{document } , k , the number of such representations is finite , and the sum over interior edge lengths is automatically cut off . with appropriate substitutions ( e.g. , quantum 6j - symbols ) , the ponzano - regge amplitude ( 67 ) becomes well - defined without any regularization . the construction of physical states as appropriate functions of boundary edge lengths is described in section 11.2 of . the resulting amplitudes can be computed for simple topologies [ 162 , 161 ] , and have several key features : for large but finite k , the turaev - viro quantum group regularization introduces a cosmological constant to the regge action [ 204 , 203 ] , 69\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\lambda { \left({{{4\pi } \over k } } \right)^2}.$$\end{document } correspondingly , the quantum 6j - symbols are related to spherical tetrahedra rather than flat tetrahedra .in the large k limit , the turaev - viro hilbert space is isomorphic to the space of gauge - invariant functions of flat su(2 ) connections [ 219 , 220 , 242 ] . this establishes a direct link to chern - simons quantization : just as ( 2 + 1)-dimensional lorentzian gravity can be written as an iso(2 , 1 ) chern - simons theory with a configuration space of flat so(2 , 1 ) connections , three - dimensional euclidean gravity can be written as an isu(2 ) chern - simons theory with a configuration space of flat su(2 ) connections.for a closed three - manifold m , the turaev - viro amplitude now interpreted as a partition function is equal to the absolute square of the partition function of an su(2 ) chern - simons theory with coupling constant k [ 228 , 260 , 233 ] , 70\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${z_{{\rm{tv } } } } = \vert { z_{{\rm{cs}}}}{\vert ^2}.$$\end{document } this again establishes an equivalence with euclidean gravity in first - order form : the first - order euclidean action with > 0 can be written as a difference of su(2 ) chern - simons actions , so 71\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${z_{{\text{grav } } } } = \int { [ d{a^ + } ] [ d{a^ - } ] } \;{e^{i(i[{a^ + } ] - i[{a^ - } ] ) } } = { \text { } } { \left\| { \int { [ d{a^ + } ] { e^{ii[{a^ + } ] } } } } \right\|^2},$$\end{document } in agreement with equation ( 70).a candidate for a discrete version of the wheeler - dewitt equation in three dimensions has been found , for which the ponzano - regge wave functions are solutions . although it has not been universally appreciated , the existence of a divergence in the sum ( 67 ) regulated either by an explicit cut - off or by quantum group tricks is rather mysterious , given the absence of local excitations and the general well - behavedness of gravity in three dimensions . this mystery may have recently been solved by freidel and louapre , who show that a residual piece of the diffeomorphism symmetry has not been factored out of the ponzano - regge action . because of this symmetry , the sum ( 67 ) overcounts physical configurations , and the regulator (l ) is simply the remaining gauge volume . freidel and louapre further show that the symmetry can instead be gauge - fixed , leading to a sum over a restricted and considerably simplified class of collapsed triangulations . for large but finite k , the turaev - viro quantum group regularization introduces a cosmological constant to the regge action [ 204 , 203 ] , 69\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\lambda { \left({{{4\pi } \over k } } \right)^2}.$$\end{document } correspondingly , the quantum 6j - symbols are related to spherical tetrahedra rather than flat tetrahedra . in the large k limit , the turaev - viro hilbert space is isomorphic to the space of gauge - invariant functions of flat su(2 ) connections [ 219 , 220 , 242 ] . this establishes a direct link to chern - simons quantization : just as ( 2 + 1)-dimensional lorentzian gravity can be written as an iso(2 , 1 ) chern - simons theory with a configuration space of flat so(2 , 1 ) connections , three - dimensional euclidean gravity can be written as an isu(2 ) chern - simons theory with a configuration space of flat su(2 ) connections . for a closed three - manifold m , the turaev - viro amplitude now interpreted as a partition function is equal to the absolute square of the partition function of an su(2 ) chern - simons theory with coupling constant k [ 228 , 260 , 233 ] , 70\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${z_{{\rm{tv } } } } = \vert { z_{{\rm{cs}}}}{\vert ^2}.$$\end{document } this again establishes an equivalence with euclidean gravity in first - order form : the first - order euclidean action with > 0 can be written as a difference of su(2 ) chern - simons actions , so 71\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${z_{{\text{grav } } } } = \int { [ d{a^ + } ] [ d{a^ - } ] } \;{e^{i(i[{a^ + } ] - i[{a^ - } ] ) } } = { \text { } } { \left\| { \int { [ d{a^ + } ] { e^{ii[{a^ + } ] } } } } \right\|^2},$$\end{document } in agreement with equation ( 70 ) . a candidate for a discrete version of the wheeler - dewitt equation in three dimensions has been found , for which the ponzano - regge wave functions are solutions . while the mathematics of ponzano - regge and turaev - viro models has been studied extensively , so far only a bit of attention has been given to the traditional issues of quantum gravity . a few numerical investigations of the ponzano - regge path integral have been undertaken , but the evidence of a continuum limit is thus far inconclusive . the model has been used to study conditional probabilities and the emergence of quasiclassical behavior in quantum gravity , but the cut - off dependence of these results makes their physical significance unclear . in an interesting recent paper , colosi et al . have investigated the dynamics of a single tetrahedron , showing that a quantum description of the evolution can be given in terms of a boundary amplitude . a number of observables , whose expectation values generally give topological information about the spacetime or about knots within spacetime , have been discussed in [ 23 , 54 , 261 , 139 ] . with a few exceptions , though , work in this area has remained largely mathematical in nature ; fairly little is understood about the physics of these observables , although some are probably related to length spectra and perhaps volumes [ 127 , 64 ] , and others are almost certainly connected to scattering amplitudes for test particles . the ponzano - regge and turaev - viro models are examples of spin foam models [ 35 , 222 ] , that is , a model based on simplicial complexes with faces , edges , and vertices labeled by group representations and intertwiners . it has been known for several years how to generalize the ponzano - regge action for a single tetrahedron [ 48 , 101 , 191 ] , and recently considerable progress has been made in constructing lorentzian spin foam models [ 222 , 125 , 102 ] . probably the most elegant derivation of a lorentzian spin foam description starts with the first - order action ( 14 ) , with = 0 , for a triangulated manifold [ 125 , 128 ] . one can rewrite the action in terms of a set of discrete variables : a lie algebra element e corresponding to the integral of e along the edge of a tetrahedron in the triangulation , and a holonomy g of the connection around the edge . the path integral then becomes an integral over these variables . as in the continuum path integral of section 3.10 , this translates back to the geometric statement that the constraints require the connection w to be flat , and thus to have trivial holonomy around a contractible curve surrounding an edge . for the euclidean ponzano - regge action , g su(2 ) , and the key trick is now to use the plancherel formula to express each (g ) as a sum over the characters of finite - dimensional representations of su(2 ) . fairly straightforward arguments then permit an exact evaluation of the remaining integrals over the g , reproducing the 6j symbols in the ponzano - regge action . to obtain a lorentzian version the corresponding plancherel formula involves a sum over both the ( continuous ) principle series of representations of so(2 , 1 ) and the discrete series . similar methods may be used for supergravity . the resulting rather complicated expression for the partition function the appearance of both continuous and discrete labels has a nice physical interpretation : continuous representations describe spacelike edges , and seem to imply a continuous length spectrum , while discrete representations label timelike edges , and suggest discrete time . these results should probably not yet be considered conclusive , since they require operators that do not commute with all of the constraints , but they are certainly suggestive . while spin foam models ordinarily assume a fixed spacetime topology , recent work has suggested a method for summing over all topologies as well , thus allowing quantum fluctuations of spacetime topology . these results will be discussed in section 3.11 . methods from 2 + 1 dimensions have also been generalized to higher dimensions , leading to new insights into the construction of spin foams . spin foam models are based on a fixed triangulation of spacetime , with edge lengths serving as the basic gravitational variables . an alternative scheme is dynamical triangulation , in which edge lengths are fixed and the path integral is represented as a sum over triangulations . ( for reviews of this approach in arbitrary dimensions , see [ 8 , 179 ] . ) dynamical triangulation has been proven to be quite useful in two - dimensional gravity , and some important steps have been taken in higher dimensions , especially with the recent progress in understanding lorentzian triangulations . the starting point is now a simplicial complex , diffeomorphic to a manifold m , composed of an arbitrary collection of equilateral tetrahedra , with sides of length a. metric information is no longer contained in the choice of edge lengths , but rather depends on the combinatorial pattern . such a model is not exact in 2 + 1 dimensions , but one might hope that as a becomes small and the number of tetrahedra becomes large it may be possible to approximate an arbitrary geometry . in particular , it is plausible ( although not rigorously proven ) that a suitable model lies in the same universality class as genuine ( 2 + 1)-dimensional gravity , in which case the continuum limit should be exact . the einstein - hilbert action for such a theory takes the standard regge form ( 65 ) , which for spherical spatial topology reduces to a sum 72\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$i = - { k_0}{n_0 } + { k_3}{n_3},$$\end{document } where n0 and n3 are the numbers of vertices and tetrahedra in the triangulation , k0 = a/(4 g ) , and k3 is related to the cosmological constant . as the number of tetrahedra becomes large , the number of distinct triangulations ( the entropy ) increases exponentially , while the n3 term in equation ( 72 ) provides an exponential suppression . path integral \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\sum \exp ( - i)$\end{document } should thus converge for k3 greater than a critical value \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$k_3^{\rm{c}}({k_0})$\end{document}. as k3 approaches \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$k_3^{\rm{c}}({k_0})$\end{document } from above , expectation values of n3 will diverge , and one may hope for a finite - volume continuum limit as a 0 . for ordinary the system appears to exhibit two phases a crumpled phase , in which the hausdorff dimension is extremely large , and a branched polymer phase neither of which look much like a classical spacetime . lorentzian model , introduced by ambjrn and loll [ 16 , 9 , 12 , 10 , 180 , 13 ] , however , has much nicer properties , including a continuum limit that appears numerically to match a finite - sized , spherical the lorentzian model begins with a slicing of spacetime into constant time surfaces , each of which is given an equilateral triangulation . the region between two neighboring slices is then filled in by tetrahedra , which can come only in the three varieties shown in figure 3 . this setup automatically restricts spacetime to have the topology , and by declaring each slice to be spacelike and each edge joining adjacent slices to be timelike , one has a well - defined wick rotation to a riemannian signature metric with regge action ( 72 ) . note that for convergence , this method requires a positive value of k3 , and thus a positive cosmological constant . the path integral for such a system can be evaluated numerically , using monte carlo methods and a set of moves that systematically change an initial triangulation [ 12 , 10 ] . the system splits into uncorrelated two - dimensional spaces , each well - described by two - dimensional gravity . at weak coupling , however , a regime appears that resembles the picture obtained from other approaches to ( 2 + 1)-dimensional gravity . in particular , one may evaluate the expectation value a(t) of the spatial area at fixed time and the correlation a(t)a(t + 1) of successive areas ; the results match the classical de sitter behavior for a spacetime s quite well . the more local behavior the hausdorff dimension of a constant time slice , for example is not yet well - understood . neither is the role of moduli for spatial topologies more complicated than s , although initial steps have been taken for the torus universe . the lorentzian dynamical triangulation model can also be translated into a two - matrix model , the so - called abab model . the feynman diagrams of the matrix model correspond to dual graphs of a triangulation , and matrix model amplitudes become particular sums of transfer matrix elements in the gravitational theory [ 11 , 14 , 15 ] . in principle , this connection can be used to solve the gravitational model analytically . while this goal has not yet been achieved ( though see ) , for example , the matrix model connection can be used to show that newton s constant and the cosmological constant are additively renormalized , and to analyze the apparent nonrenormalizability of ordinary field theoretical approach . in principle the discrete approaches described in section 2.5 in particular , the lattice descriptions of t hooft and waelbroeck should be straightforward to quantize . in practice , there has been fairly little work in this area , and most of the literature that does exist involves point particles rather than closed universes. t hooft has emphasized that the hamiltonian in his approach is an angle , and that time should therefore be discrete , in agreement with the lorentzian spin foam analysis of section 3.6. t hooft has also found that for a particular representation of the commutation relations for a point particle in ( 2 + 1)-dimensional gravity , space may also be discrete , although it remains unclear whether these results can be generalized beyond this one special example . have examined waelbroeck s lattice hamiltonian approach for the quantized torus universe , investigating the implication of the choice of an internal time variable , and waelbroeck has studied the role of the mapping class group . the approaches to quantization of sections 3.1 , 3.2 , 3.3 , 3.4 , and 3.5 share an important feature : all are reduced phase space quantizations , quantum theories based on the true physical degrees of freedom of the classical theory . that is , the classical constraints have been solved before quantizing , eliminating classically redundant gauge degrees of freedom . in dirac s approach to quantization [ 112 , 113 , 114 ] , in contrast , one quantizes the entire space of degrees of freedom of classical theory , and only then imposes the constraints . states are initially determined from the full classical phase space ; in the adm formulation of quantum gravity , for instance , they are functionals [gij , ] of the full spatial metric . the constraints then act as operators on this auxiliary hilbert space ; the physical hilbert space consists of those states that are annihilated by the constraints , with a suitable new inner product , acted on by physical operators that commute with the constraints . for gravity , in particular , the hamiltonian constraint acting on states leads to a functional differential equation , the wheeler - dewitt equation [ 110 , 276 ] . in the first order formalism , it is straightforward to show that dirac quantization is equivalent to the chern - simons quantum theory we have already seen . details can be found in chapter 8 of , but the basic argument is fairly clear : at least for = 0 , the first order constraints coming from equations ( 15 , 16 ) are at most linear in the momenta , and are thus uncomplicated to solve . in the second order formalism , matters become considerably more complicated . we begin with a wave function [gij ] , upon which we wish to impose the constraints ( 30 ) , with momenta acting as functional derivatives , 73\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${\pi ^{ij } } = - i{\delta \over { \delta { g_{ij}}}}.$$\end{document } the first difficulty is that we are no longer allowed to choose a nice time - slicing such as york time ; that would be a form of gauge - fixing , and is not permitted in dirac quantization . we can still decompose the spatial metric and momentum as in equation ( 31 ) , but only up to a spatial diffeomorphism , which depends on an undetermined vector field y appearing in the momentum . the momentum constraint fixes in terms of the scale factor , but it does so nonlocally . as a consequence , the hamiltonian constraint becomes a nonlocal functional differential equation , and very little is understood about its solutions , even for the simplest case of the torus universe . further complications come from the fact that the inner product on the space of solutions of the wheeler - dewitt equation must be gauge - fixed [ 282 , 144 ] ; again , little is understood about the resulting hilbert space . in view of the difficulty in finding exact solutions to the wheeler - dewitt equation , it is natural to look for perturbative methods , for example an expansion in powers of newton s constant g. one can solve the momentum constraints order by order by insisting that each term depend only on ( nonlocal ) spatially diffeomorphism - invariant quantities . such an expansion has been studied by banks , fischler , and susskind for the physically trivial topology s , following much earlier work by leutwyler . even in this simple case , other attempts have been made [ 47 , 186 ] to write a discrete version of the wheeler - dewitt equation in the ponzano - regge formalism of section 3.6 . this approach has the advantage that the spatial diffeomorphisms have already been largely eliminated , removing the main source of nonlocality discussed above . the wheeler - dewitt - like equation in has been shown to agree with the the ponzano - regge model . up to now , i have mainly concentrated on approaches to quantum gravity that fall under the broad heading of canonical quantization . an alternative approach already implicit in the discussion of discrete methods starts with the feynman path integral , or in an important sense , path integral methods are less precise than those of canonical quantization : the infinite - dimensional integral over histories can rarely be rigorously defined , we do not really know what classes of paths to sum over , and ordering ambiguities in the operator formalism reemerge as ambiguities in the integration measure . on the other hand , path integrals allow us to ask questions for example , about amplitudes for spatial topology change that are difficult or impossible to formulated in a canonical theory . the simplest path integral approach to ( 2 + 1)-dimensional quantum gravity is the phase space path integral , in which the action is written in the adm form ( 29 , 30 ) , and the spatial metric gij and momentum are treated as independent integration variables . the lapse n and shift appear as lagrange multipliers , and the integrals over these quantities yield delta functionals for the constraints and i . one might therefore expect the result to be equivalent to the canonical quantization of section 3.1 , in which the constraints are set to zero and solved for the physical degrees of freedom . this is indeed true , as shown in [ 77 , 245 ] for spatially closed universes and for geometries with point particles . the main subtlety comes from the appearance of many different determinants , arising from gauge - fixing and from the delta functionals , which must be shown to cancel . the phase space path integral for the first order formulation similarly reproduces the corresponding canonically quantized theory . it is perhaps more interesting to look at the covariant metric path integral , in which one starts with the ordinary einstein - hilbert action and gauge - fixes the full ( 2 + 1)-dimensional diffeomorphism group . this approach does not require a topology , and could potentially describe topology - changing amplitudes . section 9.2 of describes a partial gauge - fixing , which takes advantage of the fact that every metric on a three - manifold is conformal to one of constant scalar curvature . but while this leads to some simplification , we are still left with an infinite - dimensional integral about which very little can yet be said . by far the most useful results in the path integral approach to ( 2 + 1)-dimensional quantum gravity have come from the covariant first - order action ( 14 ) . the path integral for this action was first fully analyzed in two seminal papers by witten [ 277 , 279 ] , who showed that it reduced to a ratio of determinants that has an elegant topological interpretation as the analytic or ray - singer torsion . the partition function for a closed three - manifold with = 0 takes the form 74\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$${z_m } = { { { { \left\vert { \det \delta _ \varpi ^{(1 ) } } \right\vert}^{1/2 } } } \over { \left\vert { \det \delta _ \varpi ^{(2 ) } } \right\vert}},$$\end{document } where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\delta _ { \bar \omega}^{(n ) } = { d_{\bar \omega}}{d_{\bar \omega } } + * { d_{\bar \omega}}*{d_{\bar \omega}}$\end{document } is the gauge - covariant laplacian acting on n - forms and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}${\bar \omega}$\end{document } is a flat so(2 , 1 ) connection . when m admits more than one such flat connection , equation ( 74 ) must be integrated over the moduli space of such connections . although it was originally derived for closed manifolds , equation ( 74 ) can be extended to manifolds with boundary in a straightforward manner . the path integral then gives a transition function that depends on specified boundary data most simply , the induced spin connection , with some additional restrictions on the normal component of and the triad e [ 283 , 84 ] . for a manifold with the topology , the results agree with those of covariant canonical quantization : the transition amplitude between two surfaces with prescribed spin connections is nonzero only if the holonomies agree . but the path integral can also give transition amplitudes between states on surfaces i and f with different topologies . if we demand that the initial and final surfaces be nondegenerate and spacelike , their topologies are severely restricted : amano and higuchi have shown that i and f must have equal euler numbers . for such manifolds , concrete computations can exploit the topological invariance of the ray - singer torsion . carlip and cosgrove , for example , explicitly compute amplitudes for a transition between a genus three surface and a pair of genus two surfaces . lorentzian path integrals allow us to compute interesting topology - changing amplitudes , in which the universe tunnels from one spatial topology to another . they do not , however , directly address a principle issue of quantum cosmology , the problem of describing the birth of a universe from nothing . here , most of the literature has focused on the hawking s euclidean path integral and the hartle - hawking no boundary proposal , which describes the universe in terms of a path integral over riemannian metrics on manifolds with a single , connected boundary . as in 3 + 1 dimensions , most of the work in 2 + 1 dimensions has concentrated on the saddle point approximation . so far , the main benefit of the lower - dimensional model has been the possibility of treating topology more systematically , revealing interesting effects that are only now being explored in 3 + 1 dimensions . in the hartle - hawking approach to quantum cosmology , the initial wave function of the universe is described by a path integral for a compact manifold m with a single spatial boundary , as in figure 4 . figure 4a manifold m with a single boundary describes the birth of a universe in the hartle - hawking approach to quantum cosmology . a manifold m with a single boundary describes the birth of a universe in the hartle - hawking approach to quantum cosmology . in 2 + 1 dimensions , the selection rules of imply that such a process can be described in lorentzian signature only if ( ) = 0 , that is , only for a torus . moreover , the known examples of such metrics always yield a degenerate metric on . if one allows riemannian signature , on the other hand , such a path integral makes sense for any spatial topology , and if one further requires that be totally geodesic that is , that the extrinsic curvature of vanish one can smoothly join on a lorentzian metric at . wave function 75\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\psi [ h,\varphi { \vert _ \sigma};\sigma ] = \sum\limits_{m:\partial m = \sigma } { \int { [ dg][d\varphi ] } \;\exp \;(- { i_{\rm{e}}}[g,\varphi ] ) } , $ $ \end{document } where the value of the path integral is determined by a specified induced metric h and matter configuration on the boundary . the summation represents a sum over topologies of m ; in the absence of any basis for picking out a preferred topology , all manifolds with a given boundary are assumed to contribute . the wave function is to be interpreted as an amplitude for finding a universe , with a prescribed spatial topology , characterized by an initial this approach finesses the question of initial conditions for the universe by simply omitting an initial boundary , and it postpones the question of the nature of time in quantum gravity : information about time is hidden in the boundary geometry h , but the path integral can be formulated without making a choice of time explicit . the path integral ( 75 ) can not , in general , be evaluated exactly , even in 2 + 1 dimensions . indeed , there are general reasons to expect the expression to be ill - defined : a conformal excitation g eg contributes to ie with the wrong sign , and the action is unbounded below . in the ( 2 + 1)-dimensional lorentzian dynamical triangulation models of section 3.7 , however , it is known that these wrong sign contributions are unimportant ; they are overwhelmed by the much larger number of well - behaved geometries in the path integral . this has led to a suggestion [ 100 , 99 ] that the conformal contribution is canceled by a faddeev - popov determinant ( see also ) , and some preliminary supporting computations have been made in a proper time gauge . assuming that the conformal factor problem is solved , a saddle point evaluation of the path integral is arguably a good approximation . for simplicity , an easy computation shows that the leading contribution to equation ( 75 ) is a sum of terms of the form 76\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$$\exp \;(- { \bar{i } _ e } ) = { \delta _ m}\exp \left({{\rm{sign}}(\lambda){{{\rm{vol}}_{\bar g}(m ) } \over { 4\pi g\hbar \vert \lambda { \vert ^{1/2 } } } } } \right),$$\end{document } where is an einstein metric on m , vol(m ) is the volume of m with the metric rescaled to constant curvature 1 , and the prefactor m is related , as in section 3.10 , to the ray - singer torsion of m. for > 0 , three - manifolds that admit einstein metrics are all elliptic that is , they have constant positive curvature , and can be described as quotients of the three - sphere by discrete groups of isometries . the largest value of vol(m ) comes from the three - sphere itself , and one might expect it to dominate the sum over topologies . as shown in , though , the number of topologically distinct lens spaces with volumes less than vol(s ) grows fast enough that these spaces dominate , leading to a divergent partition function for closed three - manifolds . the implications for the hartle - hawking wave function have not been examined explicitly , but it seems likely that a divergence will appear there as well . for < 0 , three - manifolds that admit einstein metrics are hyperbolic , and the single largest contribution to equation ( 76 ) comes from the smallest such manifold . this contribution has been worked out in detail , for a genus 2 boundary , in . here , too , however , manifolds with larger volumes although individually exponentially suppressed are numerous enough to lead to a divergence in the partition function . in this case , the hartle - hawking wave function has been examined as well , and it has been shown that the wave function acquires infinite peaks at certain specific spatial geometries : again , topologically complicated manifolds whose individual contributions are small occur in large enough numbers to dominate the path integral , and entropy the benefit of restricting to 2 + 1 dimensions here is a bit different from the advantages seen earlier . we are now helped not so much by the simplicity of the geometry ( although this helps in the computation of the prefactors m ) , but by the fact that three - manifold topology is much better understood than four - manifold topology . it is only quite recently that similar results for sums over topologies have been found in four dimensions [ 79 , 80 , 229 , 19 ] . as noted in section 3.6 , recent work on spin foams has also suggested a new nonperturbative approach to evaluating the sum over topologies . building on work by boulatov , freidel and loupre have recently considered a variant of the ponzano - regge model , and have shown that although the sum over topologies diverges , it is borel summable . this result involves a clever representation of a spacetime triangulation as a feynman graph in a field theory on a group manifold , allowing the sum over topologies to be reexpressed as a sum of field theory feynman diagrams . the model considered in is not exactly the ponzano - regge model , and it is not clear that it is really ordinary quantum gravity . moreover , study of the physical meaning of the borel resummed partition function has barely begun . nonetheless , these results suggest that a full treatment of the sum over topologies in ( 2 + 1)-dimensional quantum gravity may not be hopelessly out of reach . there are also indications that string theory might have something to say about the sum over topologies . in particular , the ads / cft correspondence may impose boundary conditions that limit the topologies allowed in the sum . whether such results can be extended to spatially closed manifolds the world is not ( 2 + 1)-dimensional , and ( 2 + 1)-dimensional quantum gravity is certainly not a realistic model of our universe . nonetheless , the ( 2 + 1)-dimensional model reflects many of the fundamental conceptual issues of real world quantum gravity , and work in this field has provided some valuable insights . existence and nonuniqueness perhaps the most important lesson of ( 2 + 1)-dimensional quantum gravity is that general relativity can , in fact , be quantized . while additional ingredients strings , for instance may have their own attractions , they are evidently not necessary for the existence of quantum gravity . more than an existence theorem , though , the ( 2 + 1)-dimensional models also provide a nonuniqueness theorem : many approaches to the quantum theory are possible , and they are not all equivalent . this is perhaps a bit of a disappointment , since many in this field had hoped that once we found a self - consistent quantum theory of gravity , the consistency conditions might be stringent enough to make that theory unique . though , we should not be so surprised : quantum gravity is presumably more fundamental than classical general relativity , and it is not so strange to learn that more than one quantum theory can have the same classical limit . ( 2 + 1)-dimensional gravity as a test bed general relativity in 2 + 1 dimensions has provided a valuable test bed for a number of specific proposals for quantum gravity . some of these are classics the wheeler - dewitt equation , for instance , and reduced phase space quantization while others , like spin foams , lorentzian dynamical triangulations , and covariant canonical quantization , are less well established.we have discovered some rather unexpected features , such as the difficulties caused by spatial diffeomorphism invariance and the consequent nonlocality in wheeler - dewitt quantization , and the necessity of understanding the representations of the group of large diffeomorphisms in almost all approaches . for particular quantization programs , ( 2 + 1)-dimensional models have also offered more specific guidance : special properties of the loop operators ( 55 ) , methods for treating noncompact groups in spin foam models , and properties of the sums over topologies described in section 3.11 have all been generalized to 3 + 1 dimensions . lorentzian dynamical triangulations a particular application of ( 2 + 1)-dimensional gravity as a test bed is important enough to deserve special mention . the program of lorentzian dynamical triangulations described in section 3.7 is a genuinely new approach to quantum gravity . euclidean dynamical triangulations , one might normally be quite skeptical of such a method . but the success in reproducing semiclassical states in 2 + 1 dimensions , although still fairly limited , provides a strong argument that the approach should be taken seriously . one of the deepest conceptual difficulties in quantum gravity has been the problem of reconstructing local , dynamical spacetime from the nonlocal diffeomorphism - invariant observables required by quantum gravity . the notorious problem of time is a special case of this more general problem of observables . as we saw in section 3.4 , ( 2 + 1)-dimensional quantum gravity points toward a solution , allowing the construction of families of local and time - dependent observables that nevertheless commute with all constraints.the idea that frozen time quantum gravity is a heisenberg picture corresponding to a fixed - time - slicing schrdinger picture is a central insight of ( 2 + 1)-dimensional gravity . in practice , though , we have also seen that the transformation between these pictures relies on our having a detailed description of the space of classical solutions of the field equations . we can not expect such a fortunate circumstance to carry over to full ( 3 + 1)-dimensional quantum gravity ; it is an open question , currently under investigation , whether one can use a perturbative analysis of classical solutions to find suitable approximate observables . singularities it has long been hoped that quantum gravity might smooth out the singularities of classical general relativity . although the ( 2 + 1)-dimensional model has not yet provided a definitive test of this idea , some progress has been made . puzio , for example , has shown that a wave packet initially concentrated away from the singular points in moduli space will remain non - singular . on the other hand , minassian has recently demonstrated that quantum fluctuations do not push singularities off to infinity ( as suggested in ) , and that several classically singular ( 2 + 1)-dimensional quantum spacetimes also have singular quantum b - boundaries . another long - standing expectation has been that quantum gravity will lead to discrete , quantized lengths , with a minimum length on the order of the planck length . partial results in quantum geometry and spin foam approaches to ( 2 + 1)-dimensional quantum gravity suggest that this may be true , but also that the problem is a bit subtle [ 197 , 251 , 241 ] . the most recent result in this area relates the spectrum of lengths to representations of the ( 2 + 1)-dimensional lorentz group , which can be discrete or continuous . argue that spacelike intervals are continuous , while timelike intervals are discrete , with a spectrum of the form \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\sqrt { n(n - 1 ) } { \ell _ { \rm{p}}}$\end{document}. the analysis is a bit tricky , since the length a first step towards defining truly invariant operators describing distances between point particles supports this picture , but the results are not yet conclusive . quantum gravity contains two fundamental dimensionful constants , the planck length p and the speed of light c. this has suggested to some that special relativity might itself be altered so that both p and c are constants . this requires a nonlinear deformation of the poincar algebra , and leads to a set of theories collectively called doubly special relativity it has recently been pointed out that ( 2 + 1)-dimensional gravity automatically displays such a deformation [ 197 , 18 , 126 ] . a few attempts have been made to connect this picture to noncommutative spacetime , mainly in the context of point particles [ 197 , 275 , 38 ] , but it seems too early to evaluate them . the answer in 2 + 1 dimensions is unequivocally no : canonical quantization gives a perfectly consistent description of a universe with a fixed spatial topology . on the other hand , the path integrals of section 3.10 seem to allow the computation of amplitudes for tunneling from one topology to another . problems with these topology - changing amplitudes remain , particularly in the regulation of divergent integrals over zero - modes . if these can be resolved , however , we will have to conclude that we have found genuinely and deeply inequivalent quantum theories of gravity.sums over topologiesin conventional descriptions of the hartle - hawking wave function , and in other euclidean path integral descriptions of quantum cosmology , it is usually assumed that a few simple contributions dominate the sum over topologies . the results of ( 2 + 1)-dimensional quantum gravity indicate that such claims should be treated with skepticism ; as discussed in section 3.11 , the sum over topologies is generally dominated by an infinite number of complicated topologies , each individually exponentially suppressed . this is a new and unexpected result , whose implications for realistic ( 3 + 1)-dimensional gravity are just starting to be explored . existence and nonuniqueness perhaps the most important lesson of ( 2 + 1)-dimensional quantum gravity while additional ingredients strings , for instance may have their own attractions , they are evidently not necessary for the existence of quantum gravity . more than an existence theorem , though , the ( 2 + 1)-dimensional models also provide a nonuniqueness theorem : many approaches to the quantum theory are possible , and they are not all equivalent . this is perhaps a bit of a disappointment , since many in this field had hoped that once we found a self - consistent quantum theory of gravity , the consistency conditions might be stringent enough to make that theory unique . in retrospect , though , we should not be so surprised : quantum gravity is presumably more fundamental than classical general relativity , and it is not so strange to learn that more than one quantum theory can have the same classical limit . ( 2 + 1)-dimensional gravity as a test bed general relativity in 2 + 1 dimensions has provided a valuable test bed for a number of specific proposals for quantum gravity . some of these are classics the wheeler - dewitt equation , for instance , and reduced phase space quantization while others , like spin foams , lorentzian dynamical triangulations , and covariant canonical quantization , are less well established . we have discovered some rather unexpected features , such as the difficulties caused by spatial diffeomorphism invariance and the consequent nonlocality in wheeler - dewitt quantization , and the necessity of understanding the representations of the group of large diffeomorphisms in almost all approaches . for particular quantization programs , ( 2 + 1)-dimensional models have also offered more specific guidance : special properties of the loop operators ( 55 ) , methods for treating noncompact groups in spin foam models , and properties of the sums over topologies described in section 3.11 have all been generalized to 3 + 1 dimensions . lorentzian dynamical triangulations a particular application of ( 2 + 1)-dimensional gravity as a test bed is important enough to deserve special mention . the program of lorentzian dynamical triangulations described in section 3.7 is a genuinely new approach to quantum gravity . euclidean dynamical triangulations , one might normally be quite skeptical of such a method . but the success in reproducing semiclassical states in 2 + 1 dimensions , although still fairly limited , provides a strong argument that the approach should be taken seriously . one of the deepest conceptual difficulties in quantum gravity has been the problem of reconstructing local , dynamical spacetime from the nonlocal diffeomorphism - invariant observables required by quantum gravity . the notorious problem of time is a special case of this more general problem of observables . as we saw in section 3.4 , ( 2 + 1)-dimensional quantum gravity points toward a solution , allowing the construction of families of local and time - dependent observables that nevertheless commute with all constraints . quantum gravity is a heisenberg picture corresponding to a fixed - time - slicing schrdinger picture is a central insight of ( 2 + 1)-dimensional gravity . in practice , though , we have also seen that the transformation between these pictures relies on our having a detailed description of the space of classical solutions of the field equations . we can not expect such a fortunate circumstance to carry over to full ( 3 + 1)-dimensional quantum gravity ; it is an open question , currently under investigation , whether one can use a perturbative analysis of classical solutions to find suitable approximate observables . it has long been hoped that quantum gravity might smooth out the singularities of classical general relativity . although the ( 2 + 1)-dimensional model has not yet provided a definitive test of this idea , some progress has been made . puzio , for example , has shown that a wave packet initially concentrated away from the singular points in moduli space will remain non - singular . on the other hand , minassian has recently demonstrated that quantum fluctuations do not push singularities off to infinity ( as suggested in ) , and that several classically singular ( 2 + 1)-dimensional quantum spacetimes also have singular quantum b - boundaries . another long - standing expectation has been that quantum gravity will lead to discrete , quantized lengths , with a minimum length on the order of the planck length . partial results in quantum geometry and spin foam approaches to ( 2 + 1)-dimensional quantum gravity suggest that this may be true , but also that the problem is a bit subtle [ 197 , 251 , 241 ] . the most recent result in this area relates the spectrum of lengths to representations of the ( 2 + 1)-dimensional lorentz group , which can be discrete or continuous . argue that spacelike intervals are continuous , while timelike intervals are discrete , with a spectrum of the form \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{mathrsfs } \usepackage{upgreek } \setlength{\oddsidemargin}{-69pt } \begin{document}$\sqrt { n(n - 1 ) } { \ell _ { \rm{p}}}$\end{document}. the analysis is a bit tricky , since the length a first step towards defining truly invariant operators describing distances between point particles supports this picture , but the results are not yet conclusive . quantum gravity contains two fundamental dimensionful constants , the planck length p and the speed of light c. this has suggested to some that special relativity might itself be altered so that both p and c are constants . this requires a nonlinear deformation of the poincar algebra , and leads to a set of theories collectively called doubly special relativity [ 17 , 185 , 171 ] . it has recently been pointed out that ( 2 + 1)-dimensional gravity automatically displays such a deformation [ 197 , 18 , 126 ] . a few attempts have been made to connect this picture to noncommutative spacetime , mainly in the context of point particles [ 197 , 275 , 38 ] , but it seems too early to evaluate them . does consistent quantum gravity require spatial topology change ? the answer in 2 + 1 dimensions is unequivocally no : canonical quantization gives a perfectly consistent description of a universe with a fixed spatial topology . on the other hand , the path integrals of section 3.10 seem to allow the computation of amplitudes for tunneling from one topology to another . problems with these topology - changing amplitudes remain , particularly in the regulation of divergent integrals over zero - modes . if these can be resolved , however , we will have to conclude that we have found genuinely and deeply inequivalent quantum theories of gravity . sums over topologiesin conventional descriptions of the hartle - hawking wave function , and in other euclidean path integral descriptions of quantum cosmology , it is usually assumed that a few simple contributions dominate the sum over topologies . the results of ( 2 + 1)-dimensional quantum gravity indicate that such claims should be treated with skepticism ; as discussed in section 3.11 , the sum over topologies is generally dominated by an infinite number of complicated topologies , each individually exponentially suppressed . this is a new and unexpected result , whose implications for realistic ( 3 + 1)-dimensional gravity are just starting to be explored . we know immensely more about ( 2 + 1)-dimensional quantum gravity than we did twenty years ago . , it is only quite recently that the general tools developed over the past few years have been brought to bear on particular physical problems the resolution of singularities , for example , and the question of whether space is discrete at the planck scale . a sketchy and rather personal list of open questions would include the following : singularities a key question in quantum gravity is whether quantized spacetime resolves the singularities of classical general relativity . this is a difficult question already classically , it is highly nontrivial to even define a singularity , and the quantum extensions of the classical definitions are far from obvious . this is an area in which ( 2 + 1)-dimensional gravity provides a natural arena , but results so far are highly preliminary [ 227 , 202 ] . sums over topologies another long - standing question in quantum gravity is whether spacetime topology can ( or must ) undergo quantum fluctuations . as we saw in section 3.11 , some real progress has been made in 2 + 1 dimensions . often , though , the results require saddle point approximations , and pick out particular classes of saddle points . the nonperturbative summation techniques discussed at the end of section 3.11 promise much deeper results , and may point toward a measure on the space of topologies analogous to the measure on the space of geometries induced by the dewitt metric . quantized geometry we saw above that there is some evidence for quantization of timelike intervals in ( 2 + 1)-dimensional gravity . a systematic exploration of this issue might teach us a good deal about differences among approaches to quantization . in particular , it would be very interesting to see whether any corresponding result appears in reduced phase space quantization , wheeler - dewitt quantization , or path integral approaches . to address this problem properly , one must introduce genuine observables for quantities such as length and area , either by adding point particles or by looking at shortest geodesics around noncontractible cycles . note that for the torus universe , the moduli can be considered as ratios of lengths , and there is no sign that these need be discrete . this does not contradict the claims of , since the lengths in question are spacelike , but it does suggest an interesting dilemma in euclidean quantum gravity , where spacelike as well as timelike intervals might naturally be quantized . euclidean vs. lorentzian gravity in the chern - simons formalism of section 2.3 , euclidean and lorentzian quantum gravity seem to be dramatically inequivalent : they have different gauge groups , different holonomies , and very different behaviors under the actions of large diffeomorphisms . in the adm approach of section 2.4 , on the other hand this suggests that further study might finally tell us whether euclideanization is merely a technical trick , analogous to wick rotation in ordinary quantum field theory , or whether it gives a genuinely different theory ; and , if the latter , just how different the euclidean and lorentzian theories are . in canonical quantization , a key step would be to relate chern - simons and adm amplitudes in the euclidean theory , perhaps using the methods of section 3.4 . in spin foam and path integral approaches , it might be possible to explicitly compare amplitudes . which approaches are equivalent ? a more general problem is to understand which of the approaches described here are equivalent . in particular , it is not obvious how much of the difference among various methods of quantization can be attributed to operator ordering ambiguities , and how much reflects a deeper inequivalence , as reflected ( for instance ) in different length spectra or different possibilities for topology change . an answer might help us understand just how nonunique quantum gravity in higher dimensions will be . higher genus most of the detailed , explicit results in ( 2 + 1)-dimensional quantum gravity hold only for the torus universe t. as noted in section 2.7 , this topology has some exceptional features , and might not be completely representative . in particular , the relationship between the adm and chern - simons quantizations in section 3.4 relied on a particularly simple operator ordering ; it is not obvious that such an ordering can be found for the higher genus case . an extension to arbitrary genus might be too difficult , but a full treatment of the genus two topology , using the relation to hyperelliptic curves or the sigma model description of , may be possible . it could also be worthwhile to further explore the case of spatially nonorientable manifolds to see whether any important new features arise . some limited progress has been made : for example , there is some evidence that ( 2 + 1)-dimensional gravity is renormalizable in the 1/n expansion when coupled to scalar fields [ 174 , 205 ] . this is apparently no longer the case when gravity is coupled to fermions and a u(1 ) chern - simons gauge theory , although anselmi has argued that if coupling constants are tuned to exact values , renormalizability can be restored , and in fact the theory can be made finite . certain matter couplings in supergravity have been studied [ 104 , 196 ] , and work on circularly symmetric midi - superspace models has led to some surprising results , including unexpected bounds on the hamiltonian [ 27 , 32 , 137 , 53 , 265 , 224 ] . but the general problem of coupling matter remains very difficult , not least because except in the special case of topological matter [ 140 , 85 ] we lose the ability to represent diffeomorphisms as iso(2 , 1 ) gauge transformations.difficult as it is , however , an understanding of matter couplings may be the key to many of the conceptual issues of quantum gravity . one can explore the properties of a singularity , for example , by investigating the reaction of nearby matter , and one can look for quantization of time by examining the behavior of physical clocks . moreover , some of the deep questions of quantum gravity can be answered only in the presence of matter . this idea is an old one [ 109 , 165 , 166 ] , and it gets some support from the boundedness of the hamiltonian in midi - superspace models , but it is only in the context of a full quantum field theory that a final answer can be given . the cosmological constant undoubtedly , the biggest embarrassment in quantum gravity today is the apparent prediction , at least in effective field theory , that the cosmological constant should be some 120 orders of magnitude larger than the observed limit . several attempts have been made to address this problem in the context of ( 2 + 1)-dimensional quantum gravity . first , witten has suggested a novel mechanism by which supersymmetry in 2 + 1 dimensions might cancel radiative corrections to without requiring the equality of superpartner masses , essentially because even if the vacuum is supersymmetric , the asymptotics forbid the existence of unbroken supercharges for massive states [ 281 , 52 ] . this argument requires special features of 2 + 1 dimensions , though , and it is not at all clear that it can be generalized to 3 + 1 dimensions ( although some attempts have been made in the context of deconstruction ) .second , the discovery that the sum over topologies can lead to a divergent partition function has been extended to 3 + 1 dimensions , at least for < 0 , and it has been argued that this behavior might signal a phase transition that could prohibit a conventional cosmology with a negative cosmological constant [ 79 , 80 ] . the crucial case of a positive cosmological constant is not yet understood , however , and if a phase change does indeed occur , its nature is still highly obscure . it may be that the nonperturbative summation over topologies discussed at the end of section 3.11 could cast light on this question.one might also hope that a careful analysis of the coupling of matter in 2 + 1 dimensions could reveal useful details concerning the vacuum energy contribution to , perhaps in a setting that goes beyond the usual effective field theory approach . for example , there is evidence that the matter hamiltonian is bounded above in ( 2 + 1)-dimensional gravity ; perhaps this could cut off radiative contributions to the cosmological constant at an interesting scale . again , ( 2 + 1)-dimensional gravity as a test bed as new approaches to quantum gravity are developed , the ( 2 + 1)-dimensional model will undoubtedly remain important as a simplified test bed . for example , a bit of work has been done on the null surface formulation of classical gravity in 2 + 1 dimensions ; a quantum treatment might be possible , and could tell us more about the utility of this approach in 3 + 1 dimensions . similarly , ( 2 + 1)-dimensional gravity has recently been examined as an arena in which to test for a new partially discrete , constraint - free formulation of quantum gravity . a key question in quantum gravity is whether quantized spacetime resolves the singularities of classical general relativity . this is a difficult question already classically , it is highly nontrivial to even define a singularity , and the quantum extensions of the classical definitions are far from obvious . this is an area in which ( 2 + 1)-dimensional gravity provides a natural arena , but results so far are highly preliminary [ 227 , 202 ] . sums over topologies another long - standing question in quantum gravity is whether spacetime topology can ( or must ) undergo quantum fluctuations . as we saw in section 3.11 , some real progress has been made in 2 + 1 dimensions . often , though , the results require saddle point approximations , and pick out particular classes of saddle points . the nonperturbative summation techniques discussed at the end of section 3.11 promise much deeper results , and may point toward a measure on the space of topologies analogous to the measure on the space of geometries induced by the dewitt metric . we saw above that there is some evidence for quantization of timelike intervals in ( 2 + 1)-dimensional gravity . a systematic exploration of this issue might teach us a good deal about differences among approaches to quantization . in particular , it would be very interesting to see whether any corresponding result appears in reduced phase space quantization , wheeler - dewitt quantization , or path integral approaches . to address this problem properly , one must introduce genuine observables for quantities such as length and area , either by adding point particles or by looking at shortest geodesics around noncontractible cycles . note that for the torus universe , the moduli can be considered as ratios of lengths , and there is no sign that these need be discrete . this does not contradict the claims of , since the lengths in question are spacelike , but it does suggest an interesting dilemma in euclidean quantum gravity , where spacelike as well as timelike intervals might naturally be quantized . euclidean vs. lorentzian gravity in the chern - simons formalism of section 2.3 , euclidean and lorentzian quantum gravity seem to be dramatically inequivalent : they have different gauge groups , different holonomies , and very different behaviors under the actions of large diffeomorphisms . in the adm approach of section 2.4 , on the other hand this suggests that further study might finally tell us whether euclideanization is merely a technical trick , analogous to wick rotation in ordinary quantum field theory , or whether it gives a genuinely different theory ; and , if the latter , just how different the euclidean and lorentzian theories are . in canonical quantization , a key step would be to relate chern - simons and adm amplitudes in the euclidean theory , perhaps using the methods of section 3.4 . in spin foam and path integral approaches which approaches are equivalent ? a more general problem is to understand which of the approaches described here are equivalent . in particular , it is not obvious how much of the difference among various methods of quantization can be attributed to operator ordering ambiguities , and how much reflects a deeper inequivalence , as reflected ( for instance ) in different length spectra or different possibilities for topology change . an answer might help us understand just how nonunique quantum gravity in higher dimensions will be . most of the detailed , explicit results in ( 2 + 1)-dimensional quantum gravity hold only for the torus universe t. as noted in section 2.7 , this topology has some exceptional features , and might not be completely representative . in particular , the relationship between the adm and chern - simons quantizations in section 3.4 relied on a particularly simple operator ordering ; it is not obvious that such an ordering can be found for the higher genus case . an extension to arbitrary genus might be too difficult , but a full treatment of the genus two topology , using the relation to hyperelliptic curves or the sigma model description of , may be possible . it could also be worthwhile to further explore the case of spatially nonorientable manifolds to see whether any important new features arise . some limited progress has been made : for example , there is some evidence that ( 2 + 1)-dimensional gravity is renormalizable in the 1/n expansion when coupled to scalar fields [ 174 , 205 ] . this is apparently no longer the case when gravity is coupled to fermions and a u(1 ) chern - simons gauge theory , although anselmi has argued that if coupling constants are tuned to exact values , renormalizability can be restored , and in fact the theory can be made finite . certain matter couplings in supergravity have been studied [ 104 , 196 ] , and work on circularly symmetric midi - superspace models has led to some surprising results , including unexpected bounds on the hamiltonian [ 27 , 32 , 137 , 53 , 265 , 224 ] . but the general problem of coupling matter remains very difficult , not least because except in the special case of topological matter [ 140 , 85 ] we lose the ability to represent diffeomorphisms as iso(2 , 1 ) gauge transformations . difficult as it is , however , an understanding of matter couplings may be the key to many of the conceptual issues of quantum gravity . one can explore the properties of a singularity , for example , by investigating the reaction of nearby matter , and one can look for quantization of time by examining the behavior of physical clocks . moreover , some of the deep questions of quantum gravity can be answered only in the presence of matter . this idea is an old one [ 109 , 165 , 166 ] , and it gets some support from the boundedness of the hamiltonian in midi - superspace models , but it is only in the context of a full quantum field theory that a final answer can be given . the cosmological constant undoubtedly , the biggest embarrassment in quantum gravity today is the apparent prediction , at least in effective field theory , that the cosmological constant should be some 120 orders of magnitude larger than the observed limit . several attempts have been made to address this problem in the context of ( 2 + 1)-dimensional quantum gravity . first , witten has suggested a novel mechanism by which supersymmetry in 2 + 1 dimensions might cancel radiative corrections to without requiring the equality of superpartner masses , essentially because even if the vacuum is supersymmetric , the asymptotics forbid the existence of unbroken supercharges for massive states [ 281 , 52 ] . this argument requires special features of 2 + 1 dimensions , though , and it is not at all clear that it can be generalized to 3 + 1 dimensions ( although some attempts have been made in the context of deconstruction ) . second , the discovery that the sum over topologies can lead to a divergent partition function has been extended to 3 + 1 dimensions , at least for < 0 , and it has been argued that this behavior might signal a phase transition that could prohibit a conventional cosmology with a negative cosmological constant [ 79 , 80 ] . the crucial case of a positive cosmological constant is not yet understood , however , and if a phase change does indeed occur , its nature is still highly obscure . it may be that the nonperturbative summation over topologies discussed at the end of section 3.11 could cast light on this question . one might also hope that a careful analysis of the coupling of matter in 2 + 1 dimensions could reveal useful details concerning the vacuum energy contribution to , perhaps in a setting that goes beyond the usual effective field theory approach . for example , there is evidence that the matter hamiltonian is bounded above in ( 2 + 1)-dimensional gravity ; perhaps this could cut off radiative contributions to the cosmological constant at an interesting scale . again , ( 2 + 1)-dimensional gravity as a test bed as new approaches to quantum gravity are developed , the ( 2 + 1)-dimensional model will undoubtedly remain important as a simplified test bed . for example , a bit of work has been done on the null surface formulation of classical gravity in 2 + 1 dimensions ; a quantum treatment might be possible , and could tell us more about the utility of this approach in 3 + 1 dimensions . similarly , ( 2 + 1)-dimensional gravity has recently been examined as an arena in which to test for a new partially discrete , constraint - free formulation of quantum gravity . Output:	in three spacetime dimensions , general relativity drastically simplifies , becoming a topological theory with no propagating local degrees of freedom . nevertheless , many of the difficult conceptual problems of quantizing gravity are still present . in this review , i summarize the rather large body of work that has gone towards quantizing ( 2 + 1)-dimensional vacuum gravity in the setting of a spatially closed universe .
PubmedSumm2085	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: based on world health organization statistics , falls are the second leading cause of unintentional injury - related deaths worldwide.1 around 424,000 people globally die every year from unintentional falls , second only to road traffic deaths . injuries sustained from falls are unfortunately quite common in the elderly population ; for instance , in the united kingdom 57% of all injury - related hospitalizations in the elderly are related to falls.2 multifocal glasses in the form of bifocals have been available since 1784 . benjamin franklin converted his glasses into bifocals , allowing him to see which french politician was speaking and at the same time eat his meal.3 multifocal glasses are convenient for activities such as cooking , shopping , and driving . however , multifocal glasses are a known contributor to falls in elderly people.4 when multifocal glasses are worn while negotiating steps , in outdoor environments and around the home , they increase the chance of a fall . given that 52% of elderly people wear multifocal glasses , a large proportion of falls may be preventable.5 why and how multifocal glasses increase falls is not completely understood . the reading portion may affect depth perception and contrast sensitivity;4 additionally , the clear corridor of vision is smaller in progressives due to the large amounts of astigmatism at the edges.6 of particular interest is the prismatic displacement effect of multifocal glasses on correcting postural stability . displacement of the image of an object when viewed through a prism and the illusion of false projection is due to the amount of prismatic displacement it creates . the amount a light ray will deviate when it passes through a prism depends on the apical angle , the index of refraction of the material ( and the material surrounding it ) , the wavelength of the light , and the angle from which the light approaches the prism.7 although the prismatic displacement effect of multifocal glasses is alluded to as a potential cause of falls,4,811 to our knowledge no one has tested this effect . the aim of this study was to 1 ) map the prismatic displacement of a progressive lens , and 2 ) test whether the displacement of a progressive addition lens ( pal ) impairs reaction time and accuracy . a case crossover trial was conducted in the school of physiotherapy balance clinic at the university of otago , dunedin , new zealand . participants reaction times and accuracy were compared while wearing both pals and single ( distance ) vision glasses . initial work illustrated the largest areas of prismatic displacement in a plano , + 4 d powered pal , which was mapped using the nikon projection focimeter ( nikon corporation , tokyo , japan ) . a + 4 d lens was chosen so that the area(s ) of maximal prismatic displacement was more apparent . a plastic mask , 40 mm in diameter , with a grid of holes 3 mm apart ( each hole 1 mm in diameter and 2 mm deep ) , vertically and horizontally ( seven by eleven holes , respectively ) was placed on top of the pal . figure 1 shows the area of the pal that was mapped and the prismatic displacement measurements that were recorded in diopters , and then converted to millimeters of displacement ( diopter 666/100= mm of displacement ) . of note , the measure 666 is the distance in millimeters from the participant to the target they reached for in this study and the value 100 is a factor to convert the displacement to millimeters . both the right and left pals were measured and the right pal was inverted and averaged with the left pals results . the measurements showed that pals had larger displacements at certain points . at the center of the lens the largest mean difference was 8.15 mm ( 95% confidence interval [ ci ] 6.549.75 ) of vertical displacement . at positions at the periphery of the glasses the difference was small , around 2.23 mm ( 95% ci 0.374.09 ) of horizontal displacement ( at 200 mm below center ) and 1.83 mm ( 95% ci 0.523.14 ) of vertical displacement ( at 100 mm below center ) . there was no difference in horizontal displacement at the center and vertical displacement at 300 mm below center between the two glasses ( p=0.271 and p=0.258 ) , respectively ( figures 2 and 3 ) . participants were residents 75-years - old or older from the dunedin and mosgiel communities who had acquired a new or updated prescription for a pal within the last 12 months . exclusion criteria were : 1 ) poorer than 6/12 vision in one eye or 6/12 with binocular vision when corrected with glasses;12 2 ) ocular / retinal problems ( ie , age - related macular degeneration , cataracts , diabetic retinopathy , glaucoma , field defects , and ocular - motor disorders ) ; 3 ) > 2 diopter difference of spherical anisometropia ; 4 ) > 4 diopter of oblique astigmatism ; 5 ) deformities of the arms , neck , or head ; 6 ) inability to stand ; 7 ) suffering from dizziness or a recent injury or medical condition that affected coordination and ability to grasp ; 8) not being able to understand the study requirements ; and 9 ) a mini mental state examination ( mmse)13 score lower than 26 . baseline information was collected from the participant s optometrist s notes and a home visit conducted by the investigator . information was recorded about demographics , living arrangement , medical conditions , medication use , and visual acuity plus physical functioning . the order of the tests of the two types of glasses was randomized with the investigator blinded to the order ; the position of the bar / black line at the maximal or minimal prismatic displacement was randomized . a statistician not involved with the study prepared a computer - generated randomization schedule and opaque envelopes with instructions for the order of glasses to be worn . the schedule was made available on testing day and explained to the participant by a person independent of the study , to preserve blinding of the investigator . numbered tags were placed on the frames of each pair of glasses to allow the participant to wear the glasses in the correct order . each pair of glasses were adjusted and measured so that the eye was 20 mm away from the vertex of the lens . the investigator restated the need for the participant to refrain from describing the glasses as progressives or single distance as it is imperative that the investigator did not know the order during and after the test . it was not possible for an inexperienced observer to tell if the glasses were pals or single vision glasses . because of this the investigator remained blind to the subsequent analysis of the order of glasses . the participants acquired their single vision glasses on testing day so that they did not have time to adapt to this pair of glasses . each new prescription of single vision glasses was identical and matched to the distance part of the individual s pal prescription and frame style . where this was not possible due to a lack of supply , the make , model , size , and style of frame was matched as closely as possible to the participant s chosen frame . the single vision glasses were not matched for scratch resistant lens coating and/or transitional tints , due to budget constraints . the powerlab 4sp data acquisition system and the lab - chart pro version 7 program14 ( adinstruments , dunedin , new zealand ) were used to measure reaction time and accuracy . accuracy measurements were recorded using the labchart program mentioned and a series of nine wire switches under a foam pad ( see figure 4 ) . the labchart program could measure which switch or switches were pushed first and therefore the distance from the black line ( see figure 5 ) . the participant s finger movements when pushing a black line on a foam pad were also recorded by a video camera behind the participant . this was done to ensure only their extraocular eye movements were used to visualize the bar / black line and light emitting diode ( led ) . participants were asked to sit comfortably on a chair with their arms by their side in a neutral position and stare at an led placed 1 meter in front of them . they were instructed to reach out with their dominant hand and grab the whole bar or push the black line with their index finger when the led turned on , as quickly as possible . enough strength to grip the bar or push the black line the bar / black line was placed horizontally and vertically in front and to the dominant hand side of the participant , at a distance of 666 mm . the bar and the black line were attached to a walking frame ( mobilis quad [ cubro ltd , tauranga , new zealand ] , see figure 6 ) . the areas of visual fixation through the glasses were based on the focimeter results from the preliminary investigation , which showed the areas of maximal and minimal prismatic displacement . the coordinates ( 500 , 300 ) , ( 100 , 100 ) , ( 100 , 200 ) , ( 900 , 100 ) , ( 900 , 200 ) , and ( 500 , 0 ) from figures 2 and 3 indicate where participants fixated through their glasses on the bar / black line . the bar / black line was placed at the horizontal and vertical center of the lens ( 500 , 300 ) . the bar / black line was placed vertically only at coordinate ( 100 , 100 ) or ( 900 , 100 ) depending on the participant s dominant hand ; this position was 35 peripherally and 200 mm below center . the bar was placed horizontally at coordinate ( 100 , 200 ) or ( 900 , 200 ) , once again depending on the participant s dominant hand ; this position was 35 peripherally and 100 mm below center . finally , the black line was placed horizontally at coordinate ( 500 , 0 ) , which was 300 mm below center . the main outcome was reaction time when grasping the bar and reaction time plus accuracy when pushing the black line at different points of fixation through the glasses . each test was repeated three times and the fastest time plus its associated accuracy were used in the analyses . data were analyzed using the statistical package for the social sciences version 18 ( ibm corporation , armonk , ny , usa ) and presented as mean ( standard deviation [ sd ] ) , mean difference ( 95% confidence interval [ ci ] ) , or number ( percentage ) . to test whether measures of reaction time differed between the two types of glasses , repeated measures analysis of variance and covariance were used , controlling for length of time since updating their glasses , prismatic displacement , and order of glasses . accuracy measurements were compared for the two types of glasses using the chi - squared mcnemar test . the labchart measurements were verified from the video recordings , to confirm whether the participant pushed the black line . initial work illustrated the largest areas of prismatic displacement in a plano , + 4 d powered pal , which was mapped using the nikon projection focimeter ( nikon corporation , tokyo , japan ) . a + 4 d lens was chosen so that the area(s ) of maximal prismatic displacement was more apparent . a plastic mask , 40 mm in diameter , with a grid of holes 3 mm apart ( each hole 1 mm in diameter and 2 mm deep ) , vertically and horizontally ( seven by eleven holes , respectively ) was placed on top of the pal . figure 1 shows the area of the pal that was mapped and the prismatic displacement measurements that were recorded in diopters , and then converted to millimeters of displacement ( diopter 666/100= mm of displacement ) . of note , the measure 666 is the distance in millimeters from the participant to the target they reached for in this study and the value 100 is a factor to convert the displacement to millimeters . both the right and left pals were measured and the right pal was inverted and averaged with the left pals results . the measurements showed that pals had larger displacements at certain points . at the center of the lens the largest mean difference was 8.15 mm ( 95% confidence interval [ ci ] 6.549.75 ) of vertical displacement . at positions at the periphery of the glasses the difference was small , around 2.23 mm ( 95% ci 0.374.09 ) of horizontal displacement ( at 200 mm below center ) and 1.83 mm ( 95% ci 0.523.14 ) of vertical displacement ( at 100 mm below center ) . there was no difference in horizontal displacement at the center and vertical displacement at 300 mm below center between the two glasses ( p=0.271 and p=0.258 ) , respectively ( figures 2 and 3 ) . participants were residents 75-years - old or older from the dunedin and mosgiel communities who had acquired a new or updated prescription for a pal within the last 12 months . exclusion criteria were : 1 ) poorer than 6/12 vision in one eye or 6/12 with binocular vision when corrected with glasses;12 2 ) ocular / retinal problems ( ie , age - related macular degeneration , cataracts , diabetic retinopathy , glaucoma , field defects , and ocular - motor disorders ) ; 3 ) > 2 diopter difference of spherical anisometropia ; 4 ) > 4 diopter of oblique astigmatism ; 5 ) deformities of the arms , neck , or head ; 6 ) inability to stand ; 7 ) suffering from dizziness or a recent injury or medical condition that affected coordination and ability to grasp ; 8) not being able to understand the study requirements ; and 9 ) a mini mental state examination ( mmse)13 score lower than 26 . baseline information was collected from the participant s optometrist s notes and a home visit conducted by the investigator . information was recorded about demographics , living arrangement , medical conditions , medication use , and visual acuity plus physical functioning . the order of the tests of the two types of glasses was randomized with the investigator blinded to the order ; the position of the bar / black line at the maximal or minimal prismatic displacement was randomized . a statistician not involved with the study prepared a computer - generated randomization schedule and opaque envelopes with instructions for the order of glasses to be worn . the schedule was made available on testing day and explained to the participant by a person independent of the study , to preserve blinding of the investigator . numbered tags were placed on the frames of each pair of glasses to allow the participant to wear the glasses in the correct order . each pair of glasses were adjusted and measured so that the eye was 20 mm away from the vertex of the lens . the investigator restated the need for the participant to refrain from describing the glasses as progressives or single distance as it is imperative that the investigator did not know the order during and after the test . it was not possible for an inexperienced observer to tell if the glasses were pals or single vision glasses . because of this the investigator remained blind to the subsequent analysis of the order of glasses . the participants acquired their single vision glasses on testing day so that they did not have time to adapt to this pair of glasses . each new prescription of single vision glasses was identical and matched to the distance part of the individual s pal prescription and frame style . where this was not possible due to a lack of supply , the make , model , size , and style of frame was matched as closely as possible to the participant s chosen frame . the single vision glasses were not matched for scratch resistant lens coating and/or transitional tints , due to budget constraints . the powerlab 4sp data acquisition system and the lab - chart pro version 7 program14 ( adinstruments , dunedin , new zealand ) were used to measure reaction time and accuracy . accuracy measurements were recorded using the labchart program mentioned and a series of nine wire switches under a foam pad ( see figure 4 ) . the labchart program could measure which switch or switches were pushed first and therefore the distance from the black line ( see figure 5 ) . the participant s finger movements when pushing a black line on a foam pad were also recorded by a video camera behind the participant . this was done to ensure only their extraocular eye movements were used to visualize the bar / black line and light emitting diode ( led ) . participants were asked to sit comfortably on a chair with their arms by their side in a neutral position and stare at an led placed 1 meter in front of them . they were instructed to reach out with their dominant hand and grab the whole bar or push the black line with their index finger when the led turned on , as quickly as possible . enough strength to grip the bar or push the black line the bar / black line was placed horizontally and vertically in front and to the dominant hand side of the participant , at a distance of 666 mm . the bar and the black line were attached to a walking frame ( mobilis quad [ cubro ltd , tauranga , new zealand ] , see figure 6 ) . the areas of visual fixation through the glasses were based on the focimeter results from the preliminary investigation , which showed the areas of maximal and minimal prismatic displacement . the coordinates ( 500 , 300 ) , ( 100 , 100 ) , ( 100 , 200 ) , ( 900 , 100 ) , ( 900 , 200 ) , and ( 500 , 0 ) from figures 2 and 3 indicate where participants fixated through their glasses on the bar / black line . the bar / black line was placed at the horizontal and vertical center of the lens ( 500 , 300 ) . the bar / black line was placed vertically only at coordinate ( 100 , 100 ) or ( 900 , 100 ) depending on the participant s dominant hand ; this position was 35 peripherally and 200 mm below center . the bar was placed horizontally at coordinate ( 100 , 200 ) or ( 900 , 200 ) , once again depending on the participant s dominant hand ; this position was 35 peripherally and 100 mm below center . finally , the black line was placed horizontally at coordinate ( 500 , 0 ) , which was 300 mm below center . the main outcome was reaction time when grasping the bar and reaction time plus accuracy when pushing the black line at different points of fixation through the glasses . each test was repeated three times and the fastest time plus its associated accuracy were used in the analyses . data were analyzed using the statistical package for the social sciences version 18 ( ibm corporation , armonk , ny , usa ) and presented as mean ( standard deviation [ sd ] ) , mean difference ( 95% confidence interval [ ci ] ) , or number ( percentage ) . to test whether measures of reaction time differed between the two types of glasses , repeated measures analysis of variance and covariance were used , controlling for length of time since updating their glasses , prismatic displacement , and order of glasses . accuracy measurements were compared for the two types of glasses using the chi - squared mcnemar test . the labchart measurements were verified from the video recordings , to confirm whether the participant pushed the black line . thirty - one participants were recruited for the study ( see figure 7 ) and their characteristics are summarized in table 1 . when wearing pals , participants had a significantly faster reaction time compared to when they were wearing single vision glasses . this improved reaction time was noted when localizing the bar / line through two areas : 1 ) at the center of the lens , when grasping a horizontal bar , pals were faster by 101 ms , p=0.011 ( repeated measures for order effect , time to update their pals , and prismatic displacement ) ; 2 ) at 300 mm below center , when pushing a horizontal black line , pals were faster by 80 ms , p=0.007 . this study found no statistically significant differences for reaction time for pushing a black line and grasping a bar at the other positions tested , fixating through the two different glasses ( see table 2 ) . figures 8 and 9 represent graphical interpretations of the reaction time measurements and illustrate the significant differences between the two glasses in the central visual axis . table 3 and figure 10 illustrate accuracy at the specific points that participants localized through the lens . differences between measurements did not reach statistical significance ; however , participants showed a tendency to be less accurate when wearing pals compared with single vision glasses at the edges of the glasses . at the center of the lens , pals showed a tendency to be more accurate when the black line was placed vertically ; accuracy between the two glasses was similar when the black line was placed horizontally ( see table 3 and figure 10 ) . the video tape recordings of participants pushing the foam pad with a black line on it endorsed the findings of the switches using the labchart and powerlab data acquisition systems . it has been shown that elderly people who can grasp a support stick at a distance further away from them have a lower incidence of falls.18 the ability to reach out and grab a support with accuracy and without hesitation is important in a fall . to our knowledge this is the first study to look at the prismatic displacement effect on reaction time and accuracy when an elderly person extends their arm to touch a target . in our study , participants were faster at grasping a bar and pushing a black line placed in the central visual axis when wearing pals compared with single vision glasses . this primary finding is consistent with the study by lord et al on falls among long - term multifocal wearers.4 one of the secondary measures of lord et al s trial was reaction time with a light as a stimulus and participants wearing their preferred eyewear . the study showed that participants who wore multifocal glasses had a tendency to react faster compared with non - multifocal glasses wearers ; mean ( sd ) time to react was 276 ( 57 ) and 288 ( 58 ) ms , respectively . results from our study showed that reaction time tested in the majority of areas of fixation and all of the accuracy measurements were non - significant between the two types of glasses , which is consistent with the visible trial . the visible trial suggests that swapping multifocal glasses with single vision distance glasses had no overall effect on fall- prevention ( incidence rate ratio [ irr ] 0.92 [ 95% ci 0.731.16]).8 additionally , preplanned subgroup analysis showed that by replacing multifocal with single vision glasses there was a significant increase in falls in older frail people compared with controls who continued wearing their multifocal glasses , especially in an outdoor ( unfamiliar ) environment ( irr 1.56 ; 95% ci 1.112.19).8 however , the visible trial did illustrate through the same subgroup analysis that active older people benefit from replacing their multifocal glasses with single vision glasses ( irr 0.60 ; 0.420.87 ) , specifically when in an outdoor environment ( irr 0.61 ; 95% ci 0.420.87).8 anecdotal evidence suggests that when first prescribed multifocal glasses , the person experiences a significant undulating effect , which decreases with time . the sensation of inappropriate motion ( oscillopsia ) seems to be most present around the initial time of adaptation.19 furthermore , it has been shown that people can adjust to glasses that invert the image seen ; once the glasses are removed , they simply adjust back within 30 minutes.20 it is suggested that this is due to the brain accommodating the change , through recalibration of the proprioception receptors in the upper and lower limbs and extra retinal eye movements ( egocentric and oculocentric visual directions ) , rather than a correction of the vision . with that in mind , the participants in this trial may have adapted to their pals , having worn them for a mean of 228 days ( sd 146 days ) , and were thus able to react more quickly than with a new pair of single vision glasses . in changing glasses and thus reversing the prismatic effect , this effect was not specifically tested in our study and it is not unreasonable to postulate that the effect of replacing pals with single vision glasses may work in reverse , but this would need further study . the findings from the study by cumming et al suggest caution should be taken with a sudden change in older people s vision.21 in this study the intervention was aimed at improving vision ( eg , arranging for new eyeglasses ) and this increased rather than decreased the number of falls by participants over a 12 month period . the majority of the intervention participants that required visual improvement acquired a new pair of glasses ( 92 of 146 , 63% ) . furthermore , most participants in the intervention arm had their glasses changed 13 months before the trial started ( 209 of 309 , 68% ) ; perhaps a more gradual process of updating is required to allow for adjustment to the change in vision . the increase in fall rate seen in the intervention arm was most prominent within the first 3 months of updating their glasses . however , the length of time needed to adapt to multifocal glasses has not been defined . when an older person walks into an unfamiliar environment , he or she unconsciously remembers the potential support objects in space in that room , spatial memory known as stored central visual spatial information;22 it is possible that this information is stored incorrectly when replacing glasses with different prismatic displacements . therefore , when a trip , slip , or loss of balance does occur and the older person needs to rely on that stored information , they may miss the support object . according to our study , this would be true only for the vertical displacement in the center and the horizontal bar placed at this position for reaction time . our results found a difference in reaction time when the black line was placed at a position of 300 mm down from center , but this can not be explained by the prismatic displacement effect as there was no statistically significant vertical displacement between the two glasses at this point ( p=0.258 ) . alternatively the result at this position could be explained by the reading portion of pals because the black line was placed 666 mm away from the participants . this is the optimal focus that is , around 600 mm for the reading power of multifocal glasses.4 the mean focal distance of participants pal reading portion in this study was 395 mm ( range , 500333 mm ) . therefore , the reading portion brought the line closer to the participant and the resolution may be clearer , enabling the participant to push the line more quickly . there are alternative explanations for why multifocal glasses have an effect on falls in the elderly population . timmis et al s work suggests that participants wearing single vision glasses had more control when stepping down from a block height when compared with individuals who were well adapted to multifocal lenses.9 the study compared kinematic mechanics in 20 older people and found that those participants wearing single vision glasses had reduced vertical center of mass velocity , increased single - limb support time , and reduced peak ankle and knee angular velocities . this suggested that landing occurred in a more controlled manner when wearing single vision glasses . another study by elliott et al involved ten elderly participants and their stepping kinematics onto a raised surface when wearing a positive or negative blurring.23 the study found that stepping movements were refected by the magnification effect of the lens on the position and size of the step . the step looked closer with a positive lens , which influenced the participant s trailing foot to be placed further away and increased their leading vertical toe clearance . with a negative lens the reverse was true . they recommend that partial changes in correction should be prescribed and that updating glasses should therefore occur more frequently . replacing multifocal glasses with single vision glasses could be harmful for other reasons . in the visible trial , secondary measures were recorded of non - fall - related injuries such as lacerations , lifting or twisting injuries , burns or scalds , eye injuries , collisions , and pedestrian injuries.8 they found that 26% of participants replacing multifocal glasses with single vision glasses had a non - fall related injury , compared with 17% of the control group ( p=0.01 ) . this effect could be due to the convenience of multifocal glasses when carrying out everyday tasks such as cooking , driving , shopping , or similar activities . the study population were healthy , active participants , which may not enable generalizability to the older population at greatest risk of a fall . the technique used to map the prismatic displacement was limited to the mask used to map the section of the lens . there was a balance between mapping the largest prismatic displacement at the edge of the glasses , while testing had to occur within the spectacle frame . the general trend was for the prismatic displacement to increase the further out from the center of the lens tested . therefore , if the targets used to test accuracy and reaction time were placed in a more peripheral position , they could have potentially shown a larger effect compared with the areas of fixation that were tested . however , this was not possible due to the frames of the participants glasses . although the frames of the new single vision glasses were matched to the participant s pals , there could have been a slight differences in tilt , vertex , adjustment , and size of the new pair of glasses . another limitation with this study is that a light stimulus was used instead of a trip , slip , or loss of balance . therefore , there was no heightened awareness through a fight - or - flight response . a one - off trial which involves a perturbation rather than a light stimulus would possibly be a better method to simulate a fall.22,24 our study expands on the paper by lord et al4 and examines the influence on response to a loss of balance when replacing multifocal glasses with single vision ( distance ) glasses . to our knowledge , this is the first study to test prismatic displacement and fall risk in older people . in conclusion , this study suggests that updating glasses should possibly occur more often in the elderly population to avoid abrupt changes in prismatic displacement . Output:	backgroundmultifocal glasses ( bifocals , trifocals , and progressives ) increase the risk of falling in elderly people , but how they do so is unclear . to explain why glasses with progressive addition lenses increase the risk of falls and whether this can be attributed to false projection , this study aimed to 1 ) map the prismatic displacement of a progressive lens , and 2 ) test whether this displacement impaired reaction time and accuracy.methodsthe reaction times of healthy 75-year - olds ( 31 participants ) were measured when grasping for a bar and touching a black line . participants performed each test twice , wearing their progressives and new , matched single vision ( distance ) glasses in random order . the line and bar targets were positioned according to the maximum and minimum prismatic displacement effect through the progressive lens , mapped using a focimeter.resultsprogressive spectacle lenses have large areas of prismatic displacement in the central visual axis and edges . reaction time was faster for progressives compared with single vision glasses with a centrally - placed horizontal grab bar ( mean difference 101 ms , p=0.011 [ repeated measures analysis ] ) and a horizontal black line placed 300 mm below center ( mean difference 80 ms , p=0.007 ) . there was no difference in accuracy between the two types of glasses.conclusionolder people appear to adapt to the false projection of progressives in the central visual axis . this adaptation means that swapping to new glasses or a large change in prescription may lead to a fall . frequently updating glasses may be more beneficial .
PubmedSumm2086	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: in conclusion , we report a ketyl olefin coupling for the preparation of substituted 3-benzylchroman-4-ols , promoted by visible light photoredox catalysis . importantly , we uncovered trialkylamines as a cheap and readily available alternative to the previously reported electron proton donor system consisting of hantzsch ester / brnsted acid . by employing the trialkylamine / photocatalyst system , we have been able to develop an efficient intramolecular ketyl the formal hydroacylation protocol provides the chromanol derivatives in good yield under mild reaction conditions and with low catalyst loadings . all reactions were performed with oven - dried glassware and under an inert atmosphere ( argon ) unless otherwise stated . acetonitrile was distilled from calcium hydride and stored over 4 molecular sieves under nitrogen / argon atmosphere . chromatographic purification of products was carried out using silica gel ( 230400 mesh ) . thin - layer chromatography was carried out using aluminum plates coated with silica ( 230400 mesh ) which were visualized under uv light ( 254 nm ) or by staining with aqueous potassium permanganate solutions or vanillin alcoholic solution . h nmr spectra were recorded in deuterated solvents on spectrometers at 300 , 400 , or 600 mhz with residual protic solvent as the internal standard . c nmr spectra were recorded in deuterated solvents on spectrometers at 75 , 101 , or 151 mhz with the central peak of the deuterated solvent as the internal standard . chemical shifts ( ) are given in parts per million ( ppm ) , and coupling constants ( j ) are given in hertz ( hz ) rounded to the nearest 0.1 hz . the h nmr spectra are reported as downfield from tetramethylsilane ( multiplicity , coupling constant j , number of protons ) . the c nmr spectra are reported as . for the f - containing compound ( 2c ) , a simultaneously proton and fluorine decoupled c nmr spectrum was recorded . assignments are aided by the use of dept-135 , cosy and hmqc spectra where necessary . low resolution mass spectra ( ei / ci ) were recorded on a mass spectrometer with ( ei ) or ( ci ) detectors . high - resolution mass spectrometry ( hrms ) was performed by ei using a double - focusing mass spectrometer and by esi using a hybrid ion - trap mass spectrometer . a schlenk tube was charged with aldehyde 1 ( 1 equiv ) , catalyst pc 3 ( 1 or 2.5 mol % ) , and degassed dipea ( 2.5 equiv ) . the vial was placed in a 100 ml beaker containing a blue led strip glued on the inner wall , and the reaction mixture was stirred for 15 h. after this time , the solvent was evaporated , and the reaction mixture was purified on silica gel ( gradient pentane / etoac 95:5 ) . the title compound was synthesized according to the general procedure employing 1a ( 0.12 mmol , 28.6 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 80% ( 23 mg ) ; anti / syn 1:1 ; h nmr ( 600 mhz , cdcl3 ) 7.347.18 ( m , 14h ) , 6.96 ( td , j = 7.5 , 0.9 hz , 1h ) , 6.90 ( dd , j = 10.6 , 4.2 hz , 2h ) , 6.85 ( d , j = 8.5 hz , 1h ) , 4.51 ( dd , j = 9.4 , 3.1 hz , 2h ) , 4.23 ( dd , j = 11.1 , 2.6 hz , 1h ) , 4.144.05 ( m , 2h ) , 3.98 ( dd , j = 11.1 , 4.2 hz , 1h ) , 2.89 ( dd , j = 13.8 , 8.4 hz , 1h ) , 2.752.64 ( m , 2h ) , 2.54 ( dd , j = 13.8 , 9.3 hz , 1h ) , 2.402.29 ( m , 1h ) , 2.282.17 ( m , 1h ) , 1.92 ( bs , 1h ) , 1.74 ( bs , 1h ) ; c nmr ( 151 mhz , cdcl3 ) 154.3 , 154.2 , 139.2 , 139.1 , 130.2 , 130.1 , 129.9 , 129.8 , 129.1 ( 2c ) , 128.6 , 128.5 , 126.3 ( 2c ) , 124.1 , 123.2 , 120.9 , 120.5 , 116.9 ( 2c ) , 67.6 , 64.9 ( 2c ) , 64.6 , 41.5 , 40.0 , 34.6 , 32.8 ; ft - ir max(atr ) cm 3372 , 2922 , 1582 , 1485 , 1450 , 1302 , 1264 , 1222 , 1119 , 1074 , 1039 , 909 , 748 , 698 ; m / z ( ei ) 240 ( [ m ] , 100 ) , 91 ( [ phch2 ] , 28 ) ; hrms ( esi ) calcd for [ c16h16o2 + na ] 263.10425 , found 263.10419 . the title compound was synthesized according to the general procedure employing 1b ( 0.12 mmol , 38.1 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 50% ( 19.1 mg ) ; anti / syn 1.1:1 ; h nmr ( 400 mhz , cd3od ) 7.45 ( d , j = 2.4 hz , 1h , syn ) , 7.34 ( d , j = 3.1 hz , 1.1 1h , anti ) , 7.317.13 ( m , 12.6h ) , 6.736.65 ( m , 2.1h ) , 4.47 ( d , j = 3.4 hz , 1h , syn ) , 4.36 ( d , j = 5.0 hz , 1.1 1h , anti ) , 4.13 ( dd , j = 11.2 , 2.8 hz , 1.1 1h , anti ) , 4.033.94 ( m , 2h , syn ) , 3.89 ( dd , j = 11.1 , 5.5 hz , 1.1 1h , anti ) , 2.87 ( dd , j = 13.6 , 7.1 hz , 1h , syn ) , 2.76 ( dd , j = 13.8 , 6.1 hz , 1h , syn ) , 2.55 ( dd , j = 13.6 , 8.3 hz , 1.1 1h , anti ) , 2.45 ( dd , j = 13.7 , 9.3 hz , 1.1 1h , anti ) , 2.252.17 ( m , 1h , syn ) , 2.162.08 ( m , 1.1 1h , anti ) ; c nmr ( 101 mhz , cd3od ) 153.5 ( 2c ) , 139.4 , 139.2 , 132.4 ( 2c ) , 131.60 ( 2c ) , 128.8 , 128.7 , 128.1 ( 2c ) , 127.0 , 126.1 , 126.0 , 125.8 , 118.1 ( 2c ) , 111.9 , 111.5 , 66.4 , 65.0 , 64.8 , 64.2 , 41.5 , 40.2 , 34.2 , 32.1 ; ft - ir max(atr ) cm 2926 , 1479 , 1407 , 1260 , 1224 , 1191 , 1085 , 1055 , 1021 , 819 , 748 , 699 ; m / z ( ei ) 319 ( [ m ] , 9 ) , 91 ( [ phch2 ] , 100 ) ; hrms ( ei ) calcd for [ c16h15o2br ] 318.02499 , found 318.02492 . the title compound was synthesized according to the general procedure employing 1c ( 0.12 mmol , 30.8 mg ) , 1 mol % ( 1.1 mg ) of pc 3 and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 43% ( 13.2 mg ) ; mp = 102104 c ; anti / syn 1.1:1 ; h nmr ( 600 mhz , cd3od ) 7.317.23 ( m , 6.5 h ) , 7.217.15 ( m , 4.3 h ) , 7.06 ( dd , j = 9.0 , 3.1 hz , 1.1 1h , anti ) , 6.96 ( dd , j = 8.9 , 3.1 hz , 1h , syn ) , 6.89 ( dtd , j = 11.6 , 8.5 , 3.1 hz , 2h ) , 6.786.71 ( m , 2h ) , 4.48 ( d , j = 3.5 hz , 1h , syn ) , 4.37 ( d , j = 5.1 hz , 1.1 1h , anti ) , 4.12 ( dd , j = 11.1 , 2.8 hz , 1.1 1h , anti ) , 4.013.94 ( m , 2h , syn ) , 3.87 ( dd , j = 11.1 , 5.6 hz , 1.1 1h , anti ) , 2.88 ( dd , j = 13.6 , 7.2 hz , 1h , syn ) , 2.79 ( dd , j = 13.8 , 6.0 hz , 1.1 1h , anti ) , 2.56 ( dd , j = 13.6 , 8.3 hz , 1h , syn ) , 2.46 ( dd , j = 13.8 , 9.4 hz , 1.1 1h , anti ) , 2.242.17 ( m , 1h , syn ) , 2.162.11 ( m , 1.1 1h , anti ) ; f nmr ( 564 mhz , cd3od ) 125.8 , 126.4 ; c nmr ( 101 mhz , cd3od ) 156.9 , 156.6 , 150.4 , 150.3 , 139.5 , 139.2 , 128.8 , 128.7 , 128.1 , 128.0 , 125.9 ( 2c ) , 125.8 , 125.0 , 117.2 , 117.1 , 115.5 , 115.4 , 115.3 ( 2c ) , 66.8 , 65.0 , 64.7 , 64.4 , 41.6 , 40.3 , 34.2 , 32.1 ; ft - ir max(atr ) cm 3370 , 2926 , 2493 , 1488 , 1436 , 1251 , 1201 , 1027 , 736 , 698 ; m / z ( ei ) 258 ( [ m ] , 48 ) , 91 ( [ phch2 ] , 100 ) ; hrms ( ei ) calcd for [ c16h15o2f ] 258.10506 , found 258.10537 . the title compound was synthesized according to the general procedure employing 1d ( 0.12 mmol , 35.6 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 51% ( 18.2 mg ) ; anti / syn 1.3:1 ; h nmr ( 400 mhz , cd3od ) 8.05 ( d , j = 2.1 hz , 1.3 1h , anti ) , 7.92 ( d , j = 2.2 hz , 1h , syn ) , 7.80 ( ddd , j = 14.4 , 8.6 , 2.2 hz , 2.3h ) , 7.327.11 ( m , 11.5h ) , 6.81 ( dd , j = 14.4 , 8.6 hz , 2.3h ) , 4.52 ( d , j = 3.2 hz , 1h , syn ) , 4.42 ( d , j = 4.9 hz , 1.3 1h , anti ) , 4.21 ( dd , j = 11.2 , 2.9 hz , 1.3 1h , anti ) , 4.113.99 ( m , 2h , syn ) , 3.96 ( dd , j = 11.1 , 5.3 hz , 1.3 1h , anti ) , 3.84 ( s , 1.3 3h , anti ) , 3.81 ( s , 3h , syn ) , 2.88 ( dd , j = 13.6 , 7.1 hz , 1h , syn ) , 2.75 ( dd , j = 13.7 , 6.1 hz , 1.3 1h , anti ) , 2.56 ( dd , j = 13.6 , 8.3 hz , 1h , syn ) , 2.43 ( dd , j = 13.7 , 9.3 hz , 1.3 1h , anti ) , 2.262.13 ( m , 2.3h ) ; c nmr ( 101 mhz , cd3od ) 167.0 ( 2c ) , 158.6 ( 2c ) , 139.4 , 139.1 , 132.3 , 132.2 , 130.4 , 130.3 , 128.8 , 128.7 , 128.1 ( 2c ) , 126.0 , 125.9 , 124.8 , 123.9 , 122.1 , 121.7 , 116.3 ( 2c ) , 66.4 , 65.4 , 65.2 , 64.2 , 51.0 , 50.9 , 41.4 , 40.2 , 34.2 , 32.2 ; ft - ir max(atr ) cm 2949 , 1711 , 1611 , 1496 , 1436 , 1253 , 1192 , 1125 , 1012 , 767 , 700 ; m / z ( ei ) 298 ( [ m ] , 12 ) , 91 ( [ phch2 ] , 40 ) ; hrms ( ei ) calcd for [ c18h18o4 ] 298.11996 , found 298.12009 . the title compound was synthesized according to the general procedure employing 1e ( 0.12 mmol , 42.4 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 44% ( 18.7 mg ) ; anti / syn 1.6:1 ; h nmr ( 600 mhz , cdcl3 ) 7.497.15 ( m , 15.6h ) , 7.08 ( dd , j = 8.8 , 2.6 hz , 1.6 1h , anti ) , 6.99 ( dd , j = 8.8 , 2.6 hz , 1h , syn ) , 6.80 ( d , j = 8.8 hz , 1.6 1h , anti ) , 6.76 ( d , j = 8.8 hz , 1h , syn ) , 6.456.24 ( m , 2.6h ) , 4.524.41 ( m , 2.6h ) , 4.19 ( dd , j = 11.0 , 1.9 hz , 1.6 1h , anti ) , 4.124.01 ( m , 2h , syn ) , 3.92 ( dd , j = 11.0 , 3.9 hz , 1.6 1h , anti ) , 2.86 ( dd , j = 13.5 , 8.6 hz , 1h , syn ) , 2.732.61 ( m , 2.6h ) , 2.51 ( dd , j = 13.7 , 9.5 hz , 1.6 1h , anti ) , 2.312.26 ( m , 1h , syn ) , 2.212.17 ( m , 1.6 1h , anti ) , 1.911.62 ( overlapping of two bs , 2.6h ) 1.51 ( s , 1.6 9h , anti ) , 1.48 ( s , 9h , syn ) ; c nmr ( 151 mhz , cdcl3 ) 153.2 , 153.1 , 150.3 , 150.2 , 139.2 , 139.1 , 131.4 , 131.3 , 131.0 ( 2c ) , 129.1 ( 2c ) , 128.5 ( 2c ) , 126.3 ( 2c ) , 124.4 , 123.5 , 121.2 , 120.7 , 117.2 , 117.1 , 80.4 ( 2c ) , 67.6 , 65.1 , 64.8 , 64.6 , 41.3 , 40.0 , 34.6 , 32.7 , 28.4 , 28.3 ; ft - ir max(atr ) cm : 3350 , 2981 , 2932 , 1696 , 1499 , 1454 , 1367 , 1309 , 1241 , 1218 , 1163 , 1023 , 822 , 733 , 699 ; m / z ( ei ) 298 ( [ m ( ch3)3c ] , 100 ) , 91 ( [ phch2 ] , 67 ) ; hrms ( ei ) calcd for [ c16h16o2n1 ] ( [ m boc ] ) 254.11756 , found 254.11706 . the title compound was synthesized according to the general procedure employing 1f ( 0.12 mmol , 37.7 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 61% ( 23 mg ) ; anti / syn 1.2:1 ; h nmr ( 600 mhz , cd3od ) 7.487.44 ( m , 1.2 1h , anti ) , 7.447.40 ( m , 1h , syn ) , 7.377.05 ( m , 24.2h ) , 7.006.94 ( m , 1.2 1h , anti ) , 6.926.86 ( m , 1h , syn ) , 4.49 ( d , j = 2.7 hz , 1h , syn ) , 4.44 ( d , j = 4.1 hz , 1.2 1h , anti ) , 4.15 ( dd , j = 11.0 , 2.6 hz , 1.2 1h , anti ) , 4.00 ( t , j = 10.9 hz , 1h , syn ) , 3.92 ( dd , j = 10.6 , 3.5 hz , 1h , syn ) , 3.86 ( dd , j = 11.0 , 4.5 hz , 1.2 1h , anti ) , 2.85 ( dd , j = 13.5 , 7.5 hz , 1h , syn ) , 2.70 ( dd , j = 13.7 , 6.4 hz , 1.2 1h , anti ) , 2.55 ( dt , j = 18.0 , 9.0 hz , 1h , syn ) , 2.512.44 ( m , 1.2 1h , anti ) , 2.252.18 ( m , 1h , syn ) , 2.172.12 ( m , 1.2 1h , anti ) ; c nmr ( 151 mhz , cd3od ) 151.1 ( 2c ) , 139.6 , 139.4 , 138.5 , 138.4 , 130.2 , 130.1 , 129.8 ( 2c ) , 129.7 ( 2c ) , 129.2 ( 2c ) , 128.8 , 128.7 , 128.1 ( 2c ) , 127.5 ( 2c ) , 126.4 ( 2c ) , 125.9 , 125.8 , 125.1 , 123.9 , 120.2 , 119.9 , 67.1 , 64.8 , 64.6 , 64.5 , 41.5 , 40.3 , 34.3 , 32.5 ; ft - ir max(atr ) cm 1594 , 1496 , 1469 , 1430 , 1219 , 1017 , 754 , 697 ; m / z ( ei ) 316 ( [ m ] , 63% ) , 91 ( [ phch2 ] , 27% ) ; hrms ( esi ) calcd for [ c22h20o2 + na ] 339.13555 , found 339.13525 . the title compound was synthesized according to the general procedure employing 1 g ( 0.12 mmol , 30.3 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 56% ( 17.1 mg ) ; anti / syn 1.4:1 ; h nmr ( 600 mhz , cd3od ) 7.307.21 ( m , 7.5h ) , 7.207.12 ( m , 5.5h ) , 7.057.02 ( m , 2.4h ) , 6.99 ( d , j = 7.3 hz , 1.4 1h , anti ) , 6.80 ( t , j = 7.5 hz , 1h , syn ) , 6.73 ( t , j = 7.5 hz , 1.4 1h , anti ) , 4.45 ( d , j = 3.2 hz , 1.4 1h , anti ) , 4.38 ( d , j = 4.2 hz , 1h , syn ) , 4.19 ( dd , j = 11.0 , 2.6 hz , 1h , syn ) , 4.064.02 ( m , 1.4 2h , anti ) , 3.94 ( dd , j = 11.0 , 4.6 hz , 1h , syn ) , 2.87 ( dd , j = 13.7 , 7.4 hz , 1.4 1h , anti ) , 2.68 ( dd , j = 13.7 , 6.5 hz , 1h , syn ) , 2.58 ( dd , j = 13.6 , 8.1 hz , 1.4 1h , anti ) , 2.45 ( dd , j = 13.7 , 9.1 hz , 1h , syn ) , 2.16 ( s , 3h , syn ) , 2.12 ( s , 1.4 3h , anti ) oh signals not observed ; c nmr ( 151 mhz , cd3od ) 152.3 , 152.2 , 139.7 , 139.5 , 129.9 , 129.8 , 128.8 , 128.7 , 128.1 ( 2c ) , 127.9 , 127.8 , 125.8 ( 2c ) , 125.2 ( 2c ) , 123.9 , 122.7 , 119.6 , 119.3 , 66.9 , 64.7 , 64.6 , 64.4 , 41.8 , 40.6 , 34.3 , 32.6 , 14.8 ( 2c ) ; ft - ir max(atr ) cm 3352 , 2919 , 1598 , 1472 , 1258 , 1210 , 1092 , 1023 , 747 , 698 ; m / z ( ei ) 254 ( [ m ] , 59 ) , 253 ( [ m 1 ] , 100 ) , 91 ( [ phch2 ] , 39 ) ; hrms ( ei ) calcd for [ c17h18o2 ] 254.13013 , found 254.13046 . the title compound was synthesized according to the general procedure employing 1h ( 0.12 mmol , 42.1 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 88% ( 37.5 mg ) ; anti / syn 1.2:1 ; h nmr ( 400 mhz , cd3od ) 7.337.07 ( m , 15.4h ) , 4.48 ( d , j = 3.5 hz , 1h , syn ) , 4.39 ( d , j = 3.3 hz , 1.2 1h , anti ) , 4.13 ( dd , j = 10.9 , 2.5 hz , 1.2 1h , anti ) , 4.033.94 ( m , 2h , syn ) , 3.90 ( ddd , j = 10.9 , 4.0 , 1.1 hz , 1.2 1h , anti ) , 2.88 ( dd , j = 13.5 , 7.2 hz , 1h , syn ) , 2.67 ( dd , j = 13.6 , 6.0 hz , 1.2 1h , anti ) , 2.57 ( dd , j = 13.5 , 8.3 hz , 1h , syn ) , 2.42 ( dd , j = 13.6 , 9.8 hz , 1.2 1h , anti ) , 2.232.15 ( m , 1h , syn ) , 2.142.08 ( m , 1.2 1h , anti ) , 1.37 ( s , 1.2 9h , anti ) , 1.33 ( s , 9h , syn ) , 1.29 ( s , 1.2 9h , anti ) , 1.25 ( s , 9h , syn ) ; c nmr ( 101 mhz , cd3od ) 150.6 ( 2c ) , 141.8 , 141.5 , 139.8 , 139.6 , 136.3 , 136.1 , 128.8 ( 2c ) , 128.0 ( 2c ) , 125.8 , 125.7 , 125.2 , 124.7 , 124.0 , 123.1 , 123.0 , 122.4 , 67.7 , 65.5 , 63.9 , 63.0 , 41.7 , 40.6 , 34.5 , 34.5 , 34.3 , 33.7 , 33.7 , 32.5 , 30.7 ( 2c ) , 28.9 ( 2c ) ; ft - ir max(atr ) cm 2956 , 2870 , 1479 , 1450 , 1361 , 1229 , 1171 , 1130 , 1028 , 882 , 749 , 701 , 532 ; m / z ( ei ) 352 ( [ m ] , 65 ) , 91 ( [ phch2 ] , 15 ) ; hrms ( ei ) calcd for [ c24h32o2 ] 352.23968 , found 352.23972 . the title compound was synthesized according to the general procedure employing 1i ( 0.12 mmol , 33.8 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 73% ( 25 mg ) ; anti / syn 1:1 ; h nmr ( 600 mhz , cdcl3 ) 7.367.14 ( m , 10h ) , 6.956.77 ( m , 6h ) , 4.484.47 ( m , 2h ) , 4.29 ( dd , j = 11.1 , 2.7 hz , 1h ) , 4.22 ( ddd , j = 10.7 , 3.7 , 1.2 hz , 1h ) , 4.154.04 ( m , 6h ) , 2.88 ( dd , j = 13.7 , 8.4 hz , 1h ) , 2.70 ( dd , j = 14.0 , 7.0 hz , 1h ) , 2.68 ( dd , j = 13.7 , 7.2 hz , 1h ) , 2.59 ( dd , j = 13.9 , 8.5 hz , 1h ) , 2.352.29 ( m , 1h ) , 2.22 ( dqd , j = 12.9 , 4.3 , 2.8 hz , 1h ) , 1.97 ( bs , 2h ) , 1.471.45 ( m , 6h ) ; c nmr ( 151 mhz , cdcl3 ) 147.6 , 147.5 , 144.1 , 144.0 , 139.2 , 139.1 , 129.1 ( 2c ) , 128.5 ( 2c ) , 126.3 ( 2c ) , 125.0 , 124.0 , 121.8 , 121.6 , 120.4 , 120.1 , 112.6 ( 2c ) , 67.4 , 65.4 , 65.2 , 64.7 , 64.4 , 64.3 , 41.3 , 39.8 , 34.6 , 32.9 , 14.8 ( 2c ) ; ft - ir max(atr ) cm 3506 , 2921 , 1585 , 1478 , 1391 , 1315 , 1255 , 1214 , 1080 , 1030 , 964 , 900 , 737 ; m / z ( ei ) 284 ( [ m ] , 23 ) , 91 ( [ phch2 ] , 100 ) ; hrms ( ei ) calcd for [ c18h20o3 ] 284.14070 , found 284.14077 . the title compound was synthesized according to the general procedure employing 1j ( 0.12 mmol , 32.2 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 65% ( 21 mg ) ; mp = 112113 c ; anti / syn 1:1 ; h nmr ( 400 mhz , cd3od ) 7.307.13 ( m , 10h ) , 6.88 ( d , j = 3.0 hz , 1h ) , 6.806.63 ( m , 5h ) , 4.46 ( d , j = 3.5 hz , 1h ) , 4.36 ( d , j = 4.6 hz , 1h ) , 4.09 ( dd , j = 11.0 , 2.7 hz , 1h ) , 3.993.89 ( m , 2h ) , 3.84 ( ddd , j = 11.0 , 5.1 , 0.7 hz , 1h ) , 3.73 ( s , 3h ) , 3.70 ( s , 3h ) , 2.87 ( dd , j = 13.6 , 7.2 hz , 1h ) , 2.74 ( dd , j = 13.7 , 6.2 hz , 1h ) , 2.56 ( dd , j = 13.6 , 8.3 hz , 1h ) , 2.47 ( dd , j = 13.7 , 9.3 hz , 1h ) , 2.232.15 ( m , 1h ) , 2.142.07 ( m , 1h ) ; c nmr ( 101 mhz , cd3od ) 153.7 , 153.4 , 148.2 , 148.1 , 139.7 , 139.5 , 128.8 , 128.7 , 128.0 ( 2c ) , 125.8 , 125.7 , 124.9 , 123.9 , 116.7 ( 2c ) , 115.5 ( 2c ) , 113.9 , 113.8 , 67.1 , 64.8 , 64.5 ( 2c ) , 54.7 ( 2c ) , 42.0 , 40.7 , 34.3 , 32.3 ; ft - ir max(atr ) cm 2923 , 2471 , 1490 , 1430 , 1265 , 1205 , 1161 , 1037 , 807 , 703 ; m / z ( ei ) 270 ( [ m ] , 71 ) , 91 ( [ phch2 ] , 33 ) ; hrms ( ei ) calcd for [ c17h18o3 ] 270.12505 , found 270.12551 . the title compound was synthesized according to the general procedure employing 1k ( 0.12 mmol , 32.2 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 92% ( 29.8 mg ) ; mp = 100103 c ; anti / syn 1.7:1 ; h nmr ( 400 mhz , cd3od ) 7.427.21 ( m , 7.2h ) , 7.207.12 ( m , 8h ) , 7.09 ( d , j = 8.5 hz , 1h ) , 6.51 ( dd , j = 8.5 , 2.5 hz , 1.7 1h , anti ) , 6.44 ( dd , j = 8.5 , 2.5 hz , 1h , syn ) , 6.35 ( d , j = 2.5 hz , 1.7 1h , anti ) , 6.30 ( d , j = 2.5 hz , 1h , syn ) , 4.41 ( d , j = 3.1 hz , 1h , syn ) , 4.34 ( d , j = 3.9 hz , 1.7 1h , anti ) , 4.15 ( dd , j = 11.0 , 2.6 hz , 1.7 1h , anti ) , 4.113.93 ( m , 2h , syn ) , 3.89 ( ddd , j = 11.0 , 4.2 , 0.9 hz , 1.7 1h , anti ) , 3.72 ( s , 3h , anti ) , 3.69 ( s , 3h , syn ) , 2.85 ( dd , j = 13.6 , 7.4 hz , 1h , syn ) , 2.66 ( dd , j = 13.7 , 6.6 hz , 1.7 1h , anti ) , 2.57 ( dd , j = 13.6 , 8.0 hz , 1h , syn ) , 2.46 ( dd , j = 13.7 , 9.1 hz , 1.7 1h , anti ) , 2.232.14 ( m , 1h , syn ) , 2.142.03 ( m , 1.7 1h , anti ) ; c nmr ( 101 mhz , cd3od ) 160.7 ( 2c ) , 155.2 ( 2c ) , 139.6 , 139.5 , 131.1 , 130.9 , 128.8 , 128.7 , 128.0 ( 2c ) , 125.8 , 125.7 , 117.1 , 115.6 , 107.3 , 106.8 , 100.5 ( 2c ) , 66.3 , 64.5 , 64.2 , 64.1 , 54.3 , 54.2 , 42.0 , 40.7 , 34.2 , 32.6 ; ft - ir max(atr ) cm : 3215 , 2960 , 1737 , 1667 , 1494 , 1366 , 1221 , 1092 , 1032 , 734 ; m / z ( ei ) 270 ( [ m ] , 15 ) , 91 ( [ phch2 ] , 89 ) ; hrms ( ei ) calcd for [ c17h18o3 ] 270.12505 , found 270.12534 . the title compound was synthesized according to the general procedure employing 1l ( 0.12 mmol , 32.2 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 55% ( 17.7 mg ) ; mp = 121126 c ; anti / syn 1.2:1 ; h nmr ( 600 mhz , cdcl3 ) 7.32 ( dd , j = 7.6 , 1.4 hz , 1.2 1h , anti ) , 7.257.17 ( m , 5.4h ) , 7.117.08 ( m , 1.2 1h , anti ) , 6.95 ( td , j = 7.5 , 0.9 hz , 1.2 1h , anti ) , 6.916.82 ( m , 7.4h ) , 4.50 ( t , j = 3.5 hz , 1h , syn ) , 4.48 ( t , j = 4.1 hz , 1.2 1h , anti ) , 4.21 ( dd , j = 9.2 , 4.6 hz , 1.2 1h , anti ) 4.114.06 ( m , 2h , syn ) , 3.97 ( dd , j = 11.0 , 4.0 hz , 1.2 1h , anti ) , 3.80 ( s , 3h , syn ) , 3.79 ( s , 1.2 3h , anti ) , 2.82 ( dd , j = 13.8 , 8.5 hz , 1h , syn ) , 2.682.58 ( m , 2.2h ) , 2.48 ( dd , j = 14.0 , 9.3 hz , 1.2 1h , anti ) , 2.322.24 ( m , 1h , syn ) , 2.212.14 ( m , 1.2 1h , anti ) , 1.91 ( bs , 1.2 1h , anti ) , 1.72 ( bs , 1h , syn ) ; c nmr ( 151 mhz , cdcl3 ) 158.1 ( 2c ) , 154.3 , 154.2 , 131.1 , 131.0 , 130.1 ( 2c ) , 130.0 ( 2c ) , 129.9 , 129.7 , 124.2 , 123.2 , 120.9 , 120.5 , 116.9 ( 2c ) , 114.0 , 113.9 , 67.6 , 65.0 , 64.9 , 64.6 , 55.3 ( 2c ) , 41.6 , 40.1 , 33.7 , 31.9 ; ft - ir max(atr ) cm : 3392 , 2928 , 1609 , 1582 , 1510 , 1486 , 1450 , 1298 , 1246 , 1176 , 1031 , 850 , 813 , 753 ; m / z ( ei ) 270 ( [ m ] , 27 ) , 121 ( [ 4-meophch2 ] , 100 ) ; hrms ( ei ) calcd for [ c17h18o3 ] 270.12505 , found 270.12548 . the title compound was synthesized according to the general procedure employing 1 m ( 0.12 mmol , 37.7 mg ) , 1 mol % ( 1.1 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 86% ( 32.7 mg ) ; anti / syn 1.8:1 ; h nmr ( 600 mhz , cdcl3 ) 7.43 ( d , j = 7.2 hz , 2h ) , 7.39 ( d , j = 7.2 hz , 2h ) , 7.357.18 ( m , 26.7h ) , 7.127.09 ( m , 3h ) , 6.976.85 ( m , 5.5h ) , 4.44 ( bs , 1h , syn ) , 4.41 ( bs , 1.8 1h , anti ) , 4.27 ( dd , j = 11.2 , 2.3 hz , 1.8 1h , anti ) , 4.144.06 ( m , 2h , syn ) , 4.00 ( ddd , j = 10.8 , 3.5 , 1.5 hz , 1h , syn ) , 3.963.93 ( m , 1.8 1h , anti ) , 3.70 ( d , j = 12.4 hz , 1.8 1h , anti ) , 2.91 ( ddd , j = 11.9 , 7.5 , 3.2 hz , 1h , syn ) , 2.79 ( dq , j = 12.4 , 2.3 hz , 1.8 1h , anti ) , 2.00 ( bs , 1.8 1h , anti ) , 1.80 ( bs , 1h , syn ) ; c nmr ( 151 mhz , cdcl3 ) 154.4 ( 2c ) , 142.6 , 142.4 , 142.3 ( 2c ) , 130.9 , 130.5 , 130.0 , 129.9 , 128.9 ( 2c ) , 128.8 ( 2c ) , 128.1 ( 2c ) , 127.9 , 127.8 , 126.8 , 126.7 , 126.6 , 126.6 , 124.0 , 122.5 , 121.0 , 120.5 , 117.1 , 116.9 , 65.8 , 64.1 , 64.0 , 63.1 , 49.6 , 49.2 , 43.0 , 41.9 ; ft - ir max(atr ) cm : 3036 , 2927 , 1702 , 1619 , 1516 , 1491 , 1446 , 1318 , 1226 , 1117 , 1014 , 972 , 755 ; m / z ( ei ) 167 ( [ chph2 ] , 59 ) , 151 ( [ m cph2 ] , 28 ) ; hrms ( ei ) calcd for [ c22h20o2 ] 316.14578 , found 316.14636 . the title compound was synthesized according to the general procedure employing 1n ( 0.12 mmol , 26.7 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 68% ( 14.6 mg ) ; anti / syn 2.5:1 ; h nmr ( 600 mhz , cdcl3 ) 7.417.35 ( m , 4h ) , 7.357.29 ( m , 10h ) , 7.287.21 ( m , 14.5h ) , 7.197.15 ( m , 3h ) , 5.00 ( d , j = 5.4 hz , 1h , syn ) , 4.95 ( d , j = 6.6 hz , 2.5 1h , anti ) , 3.143.07 ( m , 4.5h ) , 3.01 ( dd , j = 15.3 , 7.3 hz , 3h ) , 2.872.83 ( m , 2.5h ) , 2.812.75 ( m , 4h ) , 2.68 ( dq , j = 14.0 , 7.9 hz , 1h , syn ) , 2.622.48 ( m , 6h ) , 1.62 ( bs , 3.5h ) ; c nmr ( 151 mhz , cdcl3 ) 144.7 , 144.5 , 143.5 , 141.5 , 141.2 , 140.6 , 128.9 ( 2c ) , 128.7 , 128.5 , 128.4 , 128.2 , 126.8 ( 2c ) , 126.2 , 125.9 , 125.0 , 124.8 , 124.7 , 123.9 , 80.8 , 76.3 , 52.3 , 46.9 , 39.3 , 35.9 , 35.8 , 35.0 ; ft - ir max(atr ) cm 3363 , 2932 , 1487 , 1454 , 1257 , 1102 , 1026 , 804 , 745 , 697 ; m / z ( ei ) 223 ( [ m 1 ] , 21 ) , 132 ( m [ phch2 ] , 100 ) 91 ( [ phch2 ] , 80 ) ; hrms ( ei ) calcd for [ c16h16o ] 224.11957 , found 224.11989 . the title compound was synthesized according to the general procedure employing 1o ( 0.12 mmol , 30 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 59% ( 17.7 mg ) ; e / z 1:1 ; h nmr ( 400 mhz , cdcl3 ) 7.41 ( dd , j = 7.7 , 1.6 hz , 1h ) , 7.317.18 ( m , 6h ) , 7.167.07 ( m , 2h ) , 7.067.00 ( m , 2h ) , 7.036.8 ( m , 6h ) , 6.77 ( s , 1h ) , 5.32 ( s , 1h ) , 5.19 ( s , 1h ) , 4.974.86 ( m , 3h ) , 4.57 ( dd , j = 12.0 , 0.9 hz , 1h ) , 2.37 ( s , 3h ) , 2.35 ( s , 3h ) , 2.14 ( bs , 1h ) , 2.05 ( bs , 1h ) ; c nmr ( 101 mhz , cdcl3 ) 154.4 , 154.3 , 138.1 ( 2c ) , 135.5 , 135.4 , 134.4 , 133.4 , 131.6 , 130.3 , 130.0 , 129.8 , 129.5 ( 2c ) , 129.2 , 129.0 , 128.7 , 128.4 , 128.3 ( 2c ) , 125.9 , 125.8 , 124.9 , 124.0 , 121.1 ( 2c ) , 117.2 , 116.9 , 69.3 , 68.1 , 63.0 , 62.8 , 21.4 ( 2c ) ; ft - ir max(atr ) cm 3391 , 2922 , 1737 , 1590 , 1479 , 1370 , 1224 , 993 , 921 , 756 , 702 ; m / z ( ei ) 252 ( [ m ] , 17 ) , 104 ( [ 3-meph ] , 25 ) , 91 ( [ 3-mephch ] , 32 ) ; hrms calcd for [ c17h16o2 ] 252.11448 , found 252.11402 . the title compound was synthesized according to the general procedure employing 1p ( 0.12 mmol , 32 mg ) , 2.5 mol % ( 2.74 mg ) of pc 3 , and 2.5 equiv of dipea ( 38.8 mg ) . the product was purified by flash column chromatography : yield 57% ( 18.3 mg ) , e / z 1:1 ; h nmr ( 600 mhz , cdcl3 ) 7.467.43 ( m , 2h ) , 7.427.39 ( m , 1h ) , 7.307.28 ( m , 1h ) , 7.247.20 ( m , 2h ) , 7.207.16 ( m , 2h ) , 6.996.84 ( m , 9h ) , 6.76 ( s , 1h ) , 5.33 ( s , 1h ) , 5.18 ( s , 1h ) , 4.994.89 ( m , 2h ) , 4.87 ( dd , j = 12.0 , 1.2 hz , 1h ) , 4.594.55 ( m , 1h ) , 3.83 ( s , 3h ) , 3.82 ( s , 3h ) , 2.20 ( bs , 1h ) , 2.07 ( bs , 1h ) ; c nmr ( 151 mhz , cdcl3 ) 159.3 , 159.1 , 154.4 , 154.3 , 133.3 , 133.2 , 131.9 , 131.3 , 130.3 , 130.2 , 130.0 , 129.8 , 129.1 , 128.8 , 128.1 , 128.0 , 125.0 , 124.1 , 121.1 ( 2c ) , 117.3 , 116.9 , 113.9 ( 2c ) , 69.4 , 68.2 , 63.0 , 62.8 , 55.3 ( 2c ) ; ft - ir max(atr ) cm 3393 , 2958 , 2929 , 1606 , 1509 , 1485 , 1458 , 1246 , 1177 , 1031 , 995 , 828 , 755 ; m / z ( ei ) 268 ( [ m ] , 25 ) , 120 ( [ 3-meophch ] , 100 ) . Output:	a visible light photoredox - catalyzed aldehyde olefin cyclization is reported . the method represents a formal hydroacylation of alkenes and alkynes and provides chromanol derivatives in good yields . the protocol takes advantage of the double role played by trialkylamines ( nr3 ) which act as ( i ) electron donors for reducing the catalyst and ( ii ) proton donors to activate the substrate via a proton - coupled electron transfer .
PubmedSumm2087	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: various guidelines have been suggested for appropriate evaluation of metastatic cancers when the primary tumor site is not readily identifiable based on the clinical presentation , tissue histology and immunohistochemical staining . after such evaluation , cancer of unknown primary ( cup ) is the diagnosis made in about 45% of invasive cancers . series of patients with this diagnosis have been studied and treated with empiric combination chemotherapy regimens . both the specific recommended regimens and the efficacy of these therapies vary by the histologic subtype and immunohistochemical and clinical presentation . objective response rates in the range of 3050% have been described in many published series . for example , in a series of 71 patients treated with paclitaxel , carboplatin and oral etoposide , 46% of patients had a major objective response with a median survival of 11 months . the national comprehensive cancer network ( nccn ) has published guidelines with recommendations concerning evaluation and care for patients with cup . molecular profiling is described as an emerging diagnostic tool , with the potential of identifying the likely primary based on comparison of expression patterns in the unknown tumor relative to known data banks that include known primaries [ 510 ] . confirming the specificity of the testing in a particular patient remains challenging , if not impossible , short of autopsy series ; however , with improved chemotherapy and targeted therapies now available for several solid tumors , a more site - specific therapy is likely to be superior for cup patients diagnosed with a specific tissue of origin compared to the administration of an empiric broad - spectrum chemotherapy regimen . ultimately , the recognition of specific molecular abnormalities ( mutations ) in tumor cells which can be targeted with specific drugs is likely to be far more clinically relevant than even identifying the primary tissue of origin in terms of choosing therapies . expression profiling strongly suggested the tissue of origin to be the kidney and the patient 's tumor responded well and durably to a targeted agent which is fda approved for treating metastatic kidney cancer . pg is a 53-year - old male who presented in april 2011 with right hip pain . ct , pet / ct and mri scanning confirmed diffuse lymphadenopathy with the pet scan showing markedly increased fdg uptake involving the left neck and supraclavicular regions ( left supraclavicular lymph node measuring 4.1 2.6 cm ) , mediastinal , retroperitoneal , bilateral and common external and internal iliac lymph node chains . the mri report noted extensive retroperitoneal and retrocrural adenopathy as well as a large necrotic mass apparently metastatic nodes. left supraclavicular lymph node biopsy showed a poorly differentiated adenocarcinoma. immunohistochemical staining was done including negative staining for cytokeratin 20 , ttf-1 , calretinin , ca19 - 9 , wt-1 and d2 - 40 , and strongly positive staining for cytokeratin 7 . the phenotype was felt to be consistent with a pancreatobiliary , lung or upper gi primary site . the pathologists wrote the morphology does not suggest a renal or adrenal primary , a question that had been raised based on the scans above . cea , hcg and afp were normal , while ca19 - 9 was 64.9 units / ml ( normal range 035 ) and ldh was 214 iu / l ( 98192 ) . the tumor was sent for mrna expression profiling ( biotheranostics , san diego , calif . , usa ) and the report suggested a 95% probability of this cancer representing kidney cancer with a 94% probability of the tumor being a papillary renal cell carcinoma ( rcc ) . because of the biotheranostics expression profiling result , further immunostaining was done . the tumor stained strongly positive for racemace ( amacr ) , rcc antigen and cd10 . the immunohistochemical profile strongly supports the tumor to be renal in origin and specifically papillary renal carcinoma. the patient was treated with everolimus ( 10 mg po qd ) beginning on may 10 , 2011 , and , 2 weeks later , reported a diminution in the supraclavicular fullness . on june 23 , 2011 , a pelvic mri was repeated and the report noted a mild decrease in the size of the retroperitoneal adenopathy . on july 5 , 2011 , a pet / ct was interpreted as showing relatively decreased fdg uptake in multiple lymph node regions suggestive of some interval treatment response since the prior pet / ct . in march 2012 , scans continued to show no evidence of progression of his malignancy . with improved chemotherapy and targeted therapies now available for several solid tumors , a more site - specific therapy is likely to be superior for cup patients diagnosed with a specific tissue of origin compared to the administration of an empiric broad - spectrum chemotherapy regimen . in the case described above , kidney cancer was identified as the likely cancer type through expression profiling , and the patient was successfully treated with a targeted agent indicated for treatment of that cancer type . treating some cup patients with systemic therapies described for known counterpart primaries based on gene expression profiling has resulted in more favorable outcomes compared to those outcomes reported for historic controls of cup patients treated with the less specific regimens , where no expression profiling was completed . for example , greco et al . recently noted a 64% rr and a 22-month median survival for 32 cup patients identified by expression profiling as having probable colorectal primaries ( although colonoscopies were negative in 30 of the 32 patients ) , while historic controls with cup treated with cup regimens , rather than colorectal regimens , had a 9-month median survival . still , no prospective randomized studies have been done and , in spite of small series such as the one above , it may be that while a molecular profile can be consistent with that of a particular primary , the response to available therapies may be different for these tumors , since the biological behavior of cup may be different from their more readily identifiable counterparts . the nccn panel concluded the panel does not recommend molecular profiling as part of routine evaluation and suggests no specific circumstances in which such testing should be considered . after reviewing the literature , this case appears to represent the first case of a papillary renal carcinoma where expression profiling was used to identify the primary , more specific immunostaining was used to confirm the diagnosis , and the patient was treated with a targeted agent , which resulted in a response . everolimus is an fda approved targeted agent for the treatment of advanced renal carcinoma based on a study showing that , compared to placebo , everolimus more than doubled the time without tumor growth or death . such therapy clearly would not have been a consideration if not for the pursuit of the molecular profiling . our patient 's tumor initially responded to everolimus and has shown no progression for over 10 months . while choosing which specific patients will benefit from expression profiling remains problematic , cases such as this highlight the importance of considering such testing in cup patients . Output:	cancer of unknown primary ( cup ) is a clinical syndrome representing many types of cancers and diagnoses are typically made after review of clinical presentation , pathology ( including immunohistochemical staining ) and imaging studies . treatment with systemic chemotherapy has been shown to result in fairly reproducible objective response rates . herein , a case of a patient who was initially diagnosed with a poorly differentiated adenocarcinoma of unknown origin is reported . after mrna gene expression profiling ( commercially available cancertype i d ) , a specific diagnosis of papillary renal cell carcinoma ( rcc ) was made and then confirmed with additional immunohistochemical staining . the patient was treated with targeted therapy and an objective radiographic response was seen . a literature review suggests this to be the first patient with papillary rcc , identified by molecular profiling , and benefitting from a targeted agent that otherwise would not have been considered in the setting of cup . this case underscores the importance of considering the use of newer testing technologies in the interest of offering patients more specific , targeted therapy in order to improve efficacy and spare patients toxicities of less specific , empiric chemotherapeutic regimens .
PubmedSumm2088	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: treatment options for hepatitis c virus ( hcv ) infection in renal transplant ( rtx ) patients are limited . the conventional treatment of interferon ( ifn ) injection with ribavirin carries the risk of allograft rejection . conversely , not treating for hcv is associated with increased morbidity and mortality due to progressive liver dysfunction , infections , malignancy , vasculitis , glomerulonephritis , and new onset diabetes . the availability of second generation oral anti - viral drugs has opened new avenues for hcv treatment . although used successfully in liver transplant situation , there is no available literature for their use in rtx patient . we share our early experience of a rtx patient with hcv infection treated successfully with combination pill of sofosbuvir ( sof ) and ledipasvir ( ldv ) ( harvoni ) , heralding a new era of safe , ifn - free , oral , anti - hcv therapy a 43-year - old male renal allograft recipient , presented at the sixth year follow up with significant weight loss of 6 kg over 3 months . prior to renal transplantation he was hypertensive , nondiabetic and hcv negative ( confirmed by standard elisa and polymerase chain reaction [ pcr ] test ) . induction therapy included basiliximab 2 doses ( 20 mg each ) followed by triple drug immuno - suppression ( prednisolone , tacrolimus , mycophenolate mofetil [ mmf ] ) he received two units blood transfusion in the peritransplant period . he was receiving prednisolone 5 mg daily , mmf500 mg thrice daily , and tacrolimus 1 mg twice daily . his other medications included nifedipine 20 mg thrice daily , metoprolol 50 mg once daily and calcium carbonate 500 mg twice daily . laboratory evaluation showed de novo new onset diabetes mellitus after transplant ( fasting blood glucose 225 mg / dl , hemoglobin a1c 11.9 ) , elevated liver enzymes ( aspartate transaminase 89 iu / l [ normal < 40 ] , gamma glutamyl transferase 150 iu / l , biliburin 1 mg% , serum albumin 4 g / dl ) , normal renal functions ( serum creatinine 0.9 mg% ) , normal hemogram , and chest x - ray . furthermore , the evaluation showed hbsag negative , anti - hcv serology positive status due to hcv genotype 1 with high viral load ( 10,800,000 iu / ml ) . diabetes was initially controlled with insulin and later he was switched to oral sitagliptin - metformin combination . his serum transaminases level remained elevated over next 9 months with persistent high hcv load ( 106,074,000 iu / ml ) . the patient was counseled about the risk of allograft rejection with existing anti - hcv therapy , and lack of treatment guidelines for using oral new anti - viral agents in rtx situation . however , with recent treatment success with newer oral anti - viral agents in non - rtx situation , the patient consented to take oral anti - viral therapy under closed supervision . he received the fixed - dose combination of ldv and sof ( harvoni ) , manufactured by gilead sciences . for next 12 weeks . at 3 weeks ( after starting treatment ) doppler ultrasound and radionuclide study with technetium-99m - diethylene - triamine - pentaacetate were normal . his serum cyclosporine trough level was in the acceptable range ( 110.2 ng / ml whole blood at 3 weeks while baseline was 95 ng / ml by liquid chromatography - tandem mass spectrometry normal range 80150 on long - term ) . he continues to do clinically well at four months posttreatment in addition , after completion of oral anti - viral therapy he is off anti - diabetic treatment with maintained euglycemia and normal liver functions . liver disease is an important cause of morbidity and mortality in rtx recipients , especially with hcv infection . abnormal liver functions test are reported in 724% of rtx recipients , with liver failure as the cause of death in 828% of long - term survivors . liver disease in rtx patient may be due to viral infections ( hepatitis b virus or hcv , epstein - barr virus , or cytomegalovirus ) , drugs ( azathioprine or cyclosporine ) , iron overload ( hemosiderosis ) or alcohol abuse . hcv infection in immuno - suppressed rtx recipient is associated with significant morbidity and mortality . patients with hcv - positive status prior to transplantation have increased risk of posttransplant liver disease with a relative risk ( rr ) of 5.0 . one large meta - analysis , that included 6365 hcv positive patients reported increased risk for death ( rr 1.79 ) and allograft failure ( rr 1.56 ) . hcv infection additionally causes heightened immuno - suppression thereby increasing the risk of sepsis in the initial years post - rtx and liver disease in later years that progresses more rapidly compared to the nontransplant situation to chronic hepatitis , cirrhosis , and liver cancer . hcv infection is also associated with increased risk for new onset diabetes after transplantation ( nodat ) , de novo glomerulonephritis and mixed cryoglobulinemia - related vasculitis that may lead to graft loss or recipient death . interestingly , the present case highlights that the patient who developed nodat associated with hcv infection may have a collateral benefit of getting cured of diabetes with control of hcv infection . efforts to minimize the incidence of post - rtx hcv infection must focus on viral screening of both donors and recipients using sensitive methods in the pretransplant stage . third generation elisa serology for the detection of hcv has high sensitivity ( > 98% ) . the latter is confirmed by a gold standard pcr test that detects hcv rna antigen , which should be done in all patients prior to transplant . the index patient was negative for hcv by elisa and pcr in pre - rtx evaluation . he probably acquired hcv infection from blood transfusions ( tested negative by elisa ) received in peritransplant period though a remote possibility of missed hcv infection in pretransplant evaluation exists . the latter may occur in patients who have intermittent viremia from the sequestered virus in the liver or peripheral mononuclear cells leading to negative serum hcv status . the standard care for the patients with hcv genotype 1 infection in non - rtx scenario is pegylated ifn plus ribavirin with newer direct acting anti - viral protease inhibitor ( boceprevir or telaprevir ) for 48 weeks . the overall sustained virological response ( svr ) rate in patients infected with hcv genotype 1 is poor ( < 50% ) compared to genotype 2 or 3 ( approximately 7580% ) more so in patients on dialysis ( nearly one - third ) . current treatment option approved for patients ineligible to receive inf in hcv genotype 1 infection is 24 weeks of sof and ribavirin . rtx is a contraindication to ifn therapy due to its limited efficacy , increased risk of acute rejection , high cost , and significant side effects . however , ifn mono - therapy is to be considered in kidney transplant recipients with hcv infection having fibrosing cholestatic hepatitis or life - threatening vasculitis . ribavirin in the post - rtx state does not produce sustained virologic response despite lowering of transaminase level . current clinical strategy in the post - rtx hcv infection is manipulation of immunosuppressive therapy ( minimizing corticosteroid , avoidance of anti - lymphocyte antibody , and to replace tacrolimus with cyclosporine ) . however , there is an urgent need for newer anti - hcv therapies with greater safety and efficacy than ifn and ribavirin in rtx patient . the drugs used as add - on oral therapy in hcv treatment include ribavarine , telaprevir , boceprevir , or simeprevir in different combinations . the advent of newer second generation anti - virals like sof -ldv , ombitasvir - paritaprevir plus dasabuvir , daclatasvir - asunaprevir , and simeprevir - sof has widened the therapeutic options with reported svr above 90% . the novel oral anti - viral agents are likely to cause a paradigm shift in the treatment of hcv infection in post - rtx state by simplifying current therapy of shorter duration and also eliminating the need for response guided therapeutic approach . the choice between these agents depends primarily on the potential for drug interactions and drug toxicity . the combination therapies are preferred as they provide better viral suppression than mono - therapy with less chance of viral escape based on complementary resistance profiles . there is no additional benefit by extension of the duration of treatment to even 24 weeks . addition of ribavirin to a ldv - sof combination increases toxicity without any additional efficacy . ldv - sof combination has favorable adverse effect profile ( common side effects rated as mild and not leading to drug discontinuation like fatigue , headache , nausea , diarrhea , and insomnia reported in 1020% ) , minimal drug interactions , and easy to administer ( single pill daily ) . ldv is an inhibitor of the hcv ns5a protein that is required for viral replication , postreplication assembly , and secretion . sof , a nucleotide pro - drug and an inhibitor of the hcv ns5b rna - dependent rna polymerase , acts as a chain terminator . sofosobuvir , the backbone of the new standard of care for hcv infection , offers a number of potential benefits such as high rates of sustained virologic response , high barrier to resistance , tolerability , pan - genotype efficacy ( hcv genotype 16 ) , need for shortened duration of treatment , usefulness in cirrhosis , human immunodeficiency virus co - infection and liver transplantation . sof ( 400 mg ) in combination with ldv ( 90 mg ) ( harvoni ) has been proven to be efficacious , tolerable and safe in series of clinical trials , both in treatment - nave and treatment - experienced patients with 1224 weeks of therapy ( depending presence or absence of cirrhosis , treatment experience , and genotype ) with high sustained viral response . the index rtx patient with a very high viral load without cirrhosis showed a remarkable response as early as 3 weeks after therapy . shorter duration for 8 weeks can be considered in treatment - nave patients without cirrhosis who have pretreatment hcv rna of < 6 million iu / ml . both sof and ldv are substrates of p - glycoprotein inducers ( like rifampicin ) with implication for potential drug interactions but it does not demonstrate significant drug interactions with immunosuppressant drugs . the development of ifn free oral anti - hcv therapy may be a major breakthrough in anti - hcv management in rtx patient . the dual drug combination therapy appears to be an important arrow in our quiver now for targeting hcv infection in high - risk rtx . such combination therapy may shorten and simplify treatment regimens even in the post - rtx scenario ushering a new era of safer , simpler and more effective therapy . however , the utility versus the risk of such new therapy has to be scrutinized carefully by studies involving a sizeable cohort of rtx patients having long - term follow - up for reliable assessment of such therapeutic intervention . medwin hospital , nampally , hyderabad , andhra pradesh , india . Output:	liver disease due to hepatitis c infection in renal transplant recipients is difficult to treat and often associated with reduced patient survival . a 43-year - old male , a renal allograft recipient , presented at 6 years follow - up with significant weight loss over 3 months . he was detected to have new onset diabetes mellitus together with hepatitis c virus ( hcv ) infection ( genotype 1 ) . his hcv load remained high despite the change of immuno - suppression from tacrolimus to cyclosporine . a decision to treat with a new anti - viral combination of ledipasvir and sofosbuvir for 12 weeks was taken . within 3 weeks , his raised serum transaminases levels normalized and viral load became undetectable . at the end of 16 weeks , he continues to do well with normal renal function , has sustained remission from hepatitis c infection and resolution of diabetes .
PubmedSumm2089	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: pancreatic endocrine tumors are classified , according to the symptomatology , in secreting and nonsecreting tumors . the nonsecreting tumors ( silent , in hormonal terms ) represent about 50% of the total ; they are followed by insulinomas - 25% and gastrinomas - 15% . it may be associated with other endocrine glands tumors in the multiple endocrine neoplasia type i ( men i ) ( parathyroid , pituitary , endocrine pancreas ) . the islet cell tumors are the most common cause of hypoglycemia resulting from endogenous hyperinsulinism . the median age at diagnosis is about 47 years , except in insulinoma patients with men 1 , in whom the median age is the mid-20 s . a 61-years old man known with hypertension and alcoholic steatohepatitis , presented on 26/03/2013 for syncopes , tonic - clonic seizures , sweating , anxiety . alcohol consumption had been stopped 2 months before the onset of the symptoms , according to the patient and his relatives statements . in the patient medical history we found an emergency admission on 20/02/2013 for psychomotor agitation , episodes of loss of consciousness with tonic - clonic seizures , delirium of mixed etiology ( alcohol and hypoglycemia ) , tremor of the upper limbs , sweating , disorientation in time and space , important hypoprosexia , fixing and evocation hypoamnesia , insomnia . blood tests evidenced 41 mg / dl hypoglicemia , hyperkaliemia , twice the normal values of lactate dehydrogenase ( ldh ) . the cranial computed tomography ( ct ) did not reveal any damage except an old ischemic millimetric lesion and a degree of atrophy . the patient was admitted to the diabetes , nutritional and metabolic diseases clinic for suspected insulinoma . the abdominal ct revealed no pathological changes in the abdominal organs , no lymph node changes . due to his agitation , seizures , and delirium , the symptoms were interpreted as alcohol withdrawal and he was transferred to the department of psychiatry . during hospitalization the blood glucose fasting levels were found again between 30 and 50 mg / dl . the neurologist interpreted the old ischemic lesion shown on the ct as possibly due to hypoxic damage caused by severe hypoglycemia . the patient was discharged with the diagnosis of complicated withdrawal seizures , chronic alcohol dependence , history of disabling ischemic stroke in the left carotid territory , with recommendation for hospitalization in a medical clinic . the following were determined : the c peptide , insulinemia and glycated hemoglobin , which were normal and did not support the diagnosis of insulinoma . , hypoglycemia was attributed to the alcohol consumption and the patient was released on 03/04/2013 . after a month , on 26/03/2013 , the patient came back for persistent symptoms . blood tests showed 39 to 59 mg / dl glucose levels and mixed dyslipidemia , carbohydrate antigen ( ca)19 - 9 was normal , the other analyses were normal . we decided to repeat the abdominal magnetic resonance imaging ( mri ) with contrast , and we discovered a 21 mm hypervascular tumor of the pancreatic uncinate process , suggesting , as first hypothesis , an endocrine tumor and peripancreatic millimetre lymph nodes . we transferred the patient to the surgery department , where the uncinate process tumor was excised and peripancreatic lymph nodes were biopsied . the histopathological appearance ( fig . 3 ) was positive in 56% of tumor cells ( nuclear expression ) , findings that were supporting the diagnosis of neuroendocrine tumor . based on the result of the histopathological examinations , corroborated with the imaging and intraoperative exploration , the tumor stage was as pt3n0mx . the rule of 10 states that 10% are multiple , 10% are malignant , insulinomas are associated with men 1 in 5% of patients and it is estimated that 21% of patients with men1 develop insulinomas . the incidence is 310 cases per million people per year . once surgically removed , the insulinomas occurring in this autosomal dominant syndrome have a higher risk of relapse . long - term survival of patients with insulinoma is generally excellent ; it is considered that approximately 9095% of insulinomas have benign histological behavior , so healing with dissaperance of symptomatology after complete resection is the rule . recurrences were observed in 5.4% of cases in a series of 120 patients with benign insulinomas , over a period of 417 years . in these cases long - term survival is possible in malignant insulinomas , especially in cases strictly localized to the pancreas and with good response to adjuvant chemotherapy ; median disease - free survival after curative resection is estimated at 5 years . the diagnosis of insulinoma often comes from a hunch , from the impossibility of highlighting the cause of repeated hypoglycemia in an apparently healthy patient . but for diagnostic certainty , patient follow - up and repeated analysis are necessary . proper management for timely treatment of a patient with insulinoma involves complex medical teamwork consisting of physicians from various specialties : endocrinology , internal medicine , surgery , pathology , medical imaging , oncology and , like in our case , psychiatry . Output:	insulinoma is a rare pancreatic islet cell tumor , the most common cause of hypoglycemia related to endogenous hyperinsulinism . we present the case of an adult patient with pancreatic insulinoma . the patient presented to our clinic after prior hospitalizations in diabetes and psychiatry hospital units for repeated episodes of loss of consciousness , sweating and tonic - clonic seizures . early detection of the cause is important for prompt initiation of therapy and to reduce symptoms . we describe our diagnostic and therapeutic strategies with references to previously published reports .
PubmedSumm2090	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: we are born in organizations , educated by organizations , and most of us spend much of our lives working for organizations . we spend much of our leisure time paying , playing , and praying in organizations . most of us will die in an organization , and when the time comes for burial , the largest organization of all the state must grant official permission . we are born in organizations , educated by organizations , and most of us spend much of our lives working for organizations . we spend much of our leisure time paying , playing , and praying in organizations . most of us will die in an organization , and when the time comes for burial , the largest organization of all the state must grant official permission . in many societies , thus , bureaucracy can be viewed as the predictable extension of social organizations within the societies . an organization is defined as a social system of consciously coordinated activities or forces of two or more persons explicitly created to achieve specific ends . not surprisingly , leading social scientists hold that 3 interrelated social processes , namely , urbanization , industrialization , and bureaucratization , are the most influential factors that have transformed american community life over the generations . indeed , immigration and globalization are contributing factors for such transformations . the institutional complex of major concern to us is health care that exemplifies the structures and functions of the bureaucracy . the structural aspect seeks to explain a feature of society as the predictable consequence of certain structural characteristics of society . the functional aspect , on the other hand , seeks to explain a feature of society in terms of the beneficial consequences it has for the larger social system . it is of interest to note that institutions are crescive in nature and have a strain of consistency , which means that institutions build over time and that they are all interrelated . for example , the institutions such as the family and health care are interrelated because the family is the unit of health because it is the unit of living . in brief , institutions are socially approved patterns of behavior . a medical industrial organization as an institution , therefore , can not be fully understood unless it is examined in itself as well as in relation to other institutions such as the family , social welfare , government , education , religion , and other organizational structures in the society . a society , from one perspective , can be viewed as falling somewhere between 2 polar pure types , ideal types , or mental constructs as either gemeinschaft ( g1 ) or gesellschaft ( g2 ) in orientation , or perhaps rural and urban ( fig 1 ) . the society - culture - personality ( scp ) model from a macroscopic perspective represents the global village , a nation , a community , an institution , or a group . there are a multitude of subsystems , comparable to subsystems in the human body , operating in any of these aggregates . bureaucracy , therefore , is the typical social and cultural milieu in which health care workers carry out their individual and common tasks in a society . thus , a health care professional does not simply work in a clinic , a hospital , or a department of a medical industrial complex ; but he or she works in a bureaucratic setting of a health care institution within a particular community of a specified society located either in a g1 or a g2 type of environment ( fig 1 ) . therefore , the purpose of this article is to present select concepts and theories of bureaucratic structures and functions so that chiropractic physicians and other health care professionals can use them in their respective practices . the scp model can be applied as an organizational instrument for assisting chiropractors in the diagnosis and treatment of their patients irrespective of locality . society - culture - personality forms an interlocking social system ( fig 1 ) . by definition , a social system is a pattern relationship of roles ; and as these roles become highly complex structures , they are called institutions ( i ) , that is , ways of taking care of the basic human needs of a society . as mentioned previously , a society , from one perspective , can be viewed as falling somewhere between 2 polar pure types , ideal types , or mental constructs , such as g1 and g2 in orientation . bureaucracy is a pyramiding of unit organizations of the social system in any complex society . bureaucracy contains several subsystems within the larger social system : the authority system , the power system , the status system , the organizational system , the security system , and the role system ( fig 1 ) . these are as follows ( fig 1 ) : freedom the employee is bound by contract to perform various functions during the day 's work , but is free to do what he / she wants after hours . hierarchy the structure of a bureaucracy is in the form of a hierarchy , with each member answerable to a higher authority or authorities . the members have defined spheres of competence and authority , and the duties and obligations of each group are specifically stated . positions are filled by a contract , and both the organization and the individual are bound by its terms . the procedure in a bureaucratic setting is highly regularized ; communication takes place through channels and according to established forms . discipline the members are supposed to carry out the work of their office , keep the secrets of the organization , and maintain an in - group feeling against outsiders . hierarchical form of social organization rationally geared to the achievement of precisely specified objectives by means of a division of labor based on demonstrated competence . as bureaucracy has come to define and to regulate important functions of other institutions within this society , health care has increasingly adopted such bureaucratic characteristics as positions and official record keeping . indeed , the appearance of bureaucratic administration in almost every sphere of life has not left the health care professions untouched . bureaucracy influences relations between chiropractic physicians and patients , between physicians and nurses , and other health - related professionals , permeating and influencing the whole structure of health services and the community as a whole . the structural aspects of bureaucracy in relation to the health care institution ( fig 1 ) can be analyzed from the viewpoint of typologies . alvin gouldner suggested that 3 patterns of bureaucracy are clearly distinguishable : mock , representative , and punishment centered . in mock bureaucracy , the informal organization of personnel circumvents the formal organization ; nonenforcement and nonobedience of rules are informally arranged by superordinates and subordinates with little tension or conflict ensuring . representative bureaucracy is marked by the endorsement and support of the rules by both groups ; tension and conflict are relatively absent . punishment - centered bureaucracy involves insistence on enforcement of the rules by one group and evasion of rules by the other ( either pattern can be adopted by the superordinate or subordinate groups ) ; consequently , tension and conflict are common in this type of work situation . stable patterns of behavior based on structural roles and specialized tasks are apparent . as a general rule , any organization large enough inhibits face - to - face relationships in a bureaucracy . when large numbers of people are added to an organization so that its goals may be accomplished , a hierarchy of authority emerges as a line - ranking of positions from top to bottom . each considers his or her function indispensable to the organization and expects his or her point of view to take precedence , especially in allocating funds for example in a medical industrial complex . the concepts of individual initiative , team work , along with authority are additional structural and functional factors that need to be taken into consideration within the medical industrial organization . a. earl swift asserts that all organizational behavior is ultimately founded upon human nature . human nature , according to swift , allows for 3 distinct decision - making systems , namely , individualism , collaboration , along with authority and power . individual initiative , also known as liberty , is a crucial aspect for an effective and successful organization . collaboration , commonly referred to as teamwork , is another significant factor in organizational relationships . teamwork does not allow any individual to manipulate and to dominate other employees because the group provides a check against individuals with bad motives , denying them the power to tyrannically dominate or exploit other people . when group decisions based upon consensus are established , a democratic approach to organizational relationships is fostered . authority and power are additional factors that should be taken into consideration if an organization is to continue to fulfill its mission and objectives . the board of trustees , as the chief policy - making group of a medical industrial organization , is there to enforce the mission and culture to ensure a proper balance of individualism , community , and authority . by establishing homeostasis , it allows for a diverse workforce to work together , as a professional community , to achieve the goals of the organization . under most circumstances , however , organizations work most effectively when all three types of decision making are balanced relatively equally , and it is the authoritarian aspect of the organization that controls this balance . given the above statements on the health care enterprise , the structural aspects of bureaucracy ( fig 1 ) are indeed linked to their functional dimensions as they relate to the health care institution ( fig 2 ) . thus , the structure and function of bureaucratic organizations are essential ingredients of analysis to bring about optimum levels of quality , affordability , availability , accessibility , and continuity in a health care institution irrespective of location whether g1 ( rural ) or g2 ( urban ) in orientation ( fig 3 ) . the functional aspect of bureaucracy in relation to the health care institution of a community or a society is closely linked to the interactional changes within a health care organizational setting such as a clinic or a hospital . furthermore , functional changes within the health care bureaucracy are affected when transactional and transformational changes are initiated in the system ( fig 2 ) . before scp that catalyst is social meaningful interaction ( smi ) ( fig 2 ) . as a person internalizes the culture of a society into his or her own personality , he or she develops a personality that is at once unique and distinctive and more or less adjusted to the demands of a society . in the development of smi , the actor enters society and establishes rapport . after this has been accomplished , social contact , the simplest unit of relationship between 2 or more persons , is possible . communication , which is involved in social contact , is the basis of smi . in a sense , it is smi . social contact plus communication results in social interaction , which is a sustained , reciprocal , purposeful , meaningful , and ( within limits ) predictable series of relationships between 2 or more socialized human beings . in the smi process , each actor takes the other(s ) into account , is aware of the other(s ) , and appraises the other(s ) . for example , interaction can take place between 2 individuals , between the individual and the group , between the individual and culture , and between the individual and mass communication . the smi is linked to socialization , a learning process in a social environment where the value - attitude system of a culture is internalized . it is important that chiropractic and other health care professionals recognize the implication of smi in terms of both quality and quantity to gain an understanding of the health care enterprise . for example , a chiropractic physician and chiropractic student should recognize the fact that patients , although presently in seeming isolation , are family members , and their reactions and behaviors demonstrate their family 's influence on their level of understanding and their attitude about the meaning and purpose of health care . the family , conversely , socializes the patient into a system of health care expectations that has been developed by the bureaucratic model . in the socialization process , other agencies , peer groups , and community pressure groups influence the patients view concerning health and illness . as previously stated , health care as an institution , therefore , can not be fully understood unless it is examined in itself as well as in relation to other institutions such as the family , social welfare , government , education , religion , and other organizational structures in the society . every social institution has some sort of structure or framework that helps to put concepts or purposes of the institution into the world of action so that it can serve the interest of society and the members who compose it . the family as a social institution has its framework or structure within which it can carry out the purposes for which it exists . the first is the family of origin ( orientation ) in which our earliest experiences take place . the second is the family of marriage ( or procreation ) in which we may enact the role of parent . it is within the network of familial relationships that we develop our attitudes and values toward health and illness . attitudes are tendencies to feel and act in certain ways . values , on the other hand , are measures of desirability . the family is the most universal of all human institutions . it varies widely in structure from the consanguineal type ( ie , extended kin groups that include a wide variety of related persons ) to conjugal families consisting simply of an adult pair ( male and female ) and their children . the conjugal family acts as a source of refuge in mass society a place where the individual may engage in genuinely personal relationships in a world that is largely impersonal . in the past , many authors have given us a description of different types of families . sorokin , for example , presents 3 types : the compulsive , the contractual , and the familistic . in the compulsive family , the bond holding members together is not love but force ; and the relationship is based on exploitation , cruelty , and deprivation . the contractual type brings profit and advancement to the participants but is devoid of love and hatred . the familistic type is based on mutual love between the spouses ; and it is characterized by devotion , sacrifice , solidarity , sharing , permanence , and stability . sorokin feels that the 3 types of families have been present regardless of one 's society but have changed in proportion with time . the contractual family , however , is the largest one in today 's western world . the type of family a person comes from can help us understand the behaviors of the patient and members of the family toward the sick person . for example , in severe coronary cases , increasing demands are made on the family to adjust their customary routines to the patient 's needs . one can expect , therefore , that if the family type was close ( familistic ) in which each member was concerned about the others before the illness , then there could be a greater willingness for members of the family to adjust their roles to help the sick person . on the contrary , if the family type was contractual and/or compulsive , then the family members would be less willing to make the sacrifices to aid the sick individual . structure within the institution of the family plays an essential role in the way stress is handled during a sudden crisis . the outcome of such a crisis will depend upon the type of familial relationship before the episode . for example research done at the family medicine unit at harvard medical school have demonstrated rather clearly that : common crises such as death of grandparents , change of residence , a loss of a father 's job , and a child 's being subjected to unusual pressure , occur four times more frequently in the two week period prior to the appearance of streptococcal infection than in the 2 weeks afterward . common crises such as death of grandparents , change of residence , a loss of a father 's job , and a child 's being subjected to unusual pressure , occur four times more frequently in the two week period prior to the appearance of streptococcal infection than in the 2 weeks afterward . the same studies have shown that : age , intimacy of contact , and family organization influenced the susceptibility to streptococcal infection . children of school age were most susceptible , and a spread of infection to other family members sharing the same bedroom was likely . age , intimacy of contact , and family organization influenced the susceptibility to streptococcal infection . children of school age were most susceptible , and a spread of infection to other family members sharing the same bedroom was likely . additional research demonstrates that streptococcal and staphylococcal infections are family disorders , and successful management responses requires consideration of the family group . another kind of crisis in which the type of family plays an important role is the biological inheritance factor . if one learns that he or she is a cause of disease that affects children , or learns that he or she is a recipient of a disease of a familiar nature , this knowledge can bring about complicated emotional problems in family interaction . for example , the birth of a deformed infant may be accompanied by a guilt reaction on the part of both parents . one can speculate about the emotional disturbances of the family in the specific case of muscular dystrophy that is carried by the female and attacks the male . the need of a parent to deny knowledge about such discomforting facts is understandable ; however , there is a tendency for the parent to believe that the facts must be disproved and to continually seek out advice to get different answers . at this stage of the crisis , there is a great need to see the family as a unit of treatment . whether or not the family is viewed as such will depend partially upon the type of familial relationships before the episode . another important role of the family is the patient - physician relationship concerning the care at the end of life . although many health care professionals are often uncomfortable discussing death and dying with their patients and families and feel that such discussion would be too difficult emotionally and thus ineffective for the patient and family , a recent study has shown that the reverse is true . almost 90% of caregivers ( ie , the family ) feel that such collaboration was not stressful , whereas 20% found it beneficial . . it can be the equivalent of a full - time job for 20% of caregivers and result in further financial burden . the average annual costs for caregiving in the united states can range from $ 3 billion to $ 6 billion . these stressors often lead families to seek long - term care ( ltc ) placement . caregivers who are older ( > 65 years of age ) and who feel a greater sense of burden are more likely to have their loved one in an ltc facility . although many caregivers experience symptoms of anxiety and depression ( 15%-20% ) before placement in an ltc facility , these symptoms did not change after placement , which is particularly true for spouses . two recent studies found that caregivers often experience a sense of relief after the passing of a loved one when it was preceded by ongoing suffering and significant burden to the caregiver . one study suggests that , in addition to the known risk of psychiatric morbidity of caregiving , there is a 60% higher risk of caregiver 's death when compared with noncaregiver 's controls . in recognizing the burden of caregiving , rabow et al recently proposed 5 areas of opportunities for caregivers to be of service to the family these are ( a ) promote communication , ( b ) promote advanced care planning and decision making , ( c ) support home care , ( d ) demonstrate empathy for patients and their family , and ( e ) participate in family grief and bereavement . in providing compassion and empathy , health care , an institution within american society , is like other institutions whereby structural subsystems and functional components are intertwined ( fig 3 ) . using a medical industrial complex , for example , one could identify certain organizational and functional goals such as education , research , and health care services . the organizational structure and functional components of the medical industrial bureaucracy are composed of a hierarchy of offices and positions . the board of trustees is the chief policy - making group within the medical industrial complex . it is made up partially of community representatives who safeguard the interests of the community within the organization . every department and service within the medical industrial complex have a hierarchy . within the hierarchy , the title and office that one holds confer status upon one , which varies depending upon the position in the organization . from the functional perspective , a professional person in a health care organization is exposed to the laws of authority , professional and bureaucratic ; both are essential for the functioning of the intricate health care enterprise . professional authority rightly has the freedom to act on the basis of professional skill and judgment on behalf of the individual patient in the particular situation regardless of bureaucratic rules . skipper and mumford asserted that the professional person derives his or her basic authority from outside the bureaucracy . for example , the health care professionals come to work in a hospital ( or a clinic ) with their own professional license to practice . on the other hand , the bureaucratic authority of the organization rightly demands regularity and conformity to regulations that make for dependability and predictability within the organizational network . thus , the medical staff usually defines the line in its activities , whereas the management authority is often restricted to matters of providing the means by which the doctor 's orders may be successfully carried out . from the viewpoints of the functions of bureaucracy in the health care institution , it is important to note that max weber 's approach is essentially formal , outlining the blueprint character of bureaucracy the rule of rules rather than of men , the role of hierarchy , specialization , and experience , the development of the office as a career . the functions of bureaucracy ( fig 2 ) are also linked to 5 factors that are directly related to the positive outcomes in any health care organization . these variables are quality , availability , accessibility , affordability , and continuity . in the functioning of the health care bureaucracy , the factors noted above may bring about sociopsychological pressures to the professionals within the organization . the acronym sara ( surprise - anxiety - rejection - acceptance ) is most relevant in analyzing the functional changes of personality within the system . thus , it may be hypothesized that the interaction between the various units in a health care bureaucratic system will either be associative , tending toward a mutual sharing of responsibility , power , and authority , or disassociative , tending toward the tension and conflict situations ( fig 2 ) . if one takes the variable of power into consideration , certain associative and dissassociative patterns are possible . one of the significant patterns today is the relationship between and among health care professionals . as the recipients of the chiropractic physician 's orders for his or her patterns , the nurse is obligated to carry out these orders in a professional competent manner ; but at the same time , he or she is a hired employee of the hospital , clinic , or doctor 's office and consequently subject to all the rules and regulations of the administrative organization . * furthermore , the demands of patient care , especially those of an emergency nature , can not be accomplished within the framework of administrative rules . thus , the nurse can be in a conflict situation between the expectations of the chiropractic physician that his or her orders be carried out and the expectations of the administration that administrative procedures will be complied with . the dysfunctions of bureaucracy were pointed out by men like veblen , dewey , and warnotte who documented concepts of trained incapacity , occupational psychosis , and professional deformation respectively . refers to that condition in which one 's abilities function as inadequacies or blind spots , preventing one from adjusting correctly to the changed situation . thus , skills and training that have been successfully applied in the past may under changed conditions result in inappropriate responses . dewey 's concept of occupational psychosis is founded upon some observations from the humdrum activities of life in which people develop certain preferences , antipathies , discriminations , and points of interest . by psychosis , dewey means a profound character of the mind . these psychoses originate from demands put upon the individual by the organizing of his occupation role . however , there are forces at work that tend to lessen the severity of the potential problems resulting from the social structure of a medical industrial complex . coe asserts that : one of these is the ideology of service , an historical legacy of the middle ages , expressed in modern terms as commitment to a job . a second force is the development of informal groups within the formal organization which permit certain activities to be accomplished regardless of the potential blocks of the social structure . one of these is the ideology of service , an historical legacy of the middle ages , expressed in modern terms as commitment to a job . a second force is the development of informal groups within the formal organization which permit certain activities to be accomplished regardless of the potential blocks of the social structure . insofar as the chiropractic physician and nurse are concerned , the relationship shifts from time to time and from place to place . it varies according to the generation each belongs to , to the size of the community and the hospital setting , and to the field of specialization in nursing and medicine . the getzels - guba model gives us further insight into the structural and functional aspects of organizational bureaucracy . the nomothetic or normative dimension of the model is composed of institutions , roles , and expectations . the idiographic or personal dimension of the model is characterized by the individual , personality , and need - disposition . social behavior is the result of the interactions between 2 sets of motives : the nomothetic / normative dimension and the idiographic / personal dimension . the getzels - guba model asserts that , within an institution , an individual has a purposeful role and is required to meet certain expectations as a member of the organization . need - disposition and role expectation can be viewed as structural elements in patient / chiropractic physician rapport and in professional and ethical functions . if there is a lack of homeostasis between needs / structures and roles / functions within an organization , it may produce stress , strain , and anxiety that may negatively affect professional relationships and the continuity of health care . according to w. edwards deming 's out of the crisis , principles such as efficiency and effectiveness need to be aligned so that the members of the health care organization take part in shared decision making . if a health care organization lacks these principles , it may lead to identity and role confusion at work ( in a hospital or a clinic ) , creating additional stress and strain that may affect the proper functioning of the organization . the patient , who is the customer in the health care organization , can be affected positively or negatively if such issues within the health care organization remain unsolved . if stress , strain , and anxiety are created from occupational relationships ( and remain unresolved ) , this could result in what gregory bateson et al refer to as the double bind theory . given this theory , the recipient of double bind is given contradictory messages , causing confusion in self - concept that may lead to an element of uncertainty within the personality of the individual . for example , a medical organization may instill in their employees the importance of teamwork and shared decision making ; but when employees actually voices their concern , they are greeted with resistance and disapproval . this element of uncertainty along with identity and role confusion can lead to what emile durkheim called anomie ( commonly known as a lack of norms ) . this type of behavior within a medical industrial organization can increase the level of stress and strain , leading to neurosis ; and if it remains untreated , it can result in psychosis that affects the functioning within the organizational system . society - culture - personality forms an interlocking social system ( fig 1 ) . by definition , a social system is a pattern relationship of roles ; and as these roles become highly complex structures , they are called institutions ( i ) , that is , ways of taking care of the basic human needs of a society . as mentioned previously , a society , from one perspective , can be viewed as falling somewhere between 2 polar pure types , ideal types , or mental constructs , such as g1 and g2 in orientation . bureaucracy is a pyramiding of unit organizations of the social system in any complex society . bureaucracy contains several subsystems within the larger social system : the authority system , the power system , the status system , the organizational system , the security system , and the role system ( fig 1 ) . these are as follows ( fig 1 ) : freedom the employee is bound by contract to perform various functions during the day 's work , but is free to do what he / she wants after hours . hierarchy the structure of a bureaucracy is in the form of a hierarchy , with each member answerable to a higher authority or authorities . the members have defined spheres of competence and authority , and the duties and obligations of each group are specifically stated . positions are filled by a contract , and both the organization and the individual are bound by its terms . the procedure in a bureaucratic setting is highly regularized ; communication takes place through channels and according to established forms . discipline the members are supposed to carry out the work of their office , keep the secrets of the organization , and maintain an in - group feeling against outsiders . hierarchical form of social organization rationally geared to the achievement of precisely specified objectives by means of a division of labor based on demonstrated competence . as bureaucracy has come to define and to regulate important functions of other institutions within this society , health care has increasingly adopted such bureaucratic characteristics as positions and official record keeping . indeed , the appearance of bureaucratic administration in almost every sphere of life has not left the health care professions untouched . bureaucracy influences relations between chiropractic physicians and patients , between physicians and nurses , and other health - related professionals , permeating and influencing the whole structure of health services and the community as a whole . the structural aspects of bureaucracy in relation to the health care institution ( fig 1 ) can be analyzed from the viewpoint of typologies . alvin gouldner suggested that 3 patterns of bureaucracy are clearly distinguishable : mock , representative , and punishment centered . in mock bureaucracy , the informal organization of personnel circumvents the formal organization ; nonenforcement and nonobedience of rules are informally arranged by superordinates and subordinates with little tension or conflict ensuring . representative bureaucracy is marked by the endorsement and support of the rules by both groups ; tension and conflict are relatively absent . punishment - centered bureaucracy involves insistence on enforcement of the rules by one group and evasion of rules by the other ( either pattern can be adopted by the superordinate or subordinate groups ) ; consequently , tension and conflict are common in this type of work situation . stable patterns of behavior based on structural roles and specialized tasks are apparent . as a general rule , any organization large enough inhibits face - to - face relationships in a bureaucracy . when large numbers of people are added to an organization so that its goals may be accomplished , a hierarchy of authority emerges as a line - ranking of positions from top to bottom . each considers his or her function indispensable to the organization and expects his or her point of view to take precedence , especially in allocating funds for example in a medical industrial complex . the concepts of individual initiative , team work , along with authority are additional structural and functional factors that need to be taken into consideration within the medical industrial organization . a. earl swift asserts that all organizational behavior is ultimately founded upon human nature . human nature , according to swift , allows for 3 distinct decision - making systems , namely , individualism , collaboration , along with authority and power . individual initiative , also known as liberty , is a crucial aspect for an effective and successful organization . collaboration , commonly referred to as teamwork , is another significant factor in organizational relationships . teamwork does not allow any individual to manipulate and to dominate other employees because the group provides a check against individuals with bad motives , denying them the power to tyrannically dominate or exploit other people . when group decisions based upon consensus are established , a democratic approach to organizational relationships is fostered . authority and power are additional factors that should be taken into consideration if an organization is to continue to fulfill its mission and objectives . the board of trustees , as the chief policy - making group of a medical industrial organization , is there to enforce the mission and culture to ensure a proper balance of individualism , community , and authority . by establishing homeostasis , it allows for a diverse workforce to work together , as a professional community , to achieve the goals of the organization . under most circumstances , however , organizations work most effectively when all three types of decision making are balanced relatively equally , and it is the authoritarian aspect of the organization that controls this balance . given the above statements on the health care enterprise , the structural aspects of bureaucracy ( fig 1 ) are indeed linked to their functional dimensions as they relate to the health care institution ( fig 2 ) . thus , the structure and function of bureaucratic organizations are essential ingredients of analysis to bring about optimum levels of quality , affordability , availability , accessibility , and continuity in a health care institution irrespective of location whether g1 ( rural ) or g2 ( urban ) in orientation ( fig 3 ) . the functional aspect of bureaucracy in relation to the health care institution of a community or a society is closely linked to the interactional changes within a health care organizational setting such as a clinic or a hospital . furthermore , functional changes within the health care bureaucracy are affected when transactional and transformational changes are initiated in the system ( fig 2 ) . before scp that catalyst is social meaningful interaction ( smi ) ( fig 2 ) . as a person internalizes the culture of a society into his or her own personality , he or she develops a personality that is at once unique and distinctive and more or less adjusted to the demands of a society . in the development of smi , the actor enters society and establishes rapport . after this has been accomplished , social contact , the simplest unit of relationship between 2 or more persons , is possible . communication , which is involved in social contact , is the basis of smi . in a sense , it is smi . social contact plus communication results in social interaction , which is a sustained , reciprocal , purposeful , meaningful , and ( within limits ) predictable series of relationships between 2 or more socialized human beings . in the smi process , each actor takes the other(s ) into account , is aware of the other(s ) , and appraises the other(s ) . for example , interaction can take place between 2 individuals , between the individual and the group , between the individual and culture , and between the individual and mass communication . the smi is linked to socialization , a learning process in a social environment where the value - attitude system of a culture is internalized . it is important that chiropractic and other health care professionals recognize the implication of smi in terms of both quality and quantity to gain an understanding of the health care enterprise . for example , a chiropractic physician and chiropractic student should recognize the fact that patients , although presently in seeming isolation , are family members , and their reactions and behaviors demonstrate their family 's influence on their level of understanding and their attitude about the meaning and purpose of health care . the family , conversely , socializes the patient into a system of health care expectations that has been developed by the bureaucratic model . in the socialization process , other agencies , peer groups , and community pressure groups influence the patients view concerning health and illness . as previously stated , health care as an institution , therefore , can not be fully understood unless it is examined in itself as well as in relation to other institutions such as the family , social welfare , government , education , religion , and other organizational structures in the society . every social institution has some sort of structure or framework that helps to put concepts or purposes of the institution into the world of action so that it can serve the interest of society and the members who compose it . the family as a social institution has its framework or structure within which it can carry out the purposes for which it exists . the first is the family of origin ( orientation ) in which our earliest experiences take place . the second is the family of marriage ( or procreation ) in which we may enact the role of parent . it is within the network of familial relationships that we develop our attitudes and values toward health and illness . it varies widely in structure from the consanguineal type ( ie , extended kin groups that include a wide variety of related persons ) to conjugal families consisting simply of an adult pair ( male and female ) and their children . the conjugal family acts as a source of refuge in mass society a place where the individual may engage in genuinely personal relationships in a world that is largely impersonal . in the past , many authors have given us a description of different types of families . sorokin , for example , presents 3 types : the compulsive , the contractual , and the familistic . in the compulsive family , the bond holding members together is not love but force ; and the relationship is based on exploitation , cruelty , and deprivation . the contractual type brings profit and advancement to the participants but is devoid of love and hatred . the familistic type is based on mutual love between the spouses ; and it is characterized by devotion , sacrifice , solidarity , sharing , permanence , and stability . sorokin feels that the 3 types of families have been present regardless of one 's society but have changed in proportion with time . the contractual family , however , is the largest one in today 's western world . the type of family a person comes from can help us understand the behaviors of the patient and members of the family toward the sick person . for example , in severe coronary cases , increasing demands are made on the family to adjust their customary routines to the patient 's needs . one can expect , therefore , that if the family type was close ( familistic ) in which each member was concerned about the others before the illness , then there could be a greater willingness for members of the family to adjust their roles to help the sick person . on the contrary , if the family type was contractual and/or compulsive , then the family members would be less willing to make the sacrifices to aid the sick individual . structure within the institution of the family plays an essential role in the way stress is handled during a sudden crisis . the outcome of such a crisis will depend upon the type of familial relationship before the episode . research done at the family medicine unit at harvard medical school have demonstrated rather clearly that : common crises such as death of grandparents , change of residence , a loss of a father 's job , and a child 's being subjected to unusual pressure , occur four times more frequently in the two week period prior to the appearance of streptococcal infection than in the 2 weeks afterward . common crises such as death of grandparents , change of residence , a loss of a father 's job , and a child 's being subjected to unusual pressure , occur four times more frequently in the two week period prior to the appearance of streptococcal infection than in the 2 weeks afterward . the same studies have shown that : age , intimacy of contact , and family organization influenced the susceptibility to streptococcal infection . children of school age were most susceptible , and a spread of infection to other family members sharing the same bedroom was likely . age , intimacy of contact , and family organization influenced the susceptibility to streptococcal infection . children of school age were most susceptible , and a spread of infection to other family members sharing the same bedroom was likely . additional research demonstrates that streptococcal and staphylococcal infections are family disorders , and successful management responses requires consideration of the family group . another kind of crisis in which the type of family plays an important role is the biological inheritance factor . if one learns that he or she is a cause of disease that affects children , or learns that he or she is a recipient of a disease of a familiar nature , this knowledge can bring about complicated emotional problems in family interaction . for example , the birth of a deformed infant may be accompanied by a guilt reaction on the part of both parents . one can speculate about the emotional disturbances of the family in the specific case of muscular dystrophy that is carried by the female and attacks the male . the need of a parent to deny knowledge about such discomforting facts is understandable ; however , there is a tendency for the parent to believe that the facts must be disproved and to continually seek out advice to get different answers . at this stage of the crisis , there is a great need to see the family as a unit of treatment . whether or not the family is viewed as such will depend partially upon the type of familial relationships before the episode . another important role of the family is the patient - physician relationship concerning the care at the end of life . although many health care professionals are often uncomfortable discussing death and dying with their patients and families and feel that such discussion would be too difficult emotionally and thus ineffective for the patient and family , a recent study has shown that the reverse is true . almost 90% of caregivers ( ie , the family ) feel that such collaboration was not stressful , whereas 20% found it beneficial . . it can be the equivalent of a full - time job for 20% of caregivers and result in further financial burden . the average annual costs for caregiving in the united states can range from $ 3 billion to $ 6 billion . these stressors often lead families to seek long - term care ( ltc ) placement . caregivers who are older ( > 65 years of age ) and who feel a greater sense of burden are more likely to have their loved one in an ltc facility . although many caregivers experience symptoms of anxiety and depression ( 15%-20% ) before placement in an ltc facility , these symptoms did not change after placement , which is particularly true for spouses . two recent studies found that caregivers often experience a sense of relief after the passing of a loved one when it was preceded by ongoing suffering and significant burden to the caregiver . one study suggests that , in addition to the known risk of psychiatric morbidity of caregiving , there is a 60% higher risk of caregiver 's death when compared with noncaregiver 's controls . in recognizing the burden of caregiving , rabow et al recently proposed 5 areas of opportunities for caregivers to be of service to the family . these are ( a ) promote communication , ( b ) promote advanced care planning and decision making , ( c ) support home care , ( d ) demonstrate empathy for patients and their family , and ( e ) participate in family grief and bereavement . in providing compassion and empathy , health care , an institution within american society , is like other institutions whereby structural subsystems and functional components are intertwined ( fig 3 ) . using a medical industrial complex , for example , one could identify certain organizational and functional goals such as education , research , and health care services . the organizational structure and functional components of the medical industrial bureaucracy are composed of a hierarchy of offices and positions . the board of trustees is the chief policy - making group within the medical industrial complex . it is made up partially of community representatives who safeguard the interests of the community within the organization . every department and service within the medical industrial complex have a hierarchy . within the hierarchy , the title and office that one holds confer status upon one , which varies depending upon the position in the organization . from the functional perspective , a professional person in a health care organization is exposed to the laws of authority , professional and bureaucratic ; both are essential for the functioning of the intricate health care enterprise . professional authority rightly has the freedom to act on the basis of professional skill and judgment on behalf of the individual patient in the particular situation regardless of bureaucratic rules . skipper and mumford asserted that the professional person derives his or her basic authority from outside the bureaucracy . for example , the health care professionals come to work in a hospital ( or a clinic ) with their own professional license to practice . on the other hand , the bureaucratic authority of the organization rightly demands regularity and conformity to regulations that make for dependability and predictability within the organizational network . thus , the medical staff usually defines the line in its activities , whereas the management authority is often restricted to matters of providing the means by which the doctor 's orders may be successfully carried out . from the viewpoints of the functions of bureaucracy in the health care institution , it is important to note that max weber 's approach is essentially formal , outlining the blueprint character of bureaucracy the rule of rules rather than of men , the role of hierarchy , specialization , and experience , the development of the office as a career . the functions of bureaucracy ( fig 2 ) are also linked to 5 factors that are directly related to the positive outcomes in any health care organization . these variables are quality , availability , accessibility , affordability , and continuity . in the functioning of the health care bureaucracy , the factors noted above may bring about sociopsychological pressures to the professionals within the organization . the acronym sara ( surprise - anxiety - rejection - acceptance ) is most relevant in analyzing the functional changes of personality within the system . thus , it may be hypothesized that the interaction between the various units in a health care bureaucratic system will either be associative , tending toward a mutual sharing of responsibility , power , and authority , or disassociative , tending toward the tension and conflict situations ( fig 2 ) . if one takes the variable of power into consideration , certain associative and dissassociative patterns are possible . one of the significant patterns today is the relationship between and among health care professionals . as the recipients of the chiropractic physician 's orders for his or her patterns , the nurse is obligated to carry out these orders in a professional competent manner ; but at the same time , he or she is a hired employee of the hospital , clinic , or doctor 's office and consequently subject to all the rules and regulations of the administrative organization . * furthermore , the demands of patient care , especially those of an emergency nature , can not be accomplished within the framework of administrative rules . thus , the nurse can be in a conflict situation between the expectations of the chiropractic physician that his or her orders be carried out and the expectations of the administration that administrative procedures will be complied with . the dysfunctions of bureaucracy were pointed out by men like veblen , dewey , and warnotte who documented concepts of trained incapacity , occupational psychosis , and professional deformation respectively . refers to that condition in which one 's abilities function as inadequacies or blind spots , preventing one from adjusting correctly to the changed situation . thus , skills and training that have been successfully applied in the past may under changed conditions result in inappropriate responses . dewey 's concept of occupational psychosis is founded upon some observations from the humdrum activities of life in which people develop certain preferences , antipathies , discriminations , and points of interest . by psychosis , dewey means a profound character of the mind . these psychoses originate from demands put upon the individual by the organizing of his occupation role . however , there are forces at work that tend to lessen the severity of the potential problems resulting from the social structure of a medical industrial complex . coe asserts that : one of these is the ideology of service , an historical legacy of the middle ages , expressed in modern terms as commitment to a job . a second force is the development of informal groups within the formal organization which permit certain activities to be accomplished regardless of the potential blocks of the social structure . one of these is the ideology of service , an historical legacy of the middle ages , expressed in modern terms as commitment to a job . a second force is the development of informal groups within the formal organization which permit certain activities to be accomplished regardless of the potential blocks of the social structure . insofar as the chiropractic physician and nurse are concerned , the relationship shifts from time to time and from place to place . it varies according to the generation each belongs to , to the size of the community and the hospital setting , and to the field of specialization in nursing and medicine . the getzels - guba model gives us further insight into the structural and functional aspects of organizational bureaucracy . the nomothetic or normative dimension of the model is composed of institutions , roles , and expectations . the idiographic or personal dimension of the model is characterized by the individual , personality , and need - disposition . social behavior is the result of the interactions between 2 sets of motives : the nomothetic / normative dimension and the idiographic / personal dimension . the getzels - guba model asserts that , within an institution , an individual has a purposeful role and is required to meet certain expectations as a member of the organization . need - disposition and role expectation can be viewed as structural elements in patient / chiropractic physician rapport and in professional and ethical functions . if there is a lack of homeostasis between needs / structures and roles / functions within an organization , it may produce stress , strain , and anxiety that may negatively affect professional relationships and the continuity of health care . according to w. edwards deming 's out of the crisis , principles such as efficiency and effectiveness need to be aligned so that the members of the health care organization take part in shared decision making . if a health care organization lacks these principles , it may lead to identity and role confusion at work ( in a hospital or a clinic ) , creating additional stress and strain that may affect the proper functioning of the organization . the patient , who is the customer in the health care organization , can be affected positively or negatively if such issues within the health care organization remain unsolved . if stress , strain , and anxiety are created from occupational relationships ( and remain unresolved ) , this could result in what gregory bateson et al refer to as the double bind theory . given this theory , the recipient of double bind is given contradictory messages , causing confusion in self - concept that may lead to an element of uncertainty within the personality of the individual . for example , a medical organization may instill in their employees the importance of teamwork and shared decision making ; but when employees actually voices their concern , they are greeted with resistance and disapproval . this element of uncertainty along with identity and role confusion can lead to what emile durkheim called anomie ( commonly known as a lack of norms ) . this type of behavior within a medical industrial organization can increase the level of stress and strain , leading to neurosis ; and if it remains untreated , it can result in psychosis that affects the functioning within the organizational system . this discussion examined some select concepts of bureaucratic structures and functions so that chiropractic physicians and other health care professionals can use them accordingly . we have focused upon the relationships of scp and smi upon the structural and functional aspects of bureaucracy in the health care institution of a society . it is hoped that present and future chiropractic physicians and other health care professionals will internalize and make use of these select concepts and theories in their future roles as health care givers ( figs 1 and 2 ) . implicit in our analysis of the health care bureaucratic structures and functions in a society is the focus that future chiropractic and other health care professionals integrate , synthesize , and actualize values and virtues such as empathy , integrity , excellence , diversity , compassion , caring , and understanding with a deep commitment to self - reflection so that they can deliver care that involves the ingredients of quality , affordability , availability , accessibility , and continuity for their patients . if these values are taught constructively in any health care bureaucracy , then future health care professionals will be able to transmit the traditions of excellence , dedication , and creativity within the health care profession . on balance , it is asserted that chiropractic physicians recognize the implications of the structures and functions of the health care bureaucracy for the prognosis , treatment , and diagnosis of a patient especially because a bureaucracy is a phenomenon of every highly urbanized and industrialized society today . indeed , it is an environment in which chiropractic physicians and other health care professionals are educated and trained , the organization in which the overwhelming majority will work through their lives . as bureaucracy has tended to define and to regulate the important functions of all institutions within this society , health care has increasingly been transformed into a bureaucratic activity both structurally and functionally . stable patterns of behavior based on structured statuses , roles , and specialized tasks are linked to life or death issues . the explosive growth of scientific knowledge , the rapid increases in technology , and the impact of globalization produce ever more specialization to be incorporated and coordinated in the present health care structures and functions . such a concern of care epitomizes the statement promulgated by wilson hoff , we must recognize that the adequacy of health services depends as much upon the organization of health personnel and the combinations with other resources as is does upon their numbers alone . indeed , a concern for care further documented the diction noted by prof frances weld peabody of harvard medical school , namely , the secret of the care of the patient is in the caring of the patient . Output:	objectivethe purpose of this article is to present select concepts and theories of bureaucratic structures and functions so that chiropractic physicians and other health care professionals can use them in their respective practices . the society - culture - personality model can be applied as an organizational instrument for assisting chiropractors in the diagnosis and treatment of their patients irrespective of locality.discussionsociety-culture-personality and social meaningful interaction are examined in relationship to the structural and functional aspects of bureaucracy within the health care institution of a society . implicit in the examination of the health care bureaucratic structures and functions of a society is the focus that chiropractic physicians and chiropractic students learn how to integrate , synthesize , and actualize values and virtues such as empathy , integrity , excellence , diversity , compassion , caring , and understanding with a deep commitment to self-reflection.conclusionit is essential that future and current chiropractic physicians be aware of the structural and functional aspects of an organization so that chiropractic and other health care professionals are able to deliver care that involves the ingredients of quality , affordability , availability , accessibility , and continuity for their patients .
PubmedSumm2091	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: already in 1949 hebb proposed that simultaneous neuronal firing stimulates synaptic plasticity . later several studies found evidence for experience - dependent neurogenesis in the hippocampi of adult mice ( for a review see ) . today there is accumulating evidence that also the human brain continues to be shaped by experience throughout adulthood [ 35 ] . these adaptive changes have been shown to take place on structural as well as functional level [ 69 ] . a practicable approach to study experience - dependent plasticity in humans recently , a number of studies have been published that investigated the effect of training on the functional architecture of the brain by resting - state fmri ( rs - fmri ) ( for a review see [ 9 , 10 ] ) . resting - state functional connectivity is commonly defined as temporal correlations of spontaneous , low frequency fluctuations of the bold signal between brain areas during rest due to common history of coactivation . as such , it allows a task - independent assessment of training - related changes in brain function [ 1114 ] . motor training varied from joystick tracking tasks , chopstick handling , finger tapping , and force - field learning to whole - body balancing and aerobic fitness training . training duration varied from 11 minutes to several weeks or months . in the cognitive domain training comprised working memory training [ 21 , 22 ] , multitasking , and logical reasoning and duration varied from 4 weeks to 3 months . newer approaches used also video - gaming and fmri based neurofeedback [ 26 , 27 ] . within the motor domain , several research groups investigated different kinds of motor skills training with varying duration , intensity , and complexity . overall , changes in intrinsic functional connectivity were located in sensorimotor and cerebellar areas . in these areas both intrinsic functional connectivity increases [ 15 , 1719 ] and decreases [ 1619 ] have been found ; decreases were rather associated with cerebellar regions [ 16 , 18 ] . in the studies conducted by taubert et al . and intrinsic functional connectivity decreased back to baseline whereas decreases were found by the groups of yoo et al . and vahdat et al . ; there , the intrinsic functional connectivity after the training was reduced compared to before the intervention . in the cognitive domain , training rather affected intrinsic functional connectivity between frontal and parietal areas [ 21 , 22 , 24 ] . however , the precise location of training - related change in intrinsic functional connectivity differs between studies . regarding the variety of changes found by different research groups , the training effects seem to be rather specific to the content of the training , the duration , the intensity , and the timing of the resting - state quantification . however , the studies mentioned before show that changes in intrinsic functional connectivity can reliably be induced by training , that is , experience , across a variety of domains . the majority of published intervention studies investigated the effect of unimodal training . within the field of healthy aging research the question rises if combined interventions might be more successful than unimodal interventions ( for a review see ) . also in the context of studies investigating effects of physical exercise on neuroplasticity and cognition it is suggested that adding cognitive training might enhance the beneficial effect of physical training ( for a review see ) . yet , there are only few studies that focus on the effect of combined interventions . to our knowledge , there are only two studies exploring the effect of a multimodal training using neuroimaging techniques [ 30 , 31 ] . in both studies , physical and cognitive training were performed apart from each other . in li et al . 's study older adults took part in tai chi exercises at one time and memory training and supportive group counselling on another time . they found increased resting - state connectivity between the medial prefrontal cortex and the medial temporal lobe . in holzschneider however , only task - based fmri changes were quantified . after combined training , changes in brain activation and changes in cardiovascular fitness correlated positively in the medial frontal gyrus and the cuneus . here , we investigated the effect of combined whole - body motor coordination training with integrated cognitive exercises in healthy adults . lutz and colleagues ( life kinetik : http://www.lifekinetik.de/ ) developed a multimodal training that combines coordinative , cognitive , and visual tasks in a way that the physical exercise is performed while participants are cognitively challenged at the same time . the training consists of combinations of motor activity and cognitive challenges and the training of visual perception , especially the perception of the peripheral visual field . moving limbs in different unusual combinations , catching , and throwing objects , thus training the visual perception and limb - eye coordination , is a basic characteristic of the training . moreover , the training tasks are not practiced to perfection but are modified after a few minutes or whenever the performance reaches about 60% . in addition to the avoidance of boredom and frustration , this is supposed to stimulate the brain to constantly adapt to new unfamiliar challenges . our motivation was to test a training concept that is flexible and interesting for the participants and includes cognitive and motor elements . although the life kinetik training was originally designed to train the coordination of athletes ( soccer players , skiers ) the difficulty of the task can easily be adapted to the capabilities of patient populations . based on the assumption that spontaneous activity reflects the history of coactivation within a local brain network or between brain regions [ 26 , 32 ] we expect increases in resting - state connectivity of those brain regions probably involved in the exercises and tasks . its connections seem to decrease with age and diminished in mild cognitive impairment ( mci ) and alzheimer 's disease ( ad ) [ 35 , 36 ] . as the training includes unusual pattern of motoric activity in combination with cognitive task , we expect the connectivity of the thalamus to increase . all the exercises and tasks involve some motor action ; hence we expect changes in the primary motor area ( ba4 , m1 ) and the premotor area ( ba6 ) because not only the execution but also the constant alertness to perform an action is involved in the task . in particular the connectivity to the right motor areas may be increased because the exercises include a considerable amount of movement of the left limbs , which is challenging for the right handed participants . the cerebellum is highly involved in motor activity and learning and the functional connections reflect the connections of the cortex [ 37 , 38 ] so we can expect some changes in its connectivity as well . the frontal eye field ( fef ) , a brain region responsible for eye movement and gaze control , is known to be altered in the course of learning to handle moving objects [ 3941 ] , which is also part of the exercise , except that this is not trained to perfection like in juggling . the whole visual cortex is additionally challenged by the attempt to train the peripheral vision and the manipulation of different moving objects and due to the possibility of assigning the requested action via a visual stimulus ( specific gesture of the trainer or colours ) . so the primary as well as the secondary visual cortices ( ba17 , ba18 , and ba19 ) are expected to change their connection to other brain regions . each exercise or task consists of chains of movements , which alternate randomly . the prompt to change hence , we expect an increased connectivity between auditory areas ( primary and secondary ba41 and ba42 ) and other brain regions especially the motor and premotor area and as a result of repeated coactivation . the functional connectivity from and to the dorsolateral prefrontal cortex ( dlpfc ) may be increased because the working memory is involved in linking the action or movement to the assigned command or prompt . the anterior cingulate cortex ( acc ) is also expected to be involved as a region needed for error detection and impulse control and might accordingly change the connection to other brain regions . 32 right handed subjects with no history of psychiatric or neurological illness were included in the study . 21 individuals ( 12 females , mean age 48 ( 9 ) years ) participated in eleven or twelve of the 13 life kinetik training sessions ( 1 hour per week ) . the other 11 subjects ( 7 females , mean age 49 ( 8 ) years ) were interested in the training but were not able to attend due to their time schedule but completed two mri scans . the study was approved by the ethics committee of the medical faculty mannheim , university of heidelberg , and performed in accordance with the declaration of helsinki . life kinetik training pursues the goal to combine motor coordination exercises with cognitive training with an emphasis on working memory . additionally , most of the time one or more pieces of sports equipment ( e.g. , ball , racket , juggling balls , and scarves ) are used which have to be thrown , caught , bounced , or similarly manipulated . the cognitive aspect comes into play by assigning distinct motor tasks to different visual or auditory cues ( symbols / key words ) . for example , a red flag might indicate bouncing a ball with the left hand while a blue flag indicates throwing and catching a ball with the right hand . the same movement - cue coupling can be done with semantic categories , for example , city names , animals , or trees . these pairs of motor task and specific cue have to be memorized during one training session . the randomization of cues is self - evident . within one training session ( 1 hour per week ) approximately 6 different types of exercises have been performed either in groups , in pairs , or by oneself . an essential aspect of this combined training is that the exercises are not trained until automatized . as soon as participant 's performance reaches about 60% correct trials the task demands are changed and new combinations of symbols and movements are introduced . moreover , cross talk of the hemispheres is fostered by including movements where limbs purposefully cross the sagittal midline ( e.g. , to catch a ball arriving at the left side of the body with the right hand ) . in total , there were 13 training sessions of 1 hour per week of which our participants followed at least 11 ; that is , the minimal training duration was 11 hours across a period of 13 weeks . functional and anatomical data were acquired from each participant within 2 weeks before the start of the first training session and within 2 weeks after the last training session on a 3 tesla mri scanner ( magnetom trio with tim technology , siemens medical service , erlangen , germany ) equipped with a 32-channel head coil . 200 rs - fmri images were acquired with gradient echo t2-weighted echo - planar - imaging sequence ( te = 28 ms , tr = 1.79 s , fov = 192 mm 192 mm , matrix size = 64 64 , and total duration is 6 min ) . a volume comprised 34 slices in ac - pc orientation with a thickness of 3 mm and slice gap of 1 mm . a t1-weighted anatomical image was also recorded ( te = 3.03 ms , tr = 2.3 s , 192 slices and fov = 256 mm 256 mm , matrix size is 256 256 , and slice thickness is 1 mm ) . data were preprocessed and analyzed using spm12 ( the wellcome department of cognitive neurology , london , uk , http://www.fil.ion.ucl.ac.uk/spm/software/spm12/ ) . all functional images were slice - time corrected and realigned to the first volume using a six - parameter rigid body transformation . the movement was not more than 1.5 mm in each subject , so no one had to be removed . segmented gray matter and white matter images of all participants were used to construct a study specific template using dartel . the template was normalized to mni space and all images , anatomical and functional , were normalized to this template using the according flow fields . the smoothing kernel for the functional images was 8 mm and 2 mm for the anatomical image . functional connectivity analyses were carried out using the conn - fmri functional connectivity toolbox v14 ( http://www.nitrc.org/projects/conn ) . the modest test - retest reliability of the rs - fmri seems attributable to remaining noise after preprocessing , adding nonneural correlation to the bold signal . removing the noise is a possibility to increase the reliability of rs - fmri data . this method extracts principal components ( 5 each ) from wm and csf time series . these components are added as confounds in the denoising step of the conn toolbox [ 43 , 45 ] . the six head motion parameters derived from spatial motion correction were also added as confounds . we did not perform global signal regression as the discussion about the impact is still ongoing [ 44 , 46 ] and it is not available on the conn toolbox . as recommended band - pass filtering was performed with a frequency window of 0.01 to 0.1 hz . seed - to - voxel and roi - to - roi functional connectivity maps were created for each participant . the roi - to - roi analysis was used to identify possible differences between trainees and control subjects at pretraining and to verify that brain networks of control subjects did not change over time . for this analysis we used all the provided brodmann areas . bivariate - correlation analyses were used to determine the linear association of the bold time series between each pair of sources and a fisher z transformation was applied . individual seed - to - voxel and roi - to - roi maps were entered into a second - level analysis . a within group roi - to - roi analysis for the control group tested the stability of the connectivity over time . a between groups roi - to - roi analysis verified the lack of differences between the two groups for the first measurement . first we used the posterior cingulate and the medial prefrontal cortex as seed region and verified the occurrence of the default mode network in each group and to both time - points . the second seed - to - voxel analysis was used to examine differences in connectivity changes in a 2 2 factorial analysis with time by training interaction ( group time ; contrast 1 + 1 + 1 1 ) . the threshold for significant changes was set to p = 0.05 whole brain cluster level fwe corrected with a cluster building threshold of p = 0.001 uncorrected on voxel level . as we expected an increase in training participants due to coactivation and no change in control subjects we verified the direction of changes with two post hoc paired sample t - tests for the trainee and the control group separately for each significant seed - to - voxel cluster . this step was chosen to assure that the significant results were not caused by between - subject variance . the other reason for this approach was the different sample size of the two groups . we report significant results due to three criteria : ( a ) significant time by group interaction , ( b ) significant increase within the training group , and ( c ) no significant decrease in the control group . for display purposes the cluster building threshold for the result - figure was set to 0.002 uncorrected on voxel level . as we could not investigate task related activity for the exercises and it is somewhat arbitrary and prone to bias to create a region of interest out of a single coordinate and an according sphere , we used the provided rois that are based on the brodmann areas according to the wfu pickatlas ( http://fmri.wfubmc.edu/software/pickatlas ) . we used all existing areas as rois , in order to get a complete picture of possible changes within the control group . some rois are not provided by the toolbox ( e.g. , thalamus , cerebellum , and fef ) ; here we created roi using the masks provided by wfu pickatlas of the according brain region . roi - to - roi analysis for the rs - fmri at the first time - point showed no differences between trainees and controls . the default network could be shown with the medial prefrontal cortex as seed in both groups and both time - points . the impact of the training was analyzed by a 2 2 anova ( group and time ) with age and sex as covariates of no interest . all seed regions with significant positive connectivity changes in trainees and no significant decreases in controls are listed in table 1 . the training involved a great amount of motor activity and the motor region was one of the hypothesized regions changing their connectivity strength . the left primary motor area ( ba4 , m1 ) showed increased connectivity to parts of the visual cortex ( figure 1(a ) , red ) and the somatosensory association area ( ba7 , figure 1(a ) , red ) . the connectivity strength of the whole premotor areas ( ba6 ) as seed to other cortical regions did not change . the primary sensorimotor cortices ( ba1 , ba2 , and ba3 ) as part of the sensorimotor network showed few changes in connectivity strength . only the spontaneous fluctuations of the left ba1 showed higher correlation to parts of the associative visual cortex ( ba19 , figure 1(a ) , cyan ) and parts of the parietal cortex ( ba7 , figure 1(a ) , cyan ) . the functional coupling within the visual network changed for the primary sensory areas ( ba17 ) of the right hemisphere . this roi increased in functional connectivity to the ventral acc ( ba24 , figure 1(b ) , violet ) and parts of the right premotor cortex ( ba6 , figure 1(c ) , violet ) . the connection to the left premotor cortex ( figure 1(c ) , blue ) was increased for the right secondary visual cortices ( ba18 ) . the connection increase to the ventral acc ( midcingulate ; figure 1(b ) , violet and blue ) of the visual areas was overlapping . different areas of the visual cortex show changes in functional coupling to the same premotor region and the cingulate cortex . the functional connectivity strength between the primary auditory cortex ( ba41 ) as part of the auditory network and the right cerebellum ( areas viii and ix ) increased with the training as well as the connection to the somatosensory association cortex ( ba7 , figure 1(d ) , green and red ) . the connections of the secondary auditory cortex ( ba42 ) to the parietal cortex ( ba7 , figure 1(d ) , violet and blue ) changed as well , partly overlapping with the increased connectivity of the primary auditory cortex . connections from the auditory to the visual cortex did also increase ( figures 1(d ) and 1(e ) , blue and green ) . the left fef but not the right fef showed connectivity changes to several clusters in the visual cortex ( figure 1(f ) , blue ) and the ventral acc ( figure 1(f ) , blue ) . the connectivity between the right dorsolateral prefrontal cortex and the right supramarginal gyrus ( figure 1(f ) , red ) increased . a more posterior part of the cingulate gyrus showed an increased functional connectivity to the right anterior frontal cortex and partially of the dorsolateral prefrontal cortex ( ba10 and ba9 , figure 1(f ) , violet ) . the characteristic regions of the default mode network , medial prefrontal cortex , lateral parietal cortex , posterior cingulate , and superior frontal cortex , showed no change in connectivity after the training . this study is the first to investigate the impact of an integrated multimodal training on functional brain connectivity . the training combines physical and cognitive exercises and does not aim at automating but focuses on novelty . for this purpose we compared the changes of the resting - state intrinsic functional connectivity of a group of subjects attending the first standard training course of life kinetik with the rs - fmri changes of a control sample . the training offers a great variety of exercises and can easily be adapted to clinical populations . we found a considerable amount of changes in resting - state functional connectivity in the training group . the connections within the default mode network , the most prominent resting - state network , did not change . the changes in functional connectivity mirror the activation during the training and some increases in correlation occur in regions that are known to be a key region in cognitive deficits , ageing or mental illness . in particular the connections from the right motor and premotor cortex , responsible for the left part of the body , were expected to strengthen . the training involved a great amount of motor activity and all participants were right handed , thus being forced to coordinate their left hand , arm , and leg . an increase of the connectivity for the whole region was only visible for the left primary motor area . the connection to parts of the somatosensory association area changed as well as to the visual cortex . musicians , for example , who have a long history of motoric training showed increased resting - state connectivity in motor areas and multisensory cortices compared to a control group . this is in contrast to other studies that found a decrease in connectivity accompanied with cumulative performance increase after initial beginners increase of connectivity . as the life kinetik training focuses on the novelty of the exercises a decrease of connectivity was not expected . various seed regions in the visual cortex showed an increased connectivity with parts of the premotor cortex , almost overlapping ( see figure 1(c ) ) . a great number of the training tasks consisted of throwing and catching different and , in some cases , relative small objects . the most comparable task used in other studies was juggling training where an impact of training intensity was found . low intensity training resulted in increases in functional connectivity in the motor network , whereas the high intensity juggling training group showed decreased functional connectivity . the results suggest that different training regimes are associated with distinct patterns of brain change . our training on the other hand was much less intensive than the low intensity juggling training which consisted of 15 min per day and furthermore , as already mentioned , the cerebellum is mapped to the association areas of the cerebrum so that we expected some changes in connectivity , which we found but less pronounced than expected . in contrast to the analysis of buckner et al . who found that the primary sensory cortices were not represented in the cerebellum , we found a change in the correlation of time courses in the primary auditory cortex and the cerebellum . maybe this is an effect of the verbal prompts during the exercises , which indicated different movements . some subregions responsible for the retention of visual - motion information were shown to change their structure during motor training . not structural but functional changes occur in our sample of trainees in contrast to the control sample . the brain region with the most prominent connectivity changes was the secondary somatosensory association cortex ( ba7 ) . mainly the auditory cortices change their relation to parts of ba7 ( figure 1(d ) ) . these clusters are all overlapping irrespective of the according seed region . also distinct from these regions are the clusters changing the connections to the motor cortex ( figure 1(b ) , red ) and the sensorimotor cortex . this result seems to corroborate the findings of the diversification of the parietal cortex [ 52 , 53 ] . grasping and visuospatial tasks activated different parts of the parietal cortex , overlapping with the regions that showed changes during life kinetik training ( figures 1(a ) and 1(d ) ) . the functional connectivity between the left fef as seed region and clusters in the visual cortex and the ventral acc increased , but not with the dorsal attention network . the fef is responsible for eye movement and surely active during throwing and catching of objects . the change of connectivity between fef and visual cortex might be an indication for a combined activation of these regions due to the increased visual attention during the training . regions of the frontal cortex involved in working memory processes and error processing showed few connectivity increases . the acc showed , in contrast to our hypothesis , no changes in connectivity to other brain regions . the ventral part of the cingulate cortex showed increased connectivity to the anterior prefrontal cortex , to the fef , and to the visual cortex . the left dorsolateral prefrontal cortex ( ba9 ) on the other hand showed an increased connectivity to the right supramarginal gyrus ( ba40 ) . given the specific property of the training , the connectivity changes seem reasonable . the cue has to be translated to an action , in most cases movements or manipulation of objects . these regions were not distinct but overlapping thus indicating the importance of these regions for preparing the action and also for the object manipulation . the question is why the auditory areas predominantly showed increased functional connectivity to the somatosensory areas ( ba7 ) but not to the premotor area . one major effort for the trainees is to remember the according movement to the prompt . but this did not result in the expected changes in connectivity of the dorsolateral prefrontal cortex . the functional connection of the thalamus to the right inferior frontal gyrus and insula increased in trainees but also decreased in controls . the thalamus is a region with multiple connections [ 33 , 55 , 56 ] . the strength of the thalamocortical connection has been reported to predict the performance in motor learning , to change with age , and to be diminished in mci and ad [ 35 , 36 ] , and a disruption of the thalamus - cortex relation has severe implications on mental health [ 5860 ] . proper or optimal function of ba7 seems to be an indication for a cognitive reserve , preventing dementia symptoms . switching attention is an important part of the training ; regions that are activated in such a task are part of the parietal cortex as well as premotor areas and the dorsolateral pfc but the relation between these regions and especially the change in relation have not yet been investigated . with our study there is a lot of evidence for brain plasticity even in the adult and aging brain . basic research has shown that different aspects of the brain can be shaped by various types of training and tasks . resting - state connectivity seems to be relatively stable , but disturbed in psychiatric disorders , changing with age [ 65 , 66 ] , and changeable by activity . intrinsic connectivity is shown to be an indicator for efficiency and positively correlated with cognitive performance [ 68 , 69 ] and intelligence . the data on the direction of changes or alterations in terms of increase or decrease are inconsistent . for example , patients with major depression show an increased functional connectivity whereas schizophrenia seems to be accompanied by decreased functional connectivity . the functional connectivity of the motor cortex , for example , is increasing with age but different relations to performance were reported . one study interpreted the positive relation of connectivity and performance as a protection against decline . this contradicting consequence of connectivity increase with regard to performance demonstrates that an intervention leading to enhanced connectivity between brain areas might not necessarily help attenuate age related decline . the tasks and types of training used in different studies to investigate the change of brain networks are somewhat arbitrary , varying from perception tasks according to robot - hand movements to juggling or transcranial electrical or magnetic stimulation . most training concepts do not have the potential to serve as a training method or therapy approach for psychiatric patients or elderly individuals . exceptions are various types of motor training like juggling , video games , aerobic training , or the quadrato - motor training which are all aimed at perfecting the task without varying the task . our motivation was to look for a training that includes motor and cognitive exercises and has the potential to be stimulating for a patient population . test - retest reliability is a not yet completely resolved issue in fmri studies [ 7477 ] . few studies addressed this issue for rs - fmri but retest reliability was found to be robust [ 44 , 78 ] . we addressed this issue by including preprocessing steps that are known to reduce noise [ 44 , 79 , 80 ] . we carefully screened the control group for changes and reported only results with a significant post hoc test . a second limitation for our results is the whole sample size as well as the difference in the size of the trainee and control group . we tried to address this issue by verifying the occurrence of the default mode network that did not change in control subjects despite the small sample size . the subject group participating in the training showed an increase in resting - state functional connectivity but the impact on task performance is unknown and could not be monitored due to the nature of the training . a next step would be to find suitable motor and cognitive test tasks to quantify an improvement following life kinetik training . further investigations should include an active control group ( practicing either motor performance or cognitive tasks or training a limited number of tasks to perfection ) to show the benefit of the combined training compared to its isolated parts . furthermore , we will investigate the impact of the training on cognitive performance and memory , preferably in a group of impaired subjects . since it is not possible to measure the brain activity during the training , we only could assume which brain regions are activated . this study was planned as a pilot study to show that the training is able to change brain connectivity . we assume that our subjects show normal resting - state networks . in a patient group Output:	this study investigated the impact of life kinetik training on brain plasticity in terms of an increased functional connectivity during resting - state functional magnetic resonance imaging ( rs - fmri ) . the training is an integrated multimodal training that combines motor and cognitive aspects and challenges the brain by introducing new and unfamiliar coordinative tasks . twenty - one subjects completed at least 11 one - hour - per - week life kinetik training sessions in 13 weeks as well as before and after rs - fmri scans . additionally , 11 control subjects with 2 rs - fmri scans were included . the conn toolbox was used to conduct several seed - to - voxel analyses . we searched for functional connectivity increases between brain regions expected to be involved in the exercises . connections to brain regions representing parts of the default mode network , such as medial frontal cortex and posterior cingulate cortex , did not change . significant connectivity alterations occurred between the visual cortex and parts of the superior parietal area ( ba7 ) . premotor area and cingulate gyrus were also affected . we can conclude that the constant challenge of unfamiliar combinations of coordination tasks , combined with visual perception and working memory demands , seems to induce brain plasticity expressed in enhanced connectivity strength of brain regions due to coactivation .
PubmedSumm2092	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: in today 's world with high speed collision , number of patients with complex trauma injuries has increased . these patients present with unique challenges as several injuries may require contrasting physiological management . patients with traumatic brain injury ( tbi ) require maintenance of vital parameters within strict limits to prevent secondary brain insult . it is well - recognized that low mean arterial pressure ( map ) reduces cerebral perfusion resulting in poor outcome in patients with severe tbi . blunt aortic injury ( bai ) with pseudoaneurysm <3 cm have been managed conservatively in our center . they can be managed by maintaining permissive hypotension ( map of 40 - 50 mmhg or a systolic blood pressure [ sbp ] 80 ) and serial imaging . here , we report a case of tbi with bai ( pseudoaneurysm > 4 cm ) , where an urgent decompressive craniectomy was warranted . bai was managed conservatively both in intra - operative and postoperative period with permissive hypotension without causing neurological deficit in long - term . a 27-year - old male patient presented in our emergency department with a history of road traffic injury . on admission , his glasgow coma score ( gcs ) was e1v1m3 , heart rate ( hr ) was 85 beats / min and blood pressure ( bp ) was 135/82 mmhg . his computed tomography ( ct ) scan showed right temporoparietal subdural hematoma ( sdh ) with multiple underlying contusions [ figure 1 ] . he was immediately intubated with the help of injection propofol 60 mg intravenous ( iv ) and cricoid pressure , with manual in - line stabilization and cervical collar in situ . patient also had left sided chest trauma and an intercostal drain ( icd ) was inserted , that drained around 200 ml of blood . ct scan and angiography of the chest revealed a pseudoaneurysm of the aortic arch , proximal to the origin of subclavian artery , measuring 4 cm in diameter [ figures 2 and 3 ] . the low gcs caused the evacuation of sdh to take precedence over bai and an immediate evacuation was planned . non - contrast computed tomography head of the patient showing fronto - temporal subdural hematoma ( white arrow ) three - dimensional reconstruction of the computed tomography scan showing heart and the great vessels ( white arrow shows the aortic arch aneurysm ) computed tomography angiograph of the patient showing a small aneurysm proximal to the origin of the left subclavian artery ( white arrow ) the patient was taken to the operating room after obtaining high - risk consent . anesthesia was induced with propofol 80 mg and fentanyl 100 g and muscle relaxation achieved with vecuronium 4 mg iv . a central venous catheter ( central venous pressure [ cvp ] ) was placed in the right internal jugular vein and right radial artery was cannulated for invasive bp and serial arterial blood gas monitoring . anesthesia was maintained with isoflurane in a mixture of air and oxygen ( 2:1 ) along with intermittent boluses of vecuronium and fentanyl . iv fluid therapy was guided by cvp , urine output and blood loss , and hemodynamics were meticulously maintained . our aim was to maintain sbp around 100 and map of 60 mmhg to maintain cerebral blood flow and also to prevent the aortic aneurysm from rupture . surgery lasted for 4 h , at the end of which the patient was shifted to the intensive care unit ( icu ) for elective ventilation . controlled ventilation was established with a tidal volume of 400 ml , respiratory rate of 15 breaths / min , positive end - expiratory pressure of 6 cm h2o and a fio2 of 0.5 . he was sedated using continuous infusion of midazolam ( 2 mg / h ) and fentanyl ( 50 g / h ) . esmolol infusion ( 50 - 100 g / kg / min ) required for a period of 22 h on the first postoperative day to control unexplained tachycardia , following which the map and hr remained stable . intracranial pressure ( icp ) monitoring was carried out and a microdialysis catheter was also inserted to measure the lactate / pyruvate ( l / p ) ratio on day 2 of icu . both icp and the l / p ratio remained within acceptable limits throughout the icu stay ( < 20 cmh2o and < 25 , respectively ) . mean cerebral perfusion pressure was maintained around 70 mmhg . the icd was removed on day 5 and icp monitoring and microdialysis catheter were removed on day 7 of icu . the patient was tracheostomized and discharged with a gcs of e4vtm6 and a resolving aneurysm on postoperative day 10 . with extensive modernization and an exponential growth in motor vehicles , polytrauma has become increasingly common . polytrauma patients form a special cohort since the management of one injury may interfere with the optimal treatment of the other , creating a dilemma for the treating physician . such a situation arises , as in our patient , where tbi and bai coexist , which poses a challenge to the anesthesiologist during surgical management of either of the injuries . secondary injury in tbi is described as the consequence of physiological insults such as ischemia , reperfusion and hypoxia to areas of at risk brain in the period after the initial injury . it has been widely studied and reported that hypotension and hypoxia have deleterious effects on the outcomes of tbi patients . the impact study suggested that hypotension and hypoxia were significantly associated with unfavorable 6-month outcome . moreover , the duration of intra - operative hypotension was also inversely associated with functional outcome . bai have a higher mortality rate with the majority of the patients succumbing at the site of trauma . with improvement in pre hospital care the number of patients with bai reaching hospital has increased . the patients may be managed by early surgery or nonoperative therapy depending on the severity of injury . the principle of nonoperative management revolves around differentiating bai into minimal aortic injury ( mai ) and severe aortic injury ( sai ) . patients with mai ( usually lesions <3 cm ) have a low risk of progression and aortic rupture and are better managed conservatively with aggressive negative inotropic and antihypertensive therapy , routine serial imaging and close clinical observation . sai mandates surgical or endovascular repair in most of the cases if the risk - benefit ratio is favorable . surgeons at our institution follow similar guidelines for management of aortic aneurysms . in our patient the management of tbi took precedence over bai , therefore we decided to manage the bai conservatively and operate the sdh . the pseudoaneurysm , although large ( > 4 cm ) , was managed conservatively in our patient . according to the recent retrospective analysis of bai by caffarelli et al . , aortic injuries , especially pseudoaneurysms are different from nontraumatic atherosclerotic aneurysms , because in the majority of the trauma patients , the underlying pathology does not exist . this stanford philosophy implies that nonoperative management may be more successful in traumatic aortic aneurysms as compared to pathological ones . however , application of permissive hypotension in patients with neurological involvement must always be carried out in the presence of monitoring of cerebral function , such as cerebral oximetry . at our institution , intra - operative cerebral oximetry is not used , but the use of icp monitors and cerebral microdialysis catheters in the postoperative period are frequent . our patient , however , presented with the unique situation of tbi with bai ( pseudoaneurysm measuring 4 cm ) . although according to the aneurysmal size the patient required surgery , the surgical management of tbi was more urgent . the management of the intra - operative period required meticulous monitoring of bp so that we could maintain cerebral perfusion without rupturing pseudoaneurysm . patients with bai with large size pseudoaneurysm may be managed intra - operatively with permissive hypotension in the presence of appropriate monitoring of cerebral function such as cerebral oximetry . however , more cases are required to be studied for validation of this hypothesis and formulation of guidelines . Output:	with an ever - increasing incidence of high impact collisions , polytrauma is becoming increasingly common . patients with traumatic brain injury ( tbi ) may require urgent surgical intervention along with maintenance of an adequate mean arterial pressure ( map ) to maintain cerebral perfusion . on the other hand , patients who sustain blunt aortic injuries ( bai ) have a high mortality rate , many of them succumbing to their injury at the site of trauma . surgery has been the mainstay of the management strategy for the remaining survivors . however , in recent years , the paradigm has shifted from early operative management to conservative treatment with aggressive blood pressure and heart rate control , serial imaging , and close clinical monitoring . when tbi and bai coexist in a patient , it becomes crucial to maintain the map within a narrow range to prevent secondary insult to the brain as well as to prevent aortic rupture . we present the management of a case of tbi with traumatic aortic pseudoaneurysm , which required stringent monitoring and maintenance of hemodynamics during decompressive craniectomy .
PubmedSumm2093	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: dentine hypersensitivity ( dh ) is an abnormal response of the exposed vital dentine to thermal , chemical , or tactile stimuli . the prevalence of dh has been reported ranging from 4 to 57% in many studies in the literature , depending on the population samples studied [ 1 , 2 ] . in patients affected by periodontitis , however , dh prevalence is likely to increase in next years since more adults keep their teeth into later life . this condition may affect patients at any age , and both genders are equally affected [ 4 , 5 ] . although different theories have been proposed for the mechanism involved in dh etiology , recent studies gave support to brannstrom 's hydrodynamic theory , according to this a stimulus applied to open tubules dentin increases the flow of dentinal tubular fluid , with mechanical deformation of the nerves located into the inner ends of the tubules or in the outer layers of the pulp . type a delta fibers are supposed to be responsible for dentinal sensitivity being probably activated by the hydrodynamic process . the most common factors involved in dh are abrasion , caused by inadequate intensity tooth brushing ; abfraction , caused by teeth flexion due to abnormal occlusal forces ; parafunctions or occlusal disequilibrium ; erosion , secondary to the presence of acids in the oral cavity , as in bulimia nervosa or gastroesophageal reflux ; anatomic predisposition due to structural deficiency of the enamel - cement junction ; cavity preparations in vital teeth that expose dentine or badly controlled dentinal acid conditioning [ 810 ] . orchardson and gillam showed that dh affects above all the vestibule - cervical area of teeth . cervical dh has probably a multifactorial etiology , and more than a cause is related to this painful manifestation . therefore , several treatments must be associated to reduce dh to satisfactory levels . according to garone - filho the abfraction , caused by occlusal overload , furthermore , according to pashley , there are two types of dentinal permeability : intratubular , into the dentinal tubules , and intertubular , between the tubules in dentinal matrix . the sensitive dentine is permeable through its thickness ; any treatment that reduces dentinal permeability must reduce dentinal sensitivity . in fact , occlusion of the exposed dentinal tubules may decrease dentinal sensitivity level [ 914 ] . however , the dh sometimes persists despite of the effective sealing of the tubules , so indicating that further mechanisms are involved in nerves activation instead of or in addition to the hydrodynamic mechanism . many authors suggested the hypothesis concerning the release of neuropeptides from the activated nervous terminations and , subsequently , the induction of a neurogenic inflammation . this hypothesis should signify that the symptoms of dh could become self - sustainable up to a certain point [ 8 , 14 ] . another great problem related to dh is its evaluation , since pain is a highly subjective sensation . nevertheless , it is possible to classify the dh according to matsumoto 's criteria . in this classification , three degrees of dh are recognized : grade 1 mild discomfort / pain , grade 2 moderate pain , and grade 3 characterized by intense and unbearable pain . through literature examination emerges that there is no therapy that can always reduce pain at satisfactory levels , even with the combination of different protocols . according to landry and voyer , there is not an ideal desensitizing agent but any kind of treatment for dh should be effective from the first application and must satisfy these parameters established by grossman since 1934 : ( 1 ) not irritating pulp , nor causing pain , ( 2 ) easy application , ( 3 ) long - lasting effect , ( 4 ) not discoloring or staining teeth , ( 5 ) not irritating soft tissues or periodontal ligament , ( 6 ) low cost . the occlusion of dentinal tubules leads to the reduction of dentinal permeability so decreasing the degree of dh . according to the hydrodynamic theory , the effectiveness of dentine desensitization agents is directly related to their capacity of promoting the sealing of the dentinal canaliculi . conventional therapies for dh are based on the local application of desensitizing agents , either professionally or at home . the most frequently used agents can be classified as protein precipitants , tubule - occluding agents [ 19 , 20 ] , tubule sealants . the sodium fluoride gel ( naf ) , which belongs to the tubule - occluding agents family , is the most commonly used agent [ 4 , 2225 ] . its mechanism relies on the mechanical occlusion that is accomplished by precipitation of insoluble calcium fluoride crystals within the tubules without adhesion . for this reason , it can not resist to the stresses of the oral environment and its action decreases with time [ 4 , 23 ] . in the last fifteen years , the introduction of lasers gave further possibilities to dh therapy [ 22 , 2629 ] . laser photobiomodulating action in dental pulp was reported by many authors as in villa et al . , with histological studies of dental pulp of mice , after laser irradiation in teeth with exposed dentine . in this study , the authors registered a large quantity of tertiary dentine production in lased teeth , that caused the physiological obliteration of tubules , while the nonirradiated control showed intense inflammatory process that , in some cases , evolved into necrosis . focusing on the role of laser in dh therapy , it is possible to show that its action is twofold . by one side , the low - level power lasers [ 14 , 31 ] , also called soft lasers , act directly on nerve transmission , with a depolarization process that prevents the diffusion of pain to snc ; however , their effectiveness seems poorer in higher degrees of dh . by the other side , high - power lasers such as : diode 980 nm and 808 nm , ktp 532 nm , nd : yag 1064 nm , co2 10600 nm , er , cr : ysgg 2780 nm , and er : yag 2940 nm act on dh provoking a melting effect with crystallization of dentine inorganic component and the coagulation of fluids contained into the dentinal tubules . among these high power devices , diode lasers are the most studied and the ones that gave the best results in several clinical protocols even in high - grade dh cases . the aim of this study is to assess the efficacy of a diode gaalas laser alone and in combination with topical sodium fluoride gel ( naf ) in the treatment of dh in order to evaluate the possibilities of this device in the management of this painful condition . the study was conducted on 10 patients ( 8 females and 2 males ; aged from 25 to 60 years ) and in a total of 115 teeth with dh assessed by mean of both air ( figure 1 ) and tactile ( figure 2 ) stimuli measured by the numeric rating scale ( nrs ) . the inclusion criteria for patient enrollment were based on : the absence of local ( e.g. , cavities , fractures ) and/or systemic pathologies , on the absence of contraindications to the proposed therapies ( e.g. , allergies to desensitizing agents ) and on the presence of teeth with dh evaluated by pain response to both air and tactile stimuli that were registered by nrs scale ( from 0 to 10 , where 0 meant the absence of pain and 10 represented an unbearable pain and discomfort felt by the patients in their life ) ; at last no desensitizing therapy had to be previously performed , nor analgesic drugs had to be recently assumed . before any treatment , all the patients received a hygiene professional program with oral hygiene instructions and the teeth vitality of all sites was assessed . for each patient , the sensitive sites were randomly divided into three groups : group 1 ( g1 ) ( 34 teeth ) treated with 1.25% naf applied for 60 seconds on tooth surface ( figure 3 ) ; group 2 ( g2 ) ( 33 teeth ) lased by a gaalas laser ( doctorsmile , lambda s.p.a . , brindole ( vi ) , italy , 980 nm ) with these parameters : 0.5 w in pw ( t on 100 ms and t off 100 ms ) and fluence of 62.2 j / cm in no contact mode and using a fiber of 320-micron diameter . each site received 3 applications of 1 minute each ( figure 4 ) once a week for three weeks;group 3 ( g3 ) ( 48 teeth ) treated using both naf gel and diode laser at the same parameters of g2 . the naf gel was left on tooth surface for 60 seconds before the irradiation ; in this way , the laser system could favor the permanence of desensitizer for a longer time than when it was used alone ( figure 5 ) . group 1 ( g1 ) ( 34 teeth ) treated with 1.25% naf applied for 60 seconds on tooth surface ( figure 3 ) ; group 2 ( g2 ) ( 33 teeth ) lased by a gaalas laser ( doctorsmile , lambda s.p.a . , brindole ( vi ) , italy , 980 nm ) with these parameters : 0.5 w in pw ( t on 100 ms and t off 100 ms ) and fluence of 62.2 j / cm in no contact mode and using a fiber of 320-micron diameter . each site received 3 applications of 1 minute each ( figure 4 ) once a week for three weeks ; group 3 ( g3 ) ( 48 teeth ) treated using both naf gel and diode laser at the same parameters of g2 . the naf gel was left on tooth surface for 60 seconds before the irradiation ; in this way , the laser system could favor the permanence of desensitizer for a longer time than when it was used alone ( figure 5 ) . patients ' response to cold air blast was assessed by a short blast of 1-second duration at a distance of 0.5 cm for each tooth . both air and tactile stimuli evaluations were performed before and after every treatment session , for a total of 3 treatment sessions at a distance of about one week each other . a reduction of dh occurred during the treatment sessions , and the positive values were maintained after 1 month ( tables 1 , 2 , and 3 ) . comparing the three regimens , a higher decrease of dh was registered in g3 , followed by g2 and g1 , respectively , whose results seem to be almost superimposable . the nrs reduction percentage was valued for each group between the first pretreatment and the third posttreatment session ( immediate/ ) . the values were divided depending on the kind of stimulation . for the air stimulus , the reduction percentage was , respectively , 10.19% ( i ) for g1 ; 22.35% ( i ) for g2 ; 25.04% ( i ) for g3 . furthermore , the tactile stimulus took down : 4.13% ( i ) for g1 ; 6.77% ( i ) for g2 ; 9.96% ( i ) for g3 . regarding to the statistical analyses , the data relating to the probe test ( figure 6 ) were subjected to the kruskal - wallis 's test which demonstrated the reliability of the study ( p < 0.0001 ) . the comparative dunn 's test showed a statistically significant difference in g3 ( p < 0.001 ) , and in g1 ( p < 0.01 ) . in g2 the obtained improvement the results obtained with the cold blast ( figure 7 ) air were always analyzed through the kruskal - wallis 's test which demonstrated the reliability of the study ( p < 0.0001 ) and the comparative dunn 's test showed a statistically significant difference in each treated group ( p < 0.001 ) . the improvement obtained in the samples of the probe test was not statistically significative ( p > 0.05 ) instead of the samples of the cold blast air in which the improvements from the first treatment to the last one were superimposable . through literature examination , it is clarified that the ideal treatment for dh does not exist , even in case of combination of different protocols . conventional therapies for the treatment of dh comprehend the topical use of desensitizing agents , either professionally or at home such as protein precipitants , tubule - occluding agents [ 19 , 20 ] , tubule sealants , and , recently , lasers [ 22 , 2629 ] . several studies [ 3234 ] describe a synergistic action of lasers in association with desensitizing agents . in fact , the laser system can favor the permanence of the desensitizer for longer time than when they are used alone . for this reason , if laser device is used in addition to a conventional desensitizing agent , the latter remains above the tooth surface for 60 seconds before the irradiation . focusing on the effectiveness of the sole diode laser , this was investigated by several authors . matsumoto et al . showed an 85% improvement in teeth treated with laser ; aun et al . reported success in laser - irradiated teeth in 98% of their cases ; yamaguchi et al . noticed an effective improvement index of 60% in the group treated with laser compared to the 22.2% of the control nonlased group ; kumazaki et al . showed an improvement of 69.2% in the group treated with laser compared to 20% in the placebo group . , in a double - blind study , found significant values in the laser - treated group . in fact , sensitivity to thermal stimuli was reduced by 67% , whereas the placebo group had a reduction of 17% , sensitivity to tactile stimuli was reduced by 65% , while the placebo group showed a reduction of 21% . another study carried out by brugnera et al . showed the immediate analgesic effect using a diode laser . in this study , significant improvements in pain and discomfort were always registered after the session treatments ( i ) even if in no case the percentages of pain reduction arrived to the high values registered in the literature . as a first laser showed the best immediate results alone and in combination with gel , since the percentages of pain reduction in g2 and g3 were more than twice than g1 values . in our sample , the best results were obtained by the association of laser and naf gel therapy ( g3 ) . it is probable that the better performance of combined treatment was due to the higher naf gel adhesion to the dentinal tubules when combined with laser energy . in the laser group , g2 , the immediate pain reduction was very high especially at air stimulation ( 22.35% i ) ; in the same group , the improvement at tactile stimulation was poorer after the treatment ( 6.77% i ) . the lower reduction values were registered in the sole gel group , g1 , by both stimulations , in the immediate period . even in consideration of the short sample analyzed , it is possible to speculate that the laser - induced superficial melting permits to keep longer the tubules occlusion by naf gel emphasizing the reduction of dh - related pain . according to these results , the gaalas laser showed a very high capability to improve immediately the dh - related pain , both alone and even better in combination with naf gel . on the other hand , the sole gel results , these results have to be confirmed by greater samples of patients and by longer follow - up periods ( e.g. , 3 and 6 months ) to confirm or not the long - lasting action of the combined laser and gel therapy . Output:	introduction . dentine hypersensitivity ( dh ) is characterized by pain after stimuli that usually provoke no symptoms . this study compared the effectiveness of gaalas diode laser alone and with topical sodium fluoride gel ( naf ) . materials and methods . the study was conducted on 10 patients ( 8 f/2 m , age 2560 ) and 115 teeth with dh assessed by air and tactile stimuli measured by numeric rating scale ( nrs ) . teeth were randomly divided into g1 ( 34 teeth ) treated by 1.25% naf ; g2 ( 33 teeth ) lased at 0.5 w pw ( t on 100 m and t off 100 ms ) , fluence 62.2 j / cm2 in defocused mode with a 320 fiber . each tooth received three 1 applications ; g3 ( 48 teeth ) received naf gel plus laser at same g2 parameters . nrs was checked at each control . results . significant pain reduction was showed . the nrs reduction percentages were calculated , and there was a concrete decrease of dh above all in g3 than g2 and g1 . conclusion . diode laser is a useful device for dh treatment if used alone and mainly if used with naf gel .
PubmedSumm2094	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: the superfamily of cytochrome p450 enzymes ( cyps ) is one of the most versatile biocatalyst systems in nature . cyps are capable of catalyzing more than 20 distinct types of reactions including regio- and stereoselective oxidation of unactivated c h bonds ( hydroxylation and epoxidation ) , dealkylation , c c bond cleavage , aromatic coupling , and others . recently , more novel activities such as decarboxylation , nitration , farnesene synthase activity , and carbene transfer have been reported , reflecting the ability to further diversify the function of these versatile enzymes . catalytic activities of the vast majority of cyps require one or more redox partner proteins to sequentially deliver two electrons from nad(p)h to the heme iron reactive center for dioxygen activation . classically , there are two major redox partner systems , including ( 1 ) a two - component system for most bacterial and mitochondrial cyps comprising a small iron sulfur redoxin and an fad - containing redoxin reductase and ( 2 ) a single fad / fmn - containing cyp reductase that serves in eukaryotic microsomal p450s as a separate partner protein or in a small number of bacterial cyps ( e.g. , p450bm3 ) as a fused functional domain . interestingly , nonclassical redox systems have been discovered continuously , among which the fmn / fe2s2-containing reductase domain termed rhfred that is naturally fused with the p450 enzyme from rhodococcus sp . ncimb 9784 has been extensively studied in our laboratories . for practical purposes due to the difficulty of obtaining native partners , one or more surrogate redox partners , acting either in isolation or as artificially fused protein complexes , the choice of surrogate partners or their mode of action is not necessarily expected to affect the type and selectivity of reactions catalyzed by p450s . although alternative redox partners may influence catalytic efficiency and/or product distribution , the chemical identity of products themselves is apparently determined by the p450 enzyme . our previous experience constructing and characterizing various self - sufficient biosynthetic p450 enzymes ( whose activity is independent of isolated redox partners ) fused with the rhfred reductase domain also supports this postulate . notably , previous work revealed that cytochrome b5 is able to modulate the bifunctional role of human cyp17a1 as a 17-hydroxylase or a 17,20-lyase in steroid biosynthesis . this well - studied example demonstrates that the catalytic type of a p450 enzyme can be altered by a third - party protein effector via alternative interactions among the p450 enzyme , p450 reductase , effector protein , and distinct substrates . however , there have been no reports for a redoxin protein or a p450 reductase acting as the specific effector by itself to endow the serving p450 enzyme with new catalytic activity . in this work , novel demethylated mycinamicin products were generated by p450 mycg supported by a stand - alone form of the rhfred reductase domain , as opposed to rhfred fused to mycg , thus challenging the generally accepted postulate . further engineering of rhfred led to didemethylated mycinamicin macrolides , demonstrating a novel route for structural diversification of natural products . protein interactions with the redox partner may serve as a modulator of p450 function by dramatically affecting the substrate binding mode in the p450 active site . mycg is the multifunctional p450 monooxygenase involved in the biosynthetic pathway of 16-membered ring macrolide mycinamicins in the rare actinomycete micromonospora griseorubida . physiologically , it catalyzes the c14 hydroxylation , and the c12=c13 epoxidation of mycinamicin iv ( m - iv ) leading to mycinamicin v ( m - v ) and mycinamicin i ( m - i ) , respectively . m - v is subsequently epoxidized by mycg , giving rise to the major final product mycinamicin ii ( m - ii ) , while m - i can not be further modified by mycg , thus representing the other terminal product in the pathway ( figure 1a ) . all diglycosylated mycinamicins display strong antibiotic activity against gram - positive bacteria and mycoplasma , and m - ii has been developed into a veterinary anti - infective agent . the multifunctional activities of mycg were previously reconstituted in vitro by using the commercial surrogate redox partners spinach ferredoxin ( fdx ) and spinach ferredoxin - nadp reductase ( fdr ) , since the native redox system driving catalytic activity of mycg remains unknown . motivated by our recent work and that of others in enhancing catalytic activity by fusing the rhfred reductase domain to the c - terminus of various biosynthetic p450s , we constructed the mycg - rhfred fusion protein . similar to mycg supported by separate spinach fdx / fdr , this single - component self - sufficient p450 system driven by nadph achieved all physiological reactions known for mycg ( figure 1b ) . oxidation and demethylation of m - iv catalyzed by mycg using different redox systems ( 2 h reactions ) . ( b ) left panel : lc traces at 280 nm of different reaction extracts . the redox systems used are shown at the left side of the corresponding traces . m - v and m - ii were coeluted , and dme - m - iv and m - i were coeluted . since m - i and m - ii lacking of the diene moiety only have weak absorbance at 280 nm , the formation of products appears not to be proportional to substrate consumption . right panel : ms / ms analysis of each mycinamicin compound ( see figure s1 for explanations of the secondary mass spectra ) . ( c ) comparison of the h nmr spectra ( see figures s2 and s3 for full spectra ) of m - iv and dme - m - iv . the chemical shift and integrated peak area of the n - monomethyl group in dme - m - iv are apparently different from those of the n - dimethyl group in m - iv . to investigate if the free form of the rhfred domain can support the catalytic function of mycg , the ability of this two - component system to convert mycinamicin iv ( m - iv ) was tested . surprisingly , in addition to the expected oxidative products , m - i , m - ii , and m - v ( coeluted with m - ii ) , a new product formed with 18.5% isolated yield ( table 1 ) with the same retention time as m - i was observed in lc - ms analysis of the reaction mixture ( figure 1b ) . because of its strong absorbance at 280 nm , resembling that of m - iv and m - v , but distinct from m - i , we assumed that the chemical structure of the new compound likely retained an intact diene moiety . indeed , its m / z value of 682.3 is 14 amu less than that of protonated m - iv , suggesting a demethylated product , which was also supported by high - resolution mass spectrometry ( [ m + h ] : obsd , 682.4161 ; calcd , 682.4160 ) . furthermore , the ms / ms spectrum of the demethylated m - iv n - demethylated mycinamicin iv ( dme - m - iv ) showed two fragmentation ions of 144.2 and 102.2 , consistent with a demethylated desosamine moiety , while the corresponding fragmentation ions for intact desosamine are 158.2 and 116.1 ( figures 1b and s1 ) . detection of formaldehyde in the reaction by purpald reagent provides additional evidence for oxidative demethylation ( see supporting information ) . to confirm the site of demethylation , the h nmr of enzymatically prepared dme - m - iv was analyzed using m - iv as reference ( figures 1c , s2 and s3 ) . as expected , the 2.35 ppm singlet peak of m - iv that corresponds to the n - dimethyl group disappeared in the spectrum of dme - m - iv . instead , a new singlet peak was observed with half the integrated area at 2.72 ppm , indicating the nhme group resulted from the mycg - catalyzed demethylation . numbers of relative auc280 nm ( areas under curve at 280 nm ) were calculated from the integrated area of certain peaks on hplc traces coupling efficiencies were calculated as the percentage of nadph used for product formation over the total nadph consumption this observation is intriguing because a heretofore unknown dme - m - iv product was obtained following separation of the previously fused protein domains , mycg and rhfred . next , we measured the oxidation and demethylation activities of mycg under a variety of molar ratios ( 1:1 , 1:10 , and 1:100 ) between mycg and rhfred . in all cases , both oxidative and demethylated products were detected ( figure s4 ) , indicating the ability of mycg to catalyze demethylation is not the consequence of a stoichiometric effect of rhfred . a time course of the reaction ( figure s5 ) showed proportional accumulation of oxidative and demethylated mycinamicin products . sequence analysis shows that rhfred consists of two functional domains , the fmn domain and the fe2s2 domain . to analyze the impact of each domain on demethylation by mycg , the fe2s2 domain of rhfred , which is presumably involved in direct interaction with the p450 enzyme , was exchanged for the spinach fdx , yielding a hybrid rhodococcus - spinach reductase , rhfred - fdx . when partnered with this hybrid reductase , the mycg - catalyzed reaction produced even greater amounts of dme - m - iv ( 24.0% isolated yield , see table 1 ) , while the oxidized products m - i , m - v , and m - ii were also detectable ( figure 1b ) . this is in contrast to the mycg - rhfred - fdx fusion system which , similarly to mycg - rhfred , generated only oxidized products , albeit in lower quantities . these results suggest that mycg - mediated demethylation is mainly determined by the mode of interaction between mycg and its redox partner rather than by the redox partner itself . quantitatively , by fitting the substrate consumption data into the one phase exponential decay curve , the rate constants ( k ) of the four studied mycg catalytic systems were determined ( table 1 and figure 2a ) . in comparison , isolated rhfred and the m - iv oxidation mediated by mycg - rhfred and mycg - rhfred - fdx demonstrated the apparent reaction velocity at 500 m of m - iv being 26.0 6.0 m min and 8.5 2.0 m min ( k [ s ] ) , respectively . the steady - state michaelis menten kinetics of mycg could not be ascertained due to multiple reactions occurring simultaneously . ( a ) enzymatic consumption of m - iv ( calculated from hplc areas under curve , auc ) fit into the one phase exponential decay curve . ( b ) the effects of catalase ( 20 u ) , superoxide dismutase ( 2 u ) , ascorbate ( 10 mm ) , and the combination of these three ros scavengers on the activity of mycg / rhfred against m - iv . solid bar is the ratio of demethylated product / oxidized products relative to that of the control reaction without addition of any scavengers , and open bar is the overall conversion percentage calculated based on the substrate m - iv consumption . it is well - known that the use of an unnatural redox partner in a cyp catalyzed reaction likely uncouples the electron generation upon nad(p)h consumption and p450 product formation . the uncoupling process generates reactive oxygen species ( ros ) such as superoxide anions ( o2 ) and h2o2 , which could direct the p450 catalytic route into the peroxide shunt pathway . to examine whether the demethylation of mycinamicins is a consequence of the peroxygenase activity by undergoing this shunt pathway , we first determined the nadph coupling efficiency of the four catalytic systems using m - iv as substrate . as shown in table 1 , the coupling efficiencies for p450-reductase fusions ( 18.7% for mycg - rhfred and 21.3% for mycg - rhfred - fdx ) were lower than those of corresponding reactions using separated reductases ( 29.4% for mycg + rhfred and 24.2% for mycg + rhfred - fdx ) . these data indicate that separation of reductase and p450 domains did not result in greater uncoupling in these p450 catalytic systems . moreover , catalase , superoxide dismutase ( sod ) , ascorbic acid , and the combination of these three ros scavengers were added into the mycg / rhfred / m - iv reaction to remove ros possibly generated from the uncoupling process . although these ros scavengers lowered the m - iv conversion ( except for ascorbate ) as well as the ratio of demethylated product to oxidized products ( except for sod ) in the majority of reactions ( figure 2b ) , these effects were not dose dependent , and the n - demethylation activity remained even at the highest concentration of catalase , sod , and ascorbate ( figure s6 ) . these results , together with the small change of coupling efficiency upon reductase domain separation , suggest that dme - m - iv is unlikely to be a product of the peroxide shunt pathway . however , the lowered demethylated product / oxidized product ratio implies superoxide species might still contribute to some of the observed demethylation . this could be supported by detection of a low level of h2o2 in mycg reactions ( figure s7 ) and the fact that addition of ascorbic acid improved the overall conversion of m - iv ( figure 2b ) since this chemical scavenger is capable of protecting the enzyme and substrate / products from radical damage by superoxides . next , we comparatively tested the activity of mycg , driven by each of the four different redox systems , against m - iv analogues m - i , ii , iii , and v , isolated from fermentation cultures of wild - type or mutant micromonospora griseorubida . the two fused systems displayed the typical physiological set of the hydroxylation and/or epoxidation products ( figure 3 ) . however , the reactions driven by free redox partners gave rise to more diversity of demethylated products . specifically , both rhfred and rhfred - fdx efficiently supported the demethylation of m - i and m - iii by mycg , generating the new products dme - m - i and dme - m - iii , respectively . m - ii and m - v were demethylated by mycg at a low level driven by rhfred - fdx , but not by rhfred . of particular interest , mycg partnered by the stand - alone form of rhfred - fdx was able to further remove the second n - methyl group from monodemethylated dme - m - i and dme - m - iii , leading to the unique double - demethylated product d2me - m - i and d2me - m - iii ( figure 3 ) , respectively . previously , in both the in vivo(30,45 ) and in vitro reactions that employed the surrogate spinach fdx / fdr system , m - i and m - ii have been shown to be the two end products of the mycinamicin biosynthetic pathway , and m - iii bearing the monomethoxy sugar javose is oxidized by mycg at a very low level . here , the change of the redox partners and their interaction mode has led to seven novel demethylated products ( figures 1 , 3 , and s8 ) , demonstrating a new route for structural diversification of natural products . lc - ms analysis of the mycg reactions using m - i , m - ii , m - iii , and m - v as substrates . the lc traces of m - i and m - ii were recorded at 240 nm . the lc traces of m - iii and m - v were recorded at 280 nm . in the m - v reactions catalyzed by two fusion mycg enzymes , the accumulation of m - ii ( seen at 240 nm ) that is coeluted with the unreacted m - v is shown in insets . the selected ms / ms results are displayed in insets , whose colors are consistent with those of the corresponding lc peaks . the asterisked numbers indicate the mass for the secondary ion fragment of double - demethylated mycinamicins ( figure s1 ) . the structure of m - ix ( the hydroxylated m - iii labeled by the diamond symbol ) is shown in figure s8 . in this study we show for the first time the ability of the multifunctional p450 monooxygenase mycg to catalyze n - demethylation of various mycinamicin natural product molecules . this novel functionality of mycg occurs when the monooxygenase is partnered with either the free rhodococcus reductase domain , rhfred , or the rhodococcus - spinach hybrid , rhfred - fdx . in contrast , no similar demethylation activity was observed with the corresponding fused redox partner systems . the minor differences in the nadph coupling efficiency upon p450-reductase domain separation ( table 1 ) and dose - independent effects of catalase , sod , and the chemical radical scavenger ascorbate on both mycg activity and the demethylation / oxidation product distribution ( figure 2b ) strongly suggest that the unnatural demethylation activity is primarily due to heme - iron - oxo catalysis , as opposed to the peroxide shunt pathway . thus , this finding highlights a broader role of redox partner protein protein interactions in modulating catalytic activity of the p450 enzyme . it is well - known that alternative or surrogate redox partner protein(s ) might not provide the optimal supporting activity toward mismatched p450 enzymes and could influence product distribution . the modulating effect of a third party protein , cytochrome b5 , on the hydroxylase / lyase bifunction of cyp17a1 supported by cytochrome p450 reductase has also been demonstrated . however , to the best of our knowledge , the entirely new catalytic activity of a p450 enzyme associated with an alternative redox partner is unprecedented . the novel catalytic activity demonstrated by mycg likely stems from the poor accessibility of the substrate binding site , as evidenced by the x - ray structure analysis of the mycg - substrate complexes . thus , the bulky and conformationally restrained mycinamicin substrates were largely unable to penetrate the l - shaped binding site to adopt a catalytically productive binding mode . nmr modeling using paramagnetic restraints suggests that displacement of a number of the protein secondary structure elements takes place in order to accommodate mycinamicin substrates in an orientation conducive to the observed oxidation pattern . by analogy to p450cam and the human p450 3a4 , reorganization of the mycg active site may be driven by binding of the redox partner . as has been recently demonstrated by elegant work from the poulos laboratory , binding of the endogenous redox partner putidaredoxin stabilizes the open conformation of p450cam , facilitating substrate access to the active site . consistent with the structurally resolved mycg - substrate complexes , mycinamicins exhibiting pseudo two - fold symmetry enter the active site in one of the two possible orientations : mycinose - in - desosamine - out ( figure 4a ) or desosamine - in - mycinose - out ( figure 4b ) . the desosamine - in - mycinose - out entry results in the catalytically nonproductive binding mode represented by the m - iii costructure ( pdb i d code 2yca ) . remarkably , the mycinose - in - desosamine - out entry in the absence of the redox partner resulted in both the m - iv and m - v substrates being stalled in the intermediate nonproductive binding pose in all the structurally resolved complexes . if substrate relocation to the catalytically competent mode is indeed initiated by specific interactions with the endogenous redox partner , the alternative surrogates may to some extent mimic this allosteric effect , either facilitating or hampering substrate progression to the catalytic site . tethered to mycg , both rhfred and rhfred - fdx afforded physiological product profiles , with the qualification that the product yield was low for mycg - rhfred - fdx . alternatively , when used as stand - alone redox partners , both the rhfred and rhfred - fdx domains produced demethylated ( rhfred ) or didemethylated ( rhfred - fdx ) products of not only the native mycg substrates m - iv and m - v but also the m - iii precursor , an otherwise poor substrate for mycg oxidation , which was efficiently demethylated ( figure 3 ) . ( a ) a putative binding pose of m - iv leading to physiological c14 hydroxylation or / and c12c13 epoxidation , which might be derived from the noncatalytic mycinose - in - desosamine - out entry pose as observed in the 2y98 ( pdb i d ) structure . ( b ) a putative desosamine - in - mycinose / javose - out binding pose yielding n - demethylation is likely to be induced or stabilized by protein the noncatalytic conformation observed in the 2yca structure may be derived from this pose . slice through the mycg binding site shows m - iv or m - iii in virtually orthogonal orientations experimentally observed in the crystal structures . protein surface is colored by hydrophobicity , hydrophobic areas are in orange , and hydrophilic areas in blue . to explain this phenomenon we speculate that while the electron transfer between mycg and the surrogate redox partners is retained , the allosteric effect facilitating substrate progression to the normal catalytically competent binding mode ( mycinose - in - desosamine - out ) is likely diminished , if not entirely lost . this might be due to intrinsically low affinity between the surrogate partners and mycg , particularly if concentration dependence in binary complex formation becomes a rate - limiting issue in the bimolecular reaction . also , loss of the positioning effect of the linker following domain separation may be a factor in changing the interaction mode of free reductase and p450 domains . we conclude that in the absence of the allosteric influence of the redox partner , mycinamicins are more inclined to be stalled in the desosamine - in - mycinose - out noncatalytic entry pose , increasing the probability of n - demethylation occurring in an otherwise atypical reaction site . despite discovery of a new n - demethylation activity for monooxygenase mycg , caution should be exercised in interpreting reconstituted p450 activity generated in a surrogate redox system , particularly until the generality of this approach is further investigated . however , an interesting hypothesis is suggested : p450 enzymes could be even more versatile in vivo than previously considered , because these biocatalysts might interact with a variety of redox partners to gain alternative activities . this may impart evolutionary advantages to the host organisms through detoxifying a more diverse range of xenobiotics or synthesizing more secondary metabolites to adapt to ever - changing environments . clear indications that the p450-omes of microbes ( e.g. , streptomyces coelicolor ) are capable of interacting with a limited set of redox partners are consistent with a broader role for redoxin - mediated selectivity in these fascinating enzyme systems . Output:	cytochrome p450 enzymes are capable of catalyzing a great variety of synthetically useful reactions such as selective c h functionalization . surrogate redox partners are widely used for reconstitution of p450 activity based on the assumption that the choice of these auxiliary proteins or their mode of action does not affect the type and selectivity of reactions catalyzed by p450s . herein , we present an exceptional example to challenge this postulate . mycg , a multifunctional biosynthetic p450 monooxygenase responsible for hydroxylation and epoxidation of 16-membered ring macrolide mycinamicins , is shown to catalyze the unnatural n - demethylation(s ) of a range of mycinamicin substrates when partnered with the free rhodococcus reductase domain rhfred or the engineered rhodococcus - spinach hybrid reductase rhfred - fdx . by contrast , mycg fused with the rhfred or rhfred - fdx reductase domain mediates only physiological oxidations . this finding highlights the larger potential role of variant redox partner protein protein interactions in modulating the catalytic activity of p450 enzymes .
PubmedSumm2095	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: climate change impacts are being observed worldwide on human and natural systems , including water resources , forestry , food production , coastal areas , ecosystems , industry , settlements and human health ( intergovernmental panel on climate change ( ipcc ) 2007 ; stern 2006 ) . many recent publications have highlighted the health impacts of climate change on populations , as well as how these effects are expected to evolve as the climate continues to change . potential impacts include more illnesses and deaths related to poor air quality , food shortages , water- and food - borne contamination , extreme weather events and changing patterns of diseases spread by animals , ticks and insects ( chiotti and lavender 2008 ; confalonieri et al . 2007 ; haines et al . 2006 ; 2006 ; patz et al . 2005 ; sequin and clarke 2008 ) . as evidence accumulates , there are increasing calls for public health decision makers to take preventative actions or adaptations ( e.g. , vulnerability assessments , surveillance , warning systems , public education and outreach ) to reduce the health risks associated with climate change ( confalonieri et al . 2007 ; mcmichael et al . a number of generic frameworks have been developed to assist decision makers and communities in preparing for the expected impacts of climate change in various sectors ( lim bs et al . these frameworks have been instrumental in ( 1 ) raising awareness among decision makers in many climate - sensitive sectors about the need to view climate change as a risk management issue ; ( 2 ) highlighting the respective roles of policy makers and program managers in planning for climate change impacts ; ( 3 ) bringing to the forefront the need to establish institutional mechanisms for adaptation development ; and ( 4 ) explicitly incorporating adaptation and climate change considerations into policies and programs designed to reduce risks . recently , some researchers have developed climate change assessment and risk management frameworks specifically for health decision makers ( berry 2008 ; kovats et al . the focus now is on mainstreaming climate change and health adaptation considerations and information into existing decision - making processes , rather than creating new policies or policy instruments ( dougherty and elasha 2004 ; smit and wandel 2006 ) . this entails incorporating information about climate - related health risks into existing risk management activities and integrating efforts among different health sector partners to develop coordinated responses ( berry 2008 ) . to this end , knowledge is required about the expected risks to health ( knzli 2010 ) , so that policies can be developed and public health systems can incorporate and act on this new information . in canada , each province has its own organizational structure , policy direction and set of priorities for delivering public health services . in ontario , the local boards of health deliver public health programs and services based on the ontario public health standards ( government of ontario 2008 ) . the ontario public health standards represent minimum requirements for ontario s 36 boards of health , although many boards of health do provide additional programs and services beyond those stipulated in the standards . population health and health promotion activities include key risk management activities related to assessing , planning , delivering , managing and evaluating public health programs and services . several required programs and protocols relate to a number of climate - sensitive threats to health infectious disease prevention and control , rabies prevention and control , vaccine preventable diseases , child health , food safety , water safety , health hazard prevention and management , and public health emergency preparedness ( fig . 1 ) . no provisions specifically require assessments or monitoring of climate change health impacts , although the ontario public health standards state that the board of health shall increase public awareness of health risk factors associated with climate change ( government of ontario 2008 ) . for each program and protocol , generic risk management activities form the basis of efforts to protect health . as illustrated , the foundational standards include population health assessment , surveillance , research and knowledge exchange , and program evaluation.fig . 1ontario public health standards : relationship between the principles , foundational standard , and program standards . source : government of ontario 2008 ontario public health standards : relationship between the principles , foundational standard , and program standards . source : government of ontario 2008 in canada , the risk management approach is well accepted and understood by the public health community , and several risk management frameworks exist to direct the activities of public health officials ( health canada 2003 ) . the question therefore is whether health sector officials can actually benefit from the frameworks proffered by climate change adaptation researchers , and if so , in what ways . in this article , the utility of climate change and health adaptation frameworks to support the development of adaptation measures in ontario is examined by comparing them with the ontario public health standards . the results from a key informant survey of public health officials across canada about climate - related health issues are presented to further analyze the utility of the frameworks and to identify other requirements for addressing climate - related health risks . ( 1 ) a literature review was conducted to select climate change and health adaptation frameworks that could be compared with the ontario public health standards . the purpose was to identify similarities and differences among the frameworks for assessing climate - related health vulnerabilities and adaptation options . ( 2 ) thirty semi - structured key informant interviews were conducted among local health authorities in canada . the purpose was to identify awareness levels , key activities and allocated resources related to climate change and health . a systematic literature search reviewed electronic databases , reference lists , books and reports ( both published and grey literature ) on the topic of climate change and adaptation , with a specific focus on health . domestic and international sources were included ; international ones when they had a canadian component or provided a theoretical or methodological basis related to the health impacts of climate change . examples of searched keywords included climate change and human health , climate change adaptation , climate change frameworks , extreme weather , impacts of global warming , adaptation policies and climate change mainstreaming . retrieved articles were scanned for information on the actions needed to increase the capacity to manage health risks associated with a changing climate . two frameworks were selected for the comparison with the ontario public health standards based on their comprehensiveness and the fact that they were specifically designed for the health sector ( 2003 ) ( fig . 2 ) and berry ( 2008 ) ( fig . 3).fig . source : berry 2008 steps in assessing vulnerability and adaptation to the health impacts of climate change . ( 2003 ) focuses on the steps and methods for assessing human health vulnerability to climate change . the framework proposed by berry ( 2008 ) builds on the work of kovats et al . ( 2003 ) by providing information on the steps to be considered when developing adaptation options to complement existing policies , institutional structures and processes ( fig . 3 ) . furthermore , it corresponds broadly to the key steps of standard risk management frameworks that are employed in the health sector in canada . basic steps include early and continuous engagement of stakeholders and other sectors , identifying adaptation options , modifying or establishing institutional mechanisms , formulating or modifying policies , incorporating adaptation into programs and projects , and monitoring and evaluating the results . to gauge the degree to which climate change risks are currently considered in policy and planning , 30 semi - structured key informant interviews were conducted in cities of various sizes across canada in 2006 , including eight interviews in ontario ( canadian public health association 2006 ) . senior public health officials from the largest city of each province were interviewed , and , based on their recommendations , follow - up interviews were conducted with other health professionals to collect further information ( fig . source : canadian public health association 2006 breakdown of interviewees by professional role . source : canadian public health association 2006 the survey instrument ( table 1 ) was designed to examine the extent to which public health officials ( or organizations ) are aware of climate change risks to health , are integrating such risks into their policies and planning , are able to respond to multiple health vulnerabilities and have identified key indicators or factors for enhancing adaptive capacity.table 1informant interview survey instrumentquestion 1.0do you believe that weather and climate have a significant impact on the health of people in your community?question 2.0do you feel that climate change will increase risks to the health of people in your geographic jurisdiction ? if yes , in what ways?question 2.1has climate change been identified as a potential current or future public health issue for your geographical jurisdiction?question 3.0what are you currently doing to protect population health from the specific climate - related health risks that you have identified as of concern to your organization in question 2.0?question 3.1do you find it necessary within the next 10 years to take new actions , or to expand current efforts , to reduce climate - related risks to health in your geographical jurisdictions ? if no , why do you find it unnecessary?question 3.2do you use climate - related information ( e.g. , information about extreme weather events , warmer temperatures ) in planning and program development activities at your organization ? if yes , please give examples.question 4.0do particular challenges exist in your efforts to reduce the health risks identified as a concern to your organization in question 2.0?source : canadian public health association 2006 informant interview survey instrument source : canadian public health association 2006 a systematic literature search reviewed electronic databases , reference lists , books and reports ( both published and grey literature ) on the topic of climate change and adaptation , with a specific focus on health . domestic and international sources were included ; international ones when they had a canadian component or provided a theoretical or methodological basis related to the health impacts of climate change . examples of searched keywords included climate change and human health , climate change adaptation , climate change frameworks , extreme weather , impacts of global warming , adaptation policies and climate change mainstreaming . retrieved articles were scanned for information on the actions needed to increase the capacity to manage health risks associated with a changing climate . two frameworks were selected for the comparison with the ontario public health standards based on their comprehensiveness and the fact that they were specifically designed for the health sector ( 2003 ) ( fig . 2 ) and berry ( 2008 ) ( fig . source : berry 2008 steps in assessing vulnerability and adaptation to the health impacts of climate change . ( 2003 ) focuses on the steps and methods for assessing human health vulnerability to climate change . the framework proposed by berry ( 2008 ) builds on the work of kovats et al . ( 2003 ) by providing information on the steps to be considered when developing adaptation options to complement existing policies , institutional structures and processes ( fig . 3 ) . furthermore , it corresponds broadly to the key steps of standard risk management frameworks that are employed in the health sector in canada . basic steps include early and continuous engagement of stakeholders and other sectors , identifying adaptation options , modifying or establishing institutional mechanisms , formulating or modifying policies , incorporating adaptation into programs and projects , and monitoring and evaluating the results . to gauge the degree to which climate change risks are currently considered in policy and planning , 30 semi - structured key informant interviews were conducted in cities of various sizes across canada in 2006 , including eight interviews in ontario ( canadian public health association 2006 ) . senior public health officials from the largest city of each province were interviewed , and , based on their recommendations , follow - up interviews were conducted with other health professionals to collect further information ( fig . source : canadian public health association 2006 breakdown of interviewees by professional role . source : canadian public health association 2006 the survey instrument ( table 1 ) was designed to examine the extent to which public health officials ( or organizations ) are aware of climate change risks to health , are integrating such risks into their policies and planning , are able to respond to multiple health vulnerabilities and have identified key indicators or factors for enhancing adaptive capacity.table 1informant interview survey instrumentquestion 1.0do you believe that weather and climate have a significant impact on the health of people in your community?question 2.0do you feel that climate change will increase risks to the health of people in your geographic jurisdiction ? if yes , in what ways?question 2.1has climate change been identified as a potential current or future public health issue for your geographical jurisdiction?question 3.0what are you currently doing to protect population health from the specific climate - related health risks that you have identified as of concern to your organization in question 2.0?question 3.1do you find it necessary within the next 10 years to take new actions , or to expand current efforts , to reduce climate - related risks to health in your geographical jurisdictions ? if yes , what actions ? why do you find it necessary ? if no , why do you find it unnecessary?question 3.2do you use climate - related information ( e.g. , information about extreme weather events , warmer temperatures ) in planning and program development activities at your organization ? if yes , please give examples.question 4.0do particular challenges exist in your efforts to reduce the health risks identified as a concern to your organization in question 2.0?source : canadian public health association 2006 informant interview survey instrument source : canadian public health association 2006 table 2 shows the comparative analysis of the two selected frameworks and the ontario public health standards . considerable overlap exists between the activities required by the ontario public health standards and the steps in kovats et al . ( 2003 ) and berry ( 2008).table 2comparative analysis of the two climate change and health adaptation frameworks and the ontario public health standardsframework stepskovats et al . ( 2003)berry ( 2008)ontario public health standards ( government of ontario 2008)engagement of stakeholdersyesyesyesidentification of burden of climate - related illness or injuryyesyespartialidentification of vulnerable populationsyesnoyesprojection of future health impacts from climate changeyesnonoassessment of the effectiveness of programs and activities to reduce climate - related health risksyesyesyesidentification and prioritization of policy and program options to meet health needsyesyesyesestablishment of policy mechanismsnoyesyestailoring and implementation of programs and policiesnoyesyessurveillance and monitoringpartialpartialpartialknowledge exchange and raising awareness of climate - related health riskspartialpartialyes comparative analysis of the two climate change and health adaptation frameworks and the ontario public health standards the ontario public health standards do not explicitly call for surveillance and monitoring of climate change health impacts , as is advocated for in the adaptation frameworks . however , they provide an institutional mechanism with generic program requirements that are able to manage climate - related risks . therefore public health officials in ontario do not require additional guidance , although specific information on how to integrate findings from climate scenarios and models to understand future risks and vulnerabilities ( kovats et al . the findings from the interviews of canadian public health authorities suggest that climate change adaptations have yet to be implemented in many regions across canada ( canadian public health association 2006 ) . this suggests that factors other than institutional mechanisms are necessary for public health authorities to address climate change impacts . elements identified as being essential for implementing climate change adaptation measures into public health policy and programs include additional resources , increased awareness and climate specific information about risks to health ( fussel and klein 2004 ) . the key informant interviews revealed a broad awareness of climate change and related health issues . all respondents indicated that weather and climate do have a significant impact on health ; most ( 76% ) believed that climate change will increase risks to health in their jurisdictions ( canadian public health association 2006 ) . however , about half ( 54% ) indicated that climate change had not been identified as a current or future public health issue in their jurisdictions . respondents who did report that climate change was considered an important public health issue in their jurisdiction also indicated that it was not considered a high priority ( canadian public health association 2006 ) . key challenges in addressing climate - related health risks included a lack of resources , a lack of prioritization of climate impacts on public health and a lack of adequate knowledge about potential linkages between climate change and health ( canadian public health association 2006 ) . many identified that climate - related information pertaining to extreme weather warnings , air quality reports , west nile virus surveillance and monitoring for flooding is used in planning and program development . however , most said they did not have adequate knowledge to make informed decisions about whether they need to take new actions , or extend current efforts , within the next 10 years . when identifying the various challenges faced in reducing climate - related health risks , the respondents did not identify a lack of tools or frameworks for managing risks ( canadian public health association 2006 ) . the comparative analysis and key informant interviews suggest that regular population health assessments and risk management activities by local public health units in ontario can address many risks related to climate hazards and emergencies , when information about such risks is made available to health officials . many activities identified in the ontario public health standards mirror those proposed by the climate change and health adaptation frameworks . the overlap between these standards and the frameworks therefore suggests that public health practices in ontario are theoretically able to accommodate many of the proposed climate considerations and adaptation strategy development measures . for example , emergency management activities such as planning and preparedness for weather - related emergencies , which are expected to increase in frequency and severity with climate change ( intergovernmental panel on climate change ( ipcc ) 2007)flow directly from the hazards identification phase within the ontario public health standards . table 2 shows the comparative analysis of the two selected frameworks and the ontario public health standards . considerable overlap exists between the activities required by the ontario public health standards and the steps in kovats et al . ( 2003 ) and berry ( 2008).table 2comparative analysis of the two climate change and health adaptation frameworks and the ontario public health standardsframework stepskovats et al . ( 2003)berry ( 2008)ontario public health standards ( government of ontario 2008)engagement of stakeholdersyesyesyesidentification of burden of climate - related illness or injuryyesyespartialidentification of vulnerable populationsyesnoyesprojection of future health impacts from climate changeyesnonoassessment of the effectiveness of programs and activities to reduce climate - related health risksyesyesyesidentification and prioritization of policy and program options to meet health needsyesyesyesestablishment of policy mechanismsnoyesyestailoring and implementation of programs and policiesnoyesyessurveillance and monitoringpartialpartialpartialknowledge exchange and raising awareness of climate - related health riskspartialpartialyes comparative analysis of the two climate change and health adaptation frameworks and the ontario public health standards the ontario public health standards do not explicitly call for surveillance and monitoring of climate change health impacts , as is advocated for in the adaptation frameworks . however , they provide an institutional mechanism with generic program requirements that are able to manage climate - related risks . therefore public health officials in ontario do not require additional guidance , although specific information on how to integrate findings from climate scenarios and models to understand future risks and vulnerabilities ( kovats et al . the findings from the interviews of canadian public health authorities suggest that climate change adaptations have yet to be implemented in many regions across canada ( canadian public health association 2006 ) . this suggests that factors other than institutional mechanisms are necessary for public health authorities to address climate change impacts . elements identified as being essential for implementing climate change adaptation measures into public health policy and programs include additional resources , increased awareness and climate specific information about risks to health ( fussel and klein 2004 ) . the key informant interviews revealed a broad awareness of climate change and related health issues . all respondents indicated that weather and climate do have a significant impact on health ; most ( 76% ) believed that climate change will increase risks to health in their jurisdictions ( canadian public health association 2006 ) . however , about half ( 54% ) indicated that climate change had not been identified as a current or future public health issue in their jurisdictions . respondents who did report that climate change was considered an important public health issue in their jurisdiction also indicated that it was not considered a high priority ( canadian public health association 2006 ) . key challenges in addressing climate - related health risks included a lack of resources , a lack of prioritization of climate impacts on public health and a lack of adequate knowledge about potential linkages between climate change and health ( canadian public health association 2006 ) . many identified that climate - related information pertaining to extreme weather warnings , air quality reports , west nile virus surveillance and monitoring for flooding is used in planning and program development . however , most said they did not have adequate knowledge to make informed decisions about whether they need to take new actions , or extend current efforts , within the next 10 years . when identifying the various challenges faced in reducing climate - related health risks , the respondents did not identify a lack of tools or frameworks for managing risks ( canadian public health association 2006 ) . the comparative analysis and key informant interviews suggest that regular population health assessments and risk management activities by local public health units in ontario can address many risks related to climate hazards and emergencies , when information about such risks is made available to health officials . many activities identified in the ontario public health standards mirror those proposed by the climate change and health adaptation frameworks . the overlap between these standards and the frameworks therefore suggests that public health practices in ontario are theoretically able to accommodate many of the proposed climate considerations and adaptation strategy development measures . for example , emergency management activities such as planning and preparedness for weather - related emergencies , which are expected to increase in frequency and severity with climate change ( intergovernmental panel on climate change ( ipcc ) 2007)flow directly from the hazards identification phase within the ontario public health standards . some studies have identified general health risks related to climate change for people living in ontario ( abelsohn et al . however , information is lacking about the direct and indirect climate - mediated processes involved at local scales within the time frames needed to inform decision making ( tamerius et al . challenges exist in the development of new knowledge about the expected impacts of climate change on health given the very complex causal sources and pathways and intertwined interactions between them ( forastiere 2010 ; xun et al . once current or near - term health risks related to climate variability are identified , activities to protect health before , during and after a health emergency can be adopted . the ontario public health standards can address existing diseases that are being exacerbated by climate change ( e.g. , lyme disease ) through current surveillance and public education and outreach activities . in this regard , the standards require the board of health shall work with community partners to improve public knowledge of infectious diseases of public health importance and infection prevention and control practices ( government of ontario 2008 ) . given that the requirements in ontario already provide direction for addressing many health concerns that are climate sensitive , new processes or risk management steps are not required in order to mainstream climate change . in order to mainstream climate change concerns , public health decision making requires information about climate - related health risks to inform existing risk management activities . these activities , such as population health assessment , surveillance and public education and outreach can only account for climate factors if information on climate - related risks is available to public health practitioners and researchers ( e.g. , what are the climate hazards and associated health impacts , where will they occur , when will they occur and who will be most affected ? ) . for example , while it is known that extreme heat can impact morbidity ( green et al . 2010 ) and mortality ( sequin and clarke 2008 ) , assessments of heat - health vulnerabilities require projections of the future frequency and severity of extreme heat events , information related to the geographical variation in exposure to extreme heat , identification of vulnerable populations and identification of actual temperature and morbidity / mortality thresholds . if such information is not available to be considered in the assessment step of existing frameworks , important risks to human health can not be mainstreamed . public health interventions that may be implemented to reduce climate change impacts on health within relevant time scales ( 510 years ) need to be identified through assessments ( ortz bult et al . however , information in these time scales about projected risks to health is often not available to decision makers . the need in canada for better understanding of health risks associated with climate change , as reported in the key informant interviews , is also reflected in resolutions or declarations passed by the canadian public health association ( 2001 ) , the world health organization ( world health organization department of communications may 22 , 2009 ) and the world medical association ( world medical association 2010 ) ; these call for more research into the health impacts of climate change . a lack of information about risks was also identified as a major barrier to public health adaptation in a recent survey of local health departments in the united states ( balbus et al . the development , analysis and transfer of knowledge among local , provincial , federal and international health authorities showing how climate change might impact the health of local populations is critical for facilitating adaptation efforts in the health sector , and should be made a priority area of collaborative efforts in the future ( ebi 2008 ; yohe and ebi 2005 ) . at the national level , health canada has completed a country - wide assessment of health vulnerabilities to climate change impacts that can help inform the development of needed adaptations by health authorities at local and regional levels ( sequin and clarke 2008 ) . moreover , new knowledge is being developed at the national level on climate - related health risks pertaining to extreme heat events ( health canada 2011 ) , vector - borne diseases and challenges faced by northern populations ( government of canada 2009 ; health canada 2009 ) . a key finding of this study is that a strong foundation for developing and implementing adaptive actions to protect health already exists through current public health requirements and activities in ontario . existing risk management practices can be applied to reduce some climate - related risks , such as air pollution , extreme weather events , infectious diseases ( e.g. , lyme disease ) , and water and food contamination . thus , the current risk management practices already allow for the mainstreaming of most climate - related health risks . there is no need to implement or adopt separate climate change and health adaptation frameworks , particularly given limited resources and competing demands faced by public health officials . for this reason , frameworks that advocate , or propose , new climate change risk management processes and procedures for reducing health risks , will have limited applicability for public health and emergency management decision makers in ontario and other provinces in canada , as well as in other countries with similar risk management systems in place . however , countries and regions with less comprehensive population health assessment , surveillance and public education and outreach activities may benefit more from the guidance provided by the adaptation frameworks examined here . the findings also indicate that risk management systems must be informed by adequate information about climate - related health risks for public health authorities to mainstream climate change concerns and take needed actions to protect at - risk populations . other research has also indicated that adaptation efforts within and outside of canada require a better understanding of current and geographically specific risks from climate - related hazards , and projected risks due to climate change in time scales relevant to public health decision makers ( haines et al . ( 2007 ) call for the development of multidisciplinary research teams which enhance collaboration between climate modelers and the health community to provide this information . what is needed is more information , including case studies and community examples that aid public health officials in their efforts to obtain , analyze and integrate findings from climate scenarios and models to gauge future impacts on health . climate change can only be mainstreamed into health sector activities when such information is available and incorporated into existing public health risk management activities , as well as decision - making processes or standards ( e.g. , local population health assessments , surveillance , public education and outreach ) . given the critical role of information about climate - related health risks , future research and program development efforts to support greater mainstreaming would benefit from investigation of the most effective and efficient ways to provide this information to governmental and non - governmental authorities charged with protecting citizens from environmental and climate - related health risks . the complexity and uncertainty inherent in understanding climate - related health risks means that investigation into best practices for knowledge development , translation and transfer among researchers , decision makers and other stakeholders is required ( balbus et al . the findings here also strongly support the call by the world health organization for countries to take immediate measures to strengthen existing public health systems to reduce current impacts on health from climate extremes and to prepare for future climate change ( world health organization 2005 ) . robust population health assessments , surveillance and public education and outreach activities , in particular , are key to protecting populations by mainstreaming climate change considerations into existing activities . Output:	objectiveit is now recognized that climate change affects human health . the question is how to adapt . this article examines mainstreaming climate considerations into public health programs and the utility of climate change and health adaptation frameworks , using ontario , canada , as a case study.methodsa literature review identified climate change and health adaptation frameworks for comparison with the ontario public health standards . key informant interviews gauged the extent to which climate change risks are currently considered in policy and planning.resultsontarios public health standards already require many of the risk management activities identified in climate change and health adaptation frameworks . however , public health officials require additional information about linkages between climate change and health to manage risks.conclusionsrisk management activities such as population health assessments , surveillance and public education and outreach can address many key risks related to climate hazards when information about the risks , vulnerable populations and time scales is made available to health officials . the development , analysis and transfer of this information should be considered a priority at all levels within the public health sector .
PubmedSumm2096	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: alcoholic steatohepatitis ( ash ) represents the intermediate stage of liver injury in the progression of steatosis , inflammation , and hepatocellular apoptosis and necrosis that occurs in the development of alcoholic cirrhosis . this paper will address the role of ethanol - induced aberrant hepatic methionine metabolism in the pathogenesis of ash , in particular through its epigenetic effects on the expressions of genes relevant to alcoholic liver injury . initially , we will discuss the normal hepatic methionine metabolic cycle , including its interactions with folate , vitamin b12 , and vitamin b6 , and its relevance to the epigenetic regulation of gene expression . this will be followed by experimental evidence for multiple effects of ethanol on the methionine metabolic cycle that lead to liver injury through altered methylation and expressions of genes relevant to steatosis , apoptosis , and oxidative stress . hepatic methionine metabolism can be visualized in two parts , the transmethylation cycle for production of s - adenosylmethionine ( sam ) and its metabolism to homocysteine and the transsulfuration pathway for the reduction of homocysteine for the synthesis of glutathione ( gsh ) ( figure 1 ) . endogenous 5-methyltetrahydrofolate ( 5-mthf ) that is derived from dietary folate is substrate for the initial vitamin - b12-dependent reaction of methionine synthase ( ms ) that converts homocysteine to methionine . the additional pathway of betaine homocysteine methyltransferase ( bhmt ) uses the choline product betaine as substrate with homocysteine for methionine synthesis and is considered a salvage pathway when ms is compromised by toxins including ethanol . methionine is metabolized to sam by methionine adenosyltransferase ( mat ) , which is a product of the mat1a gene in adult liver . sam is utilized at a rate of 68 grams per day , primarily by different methyltransferase reactions that include phosphatidylethanolamine transferase ( pemt ) to yield phosphatidylcholine ( pc ) and many others that methylate gene - specific dna and various histone residues . in addition , sam levels are regulated by the activity of glycine - n - methyltransferase ( gnmt ) with sarcosine as its end product ( not shown in figure 1 ) . while sam is the sole substrate for methyltransferase reactions , its product s - adenosylhomocysteine ( sah ) is the principal inhibitor of the same reactions . consequently , the sam - to - sah ratio is used conveniently as an index of methylation capacity since the km and ki for most of these reactions are in similar range . sah is hydrolyzed to homocysteine by sah hydrolase ( sahh ) , which is a reversible enzyme depending upon the abundance of each product . the initial reaction in the transsulfuration pathway is facilitated by sam and involves the reduction of homocysteine through the cystathionine beta synthase ( cs ) reaction to cystathionine . both cs and the subsequent cystathionase reaction require vitamin b6 as cofactor . therefore , it may be predicted that conditions that limit the production of sam and the availability of vitamin b6 reduce the production of gsh and its antioxidant properties . summarizing the key points , homocysteine is regulated by three enzymes , ms , sahh , and cs , each of which can be compromised to result in homocysteine elevation which is involved in the pathogenesis of ash . the metabolite sam plays a key role in regulating gene methylation and expression and a second one by regulating the production of the principal antioxidant gsh . there is abundant and accumulating evidence that experimental ethanol exposure and chronic alcoholism influence the expressions and activities of several enzymes in the hepatic methionine cycle . for example , transcript expressions of ms , mat1a , and cs were decreased in liver biopsies from ash patients , while decreased expressions and activities of ms , mat1a , and sahh , but increased gnmt , were found in chronic ethanol - fed micropigs with histologically proven ash . reduced activities of ms with compensatory increase in bhmt expressions and activities were also found in ethanol - fed rats [ 68 ] . since mat1a is susceptible to oxidation and nitrosylation of its cysteine 121 residue , its reduced activity in ash may be attributed to the generation of reactive oxygen species ( ros ) that are produced by the ethanol metabolizing enzyme cytochrome p4502e1 ( cyp2e1 ) [ 9 , 10 ] . thus , since sam regulates the generation of gsh through its facilitating effect on cs , the reduced antioxidant potential caused by reduced gsh would predictably decrease the effectiveness of mat1a to generate sam . these relationships were supported by in vitro studies in chemically treated primary hepatocytes in which elevation of prooxidant cyp2e1 was associated with reduced sam levels . also , studies in ethanol - fed baboons and micropigs showed close correlations between liver levels of gsh and sam [ 12 , 13 ] . the conventional understanding of the pathogenesis of ash includes a series of events that are triggered by ethanol - induced transport of enterotoxic lipopolysaccharide ( lps ) from the intestine to hepatic kupffer cells that are in turn induced to release cytokines such as tumor necrosis factor alpha ( tnf ) which initiates pathways of hepatocyte apoptosis , inflammation , and necrosis . these processes are facilitated by the effect of the ethanol metabolizing hepatocyte enzyme cyp2e1 which is a powerful producer of ros . in addition , the metabolism of ethanol by alcohol to acetaldehyde alters the nadh / nad redox potential which promotes steatosis through its effects on fatty acid oxidation in the liver . steatosis , which is an early sign in development of ash , represents the combination of increased hepatic lipogenesis and decreased fatty acid oxidation and reduced triglyceride export . many of these avenues of injury have been linked to ethanol - induced aberrant methionine metabolism in experimental animal models , as described below . studies that used the model of the intragastric ethanol - fed c57bl6 mouse linked enhanced lipogenesis and apoptosis to hyperhomocysteinemia , which is known to trigger endoplasmic reticulum ( er ) stress pathways that include upregulation of the chaperone glucose - regulated protein ( grp78 ) , the transcription factor sterol response element binding protein ( srebp1-c ) and its downstream lipogenesis genes , and the growth arrest and dna damage ( gadd153 ) pathway for apoptosis . since the histopathology of steatosis and apoptosis and these gene activations were prevented by concomitant provision of betaine which lowered homocysteine through the bhmt salvage pathway ( figure 1 ) , the authors concluded that there is a specific role for ethanol - induced hyperhomocysteinemia in the pathogenesis of ash by activation of er stress pathways . more recent studies in the ethanol - fed rat model of ash associated apoptosis with increased liver sah and steatosis with reduced activity of pemt which regulates triglyceride export from the liver [ 1719 ] . others associated apoptosis in ethanol - fed mice with increased levels of sah which sensitized isolated primary hepatocytes to the effects of tnf , while a later study in the same model associated this effect with reduced mitochondrial sam ( msam ) caused by competitive inhibition of its mitochondrial transport by elevated sah . the original finding of a mitochondrial transporter for cytoplasmic sam that is inhibited by sah was complemented by discovery of a mitochondrial gsh transporter that is inactivated by ethanol but is sustained by coadministration of sam . a role for msam in prevention of the prooxidant effects of ethanol was supported by intervention studies that demonstrated a protective effect of sam against decreased mitochondrial respiration and increased levels of mitochondrial superoxide and inducible nitric oxide synthase ( inos ) . sam may also play a role in prevention of hepatocellular carcinoma ( hcc ) in ald by inhibition of angiogenesis genes . the pathogenesis of ash includes activations of many genes involved in liver injury , each of which are potentially and epigenetically regulated by alterations in hepatic levels of sam and sah and their methylation capacity ratio . our laboratory explored these relationships in two ethanol - fed animal models , the micropig and the genetically altered mouse . an initial study demonstrated the suitability of the micropig for study of the pathogenesis of ash , in which its characteristic histopathology was induced by feeding ethanol at 40% of kcal over 12 months together with progressive increase in serum homocysteine levels and reduced gene methylation capacity as shown by decrease in the liver sam and the sam - to - sah ratio . subsequently , we included folate deficiency in the dietary regimen in order to accentuate changes in methionine metabolism through reduction of the initial methyl donor 5-mthf ( figure 1 ) , with the additional rationale of the well - known association of folate deficiency with chronic alcoholism [ 29 , 30 ] . in contrast to the findings in the previous study only 3 months were required for the development of the histopathology of ash , while increases in timed serum homocysteine levels and lowering of the hepatic sam - to - sah ratio were the greatest in the group fed the combined folate - deficient and ethanol diet compared to groups fed either diet alone . using liver samples from the same groups of micropigs , we found progressive increase in transcript and protein levels of genes relevant to steatosis and liver injury from micropigs fed control , folate - deficient , ethanol , and ethanol with folate - deficient diets , together with increased expressions of cyp2e1 and er stress pathway genes for the marker grp 78 , lipogenesis that included srebp 1-c and fatty acid synthase ( fas ) , and apoptosis including gadd 153 . the finding that these gene expressions correlated with changes in the sam - to - sah ratio supported the relationship of the pathogenesis of ash to ethanol - induced reduction of gene methylation capacity . this concept was supported by subsequent studies in which the coadministration of sam sustained normal histology while preventing changes in the sam - to - sah ratio , and activations of the same er stress genes for lipogenesis and apoptosis , as well as genes relevant to oxidative liver injury including cyp2e1 , nicotinamide adenine dinucleotide phosphate oxidase ( nox1 ) , and inducible nitric oxide synthase ( inos ) . epigenetics refers to the effects of external factors on the expressions of genes that are unrelated to changes in their dna sequences . gene expression is regulated at the levels of both dna and its histone infrastructures that exist within nuclear chromatin . in the context of the present paper , such changes can be attributed to the regulatory effects of increased or decreased methylation on dna and histones , since folate contains the parent methyl group and sam is the principal methyl donor in all methyltransferase reactions ( figure 1 ) . as described in recent reviews , the effect of chronic ethanol exposure on the activation or suppression of selected ash - related genes can be ascribed to different methylation effects at histone 3 lysine ( k ) residues , whereby gene expression is enhanced by methylation at h3mek4 but is silenced by methylation at h3mek9 [ 34 , 35 ] . furthermore , ethanol exposure enhances gene expression by acetylation at h3ack9 , whereas sirtuin1 , a known histone de - acetylase , is reduced in livers of ethanol - fed mice . therefore , the effects of ethanol exposure on specific histone methyltransferases , acetylases , and deacetylases may be crucial to the epigenetic control of the expressions of genes relevant to liver injury . further epigenetic regulation occurs at the level of dna , where increased methylation of promoter region cytosine residues suppresses gene expression and vice versa . for example , global dna hypomethylation occurred in the ethanol - fed sam - deficient rat and mat1a knockout mouse with enhanced carcinogenesis , as well as in our folate - deficient chronic alcoholic micropig model in association with increased dna oxidation and strand breaks . others found reduced expression of the dna methyltransferase dnmt3b that correlated negatively with increased blood alcohol levels in chronic alcoholics together with increased global dna methylation . we explored epigenetic regulation of ash in a novel model of the c57bl6 cs - deficient mouse fed intragastric ethanol at 37% of kcal for 4 weeks . this model is particularly relevant since , as noted above , the same method was used to induce ash in c57bl6 wild - type mice with findings of altered methionine metabolism , all of which were corrected by the methyl donor betaine . since cs heterozygosity is known to increase serum homocysteine levels and ethanol exposure inhibits ms expression and activity [ 5 , 6 ] we reasoned that both ethanol exposure and genotype would accentuate aberrant methionine metabolism with the greatest effect in the combined group . our study showed all the histopathological features of severe ash in the combined ethanol - fed heterozygous group , which also showed the greatest reduction in the sam - to - sah ratio and the greatest increase in expressions of the er stress genes srebp 1-c , grp78 , and gadd153 . while immunohistological staining of liver slices showed that methylated histone residue h3k9 was specifically reduced in centrilobular regions which were the sites of maximal steatosis in the ethanol - fed heterozygous group , the chromatin immunoprecipitation ( chip ) assay using antibody to h3k9 showed its reduced presence in the promoter regions of the same er stress genes . since h3k9 associates with gene suppression , its reduced presence in liver from the combined ethanol - fed heterozygous group is consistent with activation of these genes . furthermore , the transcript expression of g9a , a histone methyltransferase that is required for methylation of k9 residues , was specifically reduced in both ethanol - fed groups in contrast to other k9 methyltransferases . the mechanism for reduced expression of the histone methyltransferase g9a is unclear , but others have shown effects of ethanol - induced ros on chromatin remodeling as a mechanism for changes in histone acetylation reactions . summarizing , these studies provided an epigenetic mechanism for the relationship of ethanol - induced aberrant methionine metabolism on the epigenetic regulation of expression of selected er stress genes that are relevant to lipogenesis and apoptosis in ash . using the chip assay in a mouse macrophage raw cell line and in vivo in lps - treated mice , others showed that sam inhibited the lps - induced activations of tnf and inos by blocking the binding of h3mek4 to the promoter regions of these genes . data from two groups suggest epigenetic regulation of proteosome inhibition as a mechanism for enhanced liver injury . in one study in an ethanol - fed rat model , proteosome inhibition was associated with alterations in histone acetylation and methylation , whereas methylation of a specific subunit by provision of sam prevented proteosome degradation in ethanol - exposed hepatocytes . a study of patients with cirrhosis of diverse etiologies including alcoholism found significant elevations in serum homocysteine and cystathionine , which was suggestive of a block in the transsulfuration pathway for homocysteine elimination and production of gsh . compared to healthy subjects , we found increased serum homocysteine levels in alcoholics with or without ash in association with elevated serum sah levels . while others have described elevated homocysteine levels in chronic alcoholics in association with deficiencies of folate and vitamin b6 , we found that ash patients also had marked increase in serum levels of cystathionine , the substrate for vitamin - b6-dependent cystathionase ( figure 1 ) , with comparable reduction in its product -aminobutyrate ( abu ) . furthermore , cystathionine levels and the abu - to - cystathionine ratio correlated with vitamin b6 levels and the abu - to - cystathionine ratio was a positive predictor of the presence of ash among alcohol drinkers . the growing experimental evidence linking impaired methionine metabolism with the onset and development of alcoholic liver disease prompted several clinical trials that attempted to demonstrate a role for sam in its treatment . one study showed that 15 days of intravenous sam at 2 g / day increased red blood cell gsh in 20 patients with ash . hepatic gsh levels were low in liver biopsies from 17 patients with ash or other chronic liver diseases and were normalized by sam treatment at 1.2 g / day in a 6-month controlled study . a later 2-year multicenter european trial of 123 ash patients found reduction in mortality or incidence of liver transplant from 30% in the placebo group to 16% in the sam group , which was significant after exclusion of childs class c patients from the analysis . a 2006 meta - analysis of 9 studies found insufficient evidence for the effective use of sam in the treatment of alcoholic liver disease . whereas the pc derivative polyenylphosphatidylcholine ( ppc ) was known to restore liver sam levels in ethanol fed rats , there was no benefit of this compound according to liver histopathology in 412 ash patients who underwent liver biopsies at baseline and after 24 months of treatment . we conducted a randomized treatment trial in 26 ash patients who participated in our baseline study and then received sam at 1.2 g / d or placebo daily for 24 weeks . all biochemical parameters of liver function improved over time with no differences between the groups . although serum sam levels rose over time with treatment , there were no changes in other methionine metabolites in either group . furthermore , there were no differences between groups in histopathology scores for steatosis , inflammation , or fibrosis in 14 patients who underwent liver biopsies before and after treatment . summarizing , the results from several clinical trials of sam in treatment of ash are inconclusive as to its effect . previous positive studies included patients with other causes of liver disease and showed no comprehensive histological data on the efficacy of sam . while our study is the first to provide data on potential changes in methionine metabolite levels and documents the lack of differences in histopathology , its limitations include relatively short exposure to sam and relatively small number of subjects . unlike studies in animal models that showed the efficacy of sam in prevention of ash [ 13 , 32 , 33 ] , the use of sam in the treatment of established liver disease may be compromised by lack of retention of sam by injured hepatocytes as well as decreased numbers of normally functioning hepatocytes . also , based on our methionine metabolite findings at baseline and the role of vitamin b6 in the transsulfuration pathway ( figure 1 ) , there may be a requirement for vitamin b6 supplementation in addition to sam in longer - term treatment of ash . abundant evidence now exists for the central role of aberrant hepatic methionine metabolism in the pathogenesis of ash . ethanol and its ros products inhibit key enzymes in the methionine cycle , in particular those involved in the synthesis of sam . since sam is the principal methyl donor for histones and dna and facilitates the production of gsh , its reduction influences the methylation and hence epigenetic regulation of many genes involved in alcoholic liver injury as well as the capacity for antioxidant defense in the pathogenesis of ash . whereas sam supplementation has been proven to prevent ash and its mechanisms in experimental animal models , clinical trials have failed to demonstrate a definitive effect of sam in the treatment of established ash . Output:	the pathogenesis of alcoholic steatohepatitis ( ash ) involves ethanol - induced aberrations in hepatic methionine metabolism that decrease levels of s - adenosylmethionine ( sam ) , a compound which regulates the synthesis of the antioxidant glutathione and is the principal methyl donor in the epigenetic regulation of genes relevant to liver injury . the present paper describes the effects of ethanol on the hepatic methionine cycle , followed by evidence for the central role of reduced sam in the pathogenesis of ash according to clinical data and experiments in ethanol - fed animals and in cell models . the efficacy of supplemental sam in the prevention of ash in animal models and in the clinical treatment of ash will be discussed .
PubmedSumm2097	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: . introduced through the active ingredient , excipients , or the atmosphere , it can induce phase transitions , dissolve soluble components , and increase interactions between the drug and excipients , all of which can adversely affect the physical and chemical stability of the drug substance to the detriment of drug product performance . it must therefore be accounted for at all stages of drug substance and product manufacturing . not surprisingly , of the many possible critical quality attributes of a drug substance , hygroscopicity , a measure of the water vapor taken up by a solid with the potential to effect surface and bulk properties , is routinely evaluated , typically by moisture sorption analysis , at the earliest stages of drug product development . the measurement of water vapor sorption isotherms must , however , be tied more directly to the mechanisms by which water is taken up to truly understand the impact that water sorption has on the properties of the solid . on a molecular level , water has the ability to interact with a substance in many ways , including ( i ) physisorption or binding to the surface by hydrogen bonding , ( ii ) physical entrapment to form liquid inclusions , ( iii ) absorption in localized disordered regions , ( iv ) chemical addition , and ( v ) hydrate formation . hydrates often crystallize because water improves crystal packing efficiency and satisfies the hydrogen bonding sites of the drug better than the drug itself . hydrates may be stoichiometric or nonstoichiometric.(3,1013 ) stoichiometric hydrates show step - shaped sorption / desorption isotherms characterized by a fixed water content over a defined relative humidity ( rh ) range and generally convert upon dehydration to a distinct phase , crystalline or amorphous . the waters of crystallization usually play a crucial role in stabilizing the molecular network . nonstoichiometric hydrates , however , have a continuously variable composition within a certain rh range that is not associated with a significant change in the crystal lattice , except for possibly anisotropic expansion of the network to accommodate water . dehydrating a nonstoichiometric hydrate may result in an isomorphic dehydrate ( desolvate ) , a one - component phase that exhibits the main structural features of its parent phase . desolvated solvates are usually metastable and easily take up the original solvent or sometimes other solvents to minimize free volume ( void space ) in the crystal . in some cases , nonstoichiometric hydrates lose crystallinity and become amorphous when the very last water is forced out by extreme drying conditions . whether a crystalline hydrate is targeted as the delivery vehicle for a drug in the formulated product or designed around in favor of a neat form , the dehydration and rehydration processes of a hydrate forming system can be complex and difficult to control . hydrates are generally expected to be thermodynamically more stable , hence less soluble and slower to dissolve than anhydrate forms above the critical water activity for hydrate formation . however , as particle size / shape distribution , specific surface area , and other surface properties heavily affect dissolution , a hydrate may dissolve faster than an anhydrate . additionally , some hydrates are intrinsically more soluble ( less stable ) in water than neat forms . such exceptions to the general solubility trend include ly334370 hcl , n-{[(5s)-3-(4-{6-[(1r,5s)-6-cyano-3-oxabicyclo[3.1.0]hex-6-yl]pyridin-3-yl}phenyl)-2-oxo-1,3-oxazolidin-5-yl]methyl}acetamide , ly156735 ( a melatonin agonist ) , and norfloxacin . norfloxacin is considered unusual because its more soluble hydrates are zwitterionic and at least one of its anhydrates is charge neutral . in this case , the higher solubility of the hydrate can be readily attributed to the stronger hydration of the highly charged zwitterions relative to the uncharged molecule in the anhydrous form . another compound that shows water - induced proton transfer to form zwitterionic hydrates is the 5-ht2a and h1 inverse agonist , 3-(4-dibenzo[b , f]oxepin-11-yl - piperazin-1-yl)-2,2-dimethylpropanoic acid ( ly2624803 or db7 , figure 1 ) . we previously used db7 as a model compound to test the value of combining computational crystal structure prediction ( csp ) methods with an industrial solid form screening program . from the experimental screen for solid forms , which encompassed a broad range of crystallization conditions and totaled over 300 experiments , db7 was identified in nine solid forms , including three neat polymorphs ( forms i iii ) , a stoichiometric dihydrate ( hy2 ) , a nonstoichiometric hydrate ( hya ) , three unstable isostructural solvates ( from methanol , ethanol , and 2-propanol ) , and an amorphous form . for an amphoteric molecule with measured pka values of 2.95 ( oxepine - n ) , 4.03 ( carboxylic acid ) , and 7.81 ( piperidine - n ) , application of the pka rule of three would suggest that proton transfer from the carboxylic acid to the piperidine - n is essentially complete rendering db7 zwitterionic , at least in aqueous solutions at moderate ph . however , solid state n nmr spectra provided an initial indication that only hy2 and hya contain zwitterions ( db7 ) ; the neat forms contain the neutral db7 molecule . molecular diagrams of db7 in the charge neutral anhydrous forms i iii and amorphous form and in the zwitterionic hydrates . the atomic numbering given on the charge neutral form is also used for zwitterionic phases , which are indicated with superscript z . in this study , we explore through a combination of experiment and computation the structural and thermodynamic relationships between the hydrate / anhydrate phases of db7 and their interconversion pathways as a function of temperature , water activity , and ph . a range of experimental techniques , along with computed lattice energies and chemical shift prediction of experimental and hypothetical structures , are used to understand at an atomistic level two distinct ( de)hydration mechanisms of stoichiometric and nonstoichiometric hydrates complicated by both structural disorder and proton transfer . db7 ( form ii , purity 99.6% ) was obtained from lilly research laboratories . the dihydrate ( hy2 ) was prepared by dissolving amorphous db7 ( prepared by rotovapping a dichloromethane solution of the compound ) in stirred water ( ph 5.9 ) at rt . within 15 min a thick white slurry , hy2 , had formed . the solid product was isolated by vacuum filtration and dried in a 75% rh chamber . after stirring in water at rt for 10 days hy2 transformed to the second hydrate ( hya ) . the solid product was isolated by vacuum filtration , washed / transferred with water , air - dried , and equilibrated for several days at 43% rh to further dry the sample . experiments on hy2 and hya were performed with samples that had been stored at 75% rh over a saturated nacl solution . for solubility experiments the hydrate samples were equilibrated at 92% rh ( over a saturated kno3 solution ) , for ssnmr experiments at 75 , 58 , 43 , 33 , 22 , and 11% rh and for h2o / d2o exchange ( raman ) experiments at 98 and 11% rh . the three anhydrates , forms i iii , were prepared according to ref ( 47 ) . single crystal x - ray diffraction data were collected using a bruker d8-based 3-circle geometry diffractometer equipped with a cuk ( hy2 ) or mok ( hya ) radiation source and a smart apex ii 6000 ccd area detector . hy2 and hya1.95 crystals were taken freshly from the mother liquor and hya1.73 after storing the sample at ambient conditions ( 40% rh ) . the structures were solved by direct methods using the program package wingx ( sir2011 and shelxl2013 ) . the aromatic and aliphatic hydrogen atoms were generated by a riding model on idealized geometries with uiso(h ) = 1.2ueq(c ) for aromatic and nonterminal aliphatic hydrogens and uiso(h ) = 1.5ueq(c ) for ch3 . water hydrogen atoms and hy2 n h were located from the difference map and refined with constrained o / n h bond distances and uiso(h ) = 1.5ueq(o)/uiso(h ) = 1.2ueq(n ) . the hya n h proton was identified from the difference map and refined isotropically . proton disorder of the hy2 water molecules is addressed in section 3.2.1 and hya water position occupancies / disorder in section 3.2.2 . the structure models for both hydrates were supported with electronic structure calculations ( sections 1116 of the supporting information ) . xrpd patterns were obtained using an xpert pro diffractometer ( panalytical , almelo , nl ) equipped with a theta / theta coupled goniometer in transmission geometry , programmable xyz stage with well plate holder , cu k1,2 radiation source with a focusing mirror , a 0.5 divergence slit and a 0.02 soller slit collimator on the incident beam side , a 2 mm antiscattering slit and a 0.02 soller slit collimator on the diffracted beam side , and a solid state pixcel detector . the patterns were recorded at a tube voltage of 40 kv and tube current of 40 ma , applying a step size of 2 = 0.013 with 40s/80s per step in the 2 range between 2 and 40. for nonambient rh measurements a vgi stage ( vgi 2000 m , middlesex , u.k . ) was used . the diffraction patterns were indexed using the first 20 peaks with dicvol04 , and the space group , which was determined based on a statistical assessment of systematic absences as implemented in the dash structure solution package , agreed with the single crystal data ignoring temperature effects . hastings pseudo - voigt function was used for peak shape fitting . for the rietveld refinements the db7 molecule was refined with restraints ( distances , angles , and aromatic rings flattened ) and the water molecules as oxygens only . moisture sorption and desorption studies were performed with the automatic multisample gravimetric moisture sorption analyzer sps23 - 10 ( proumid , ulm , germany ) . the measurement cycles were started at 40% with an initial stepwise desorption ( decreasing humidity ) to 0% , followed by a sorption cycle ( increasing humidity ) up to 95% and back to 0% relative humidity ( rh ) . the equilibrium condition for each step was set to a mass constancy of 0.001% over 60 min and a maximum time limit of 48 h for each step . differential scanning calorimetry ( dsc ) was conducted using a ta q1000 dsc , operated with thermal advantage release 5.4.0 software ( ta instruments , usa ) or a diamond dsc equipped with a controlled cooling accessory ( intracooler 1p ) and operated with pyris7.0 software ( perkinelmer , norwalk , ct , usa ) . a few milligrams of accurately weighed ( mettler um3 ultramicrobalance ) sample was heated in perforated or sealed al - pans . heating rates ranging from 1 to 50 c min were applied with a nitrogen purge . the temperature and heat flow of the ta q1000 were calibrated against indium melting . the diamond dsc was calibrated for temperature with pure benzophenone ( mp 48.0 c ) and caffeine ( mp 236.2 c ) , and the energy calibration was performed with indium ( mp 156.6 c , heat of fusion 28.45 j g ) . the errors on the extrapolated transition onset temperatures and enthalpy values are 95% confidence intervals ( ci ) derived from at least three measurements . isothermal calorimetry was used to derive dehydration enthalpies ( see section 10 of the supporting information ) . thermogravimetric analysis ( tga ) was carried out with a tga7 system ( perkinelmer , usa ) using pyris 2.0 software . two - point calibration of the temperature was performed with ferromagnetic materials ( alumel and ni curie - point standards , perkinelmer ) . heating rates ranging from 2 to 20 c min were applied , and dry nitrogen was used as a purge gas ( sample purge , 20 ml min ; balance purge , 40 ml min ) . raman spectra were recorded with a bruker rfs 100 raman - spectrometer ( bruker analytische messtechnik gmbh , germany ) , equipped with a nd : yag laser ( 1064 nm ) as the excitation source and a liquid - nitrogen - cooled , high sensitivity ge - detector . the spectra ( 128 scans per spectrum ) were recorded in aluminum sample holders with a laser power of 300 mw and a resolution of 2 cm . temperature conditions were adjusted with a specac ( grasebury specac limited , orpington , u.k . ) variable temperature cell and a temperature control unit . for investigating h2o / d2o exchange , the number of scans and laser power were increased to 1064 and 400 mw , respectively . samples were stored and measured in hygrostats as detailed in ref ( 66 ) and section 9 of the supporting information . principle component analysis ( pca ) , a multivariate data treatment to reduce the number of variables and provide a representation of the spectra in low dimensional space , was used to interpret changes in the raman spectra during dehydration . max normalization ( opus version 5.5 , bruker optics , ettlingen , germany ) and first derivatives were calculated using simca - p ( version 11.0 , umetrics ab , umea , sweden ) . the spectral region of 1800 to 30 cm was used for constructing the pca models . infrared spectroscopy was used to characterize the ionization state of the crystalline and amorphous db7 forms ( see section 1 of the supporting information ) . cross - polarization / magic angle spinning nmr spectra were obtained on a bruker avance iii 400 wide - bore nmr spectrometer operating at h and c frequencies of 400.131 and 100.623 mhz , respectively , and using bruker 4 mm probes . the mas rate was set to 10 khz 2 hz using a bruker mas - ii controller . h decoupling was achieved using the spinal64 decoupling sequence at a proton nutation frequency of 100 khz . spinning sidebands were suppressed using a five - pulse total sideband suppression ( toss ) sequence . a 3.4 ms linear rf power ramp was used for cross - polarization from h to c. the acquisition time was set to 34 ms , and spectra were acquired over a spectral width of 30 khz with a recycle delay of 7 s. the c chemical shifts were externally referenced ( 0.05 ppm ) to the proton - decoupled c peak of neat ( liquid ) tetramethylsilane via the high - field resonance of adamantane ( = 29.5 ppm ) . the sample temperature was regulated to 25 c in order to minimize frictional heating caused by sample spinning . the crystal16 crystallization system ( avantium , nl ) was used to determine the kinetic solubilities of forms i , ii , hya , and hy2 in an acetonitrile / water ( 1:1 ) mixture . the temperature at the point the suspension becomes a clear solution upon heating or ( at 0.1 c per minute , with the exception of form i where a heating rate of 0.3 c per minute was used to avoid transformation to form ii ) was taken as the saturation temperature of the measured sample with known concentration . to make sure that solvent - mediated transformations had not occurred during the measurements , excess solid was stirred under the same conditions , and xrpd patterns of the residual solid were recorded after reaching the highest clear point temperature derived from the solubility experiments . periodic electronic structure calculations were carried out with the castep plane wave code using the perdew burke ernzerhof ( pbe ) generalized gradient approximation ( gga ) exchange - correlation density functional and ultrasoft pseudopotentials , with the addition of a semiempirical dispersion correction , either the tkatchenko and scheffler ( ts ) model or grimme06 ( g06 ) . for further details energy difference estimates between the zwitterionic and neutral forms are not given because the pbe functional has been shown to overstabilize the proton transfer phases of ammonia monohydrate ; this systematic error should largely cancel when comparing structures with the same ionization state , as should the known limitations in modeling water water interactions . nmr shielding calculations were performed on pbe - ts optimized structural models of db7 hya using the castep nmr code and on the fly pseudopotentials . the castep computed shielding constants , calc , were converted to chemical shifts , calc , according to calc = ref calc using a reference value , ref , taken from the zero intercepts of the fits of the calculated shielding vs. experimental chemical shift plot ( castep = xexp + ref ) for hya ( section 17 of the supporting information ) . our ability to meaningfully study the structures , stability relationships , and interconversion pathways of the db7 hydrates at a molecular level relied on securing highly crystalline , phase pure samples of each form through exquisite control over crystallization , filtration , and drying . since both hy2 and hya are metastable with respect to some or all of the neat polymorphs depending on the rh , they needed to be crystallized under carefully controlled conditions , then isolated and characterized before subsequent conversion to the more stable neat forms . selecting for the metastable hydrates in crystallization required that amorphous db7 be used as the starting material to ensure that the solutions were supersaturated with respect to the hydrates and free of crystalline seeds of the neat polymorphs . even then , the filtration and drying of these materials was surprisingly tricky , with under or over drying leading to form conversions , amorphization , particle agglomeration , and mostly brittle , chunky solids . eventually , conditions were identified to selectively crystallize and recover the hydrates in highly crystalline form , each with reasonably good material handling properties . the hy2 and hya materials used to fund our experimental effort were generally composed of well - formed crystals of similar size and shape commensurate with their high degree of crystallinity , phase purity , and homogeneity ( figure 2 ) . the molecular conformation in this zwitterionic form is closely related to one of the form iii conformers ( figure s2 of the supporting information ) , but with the carboxylic acid proton moved to the piperazine n3 . thus , the protonated piperazine is intramolecularly hydrogen bonded to the carboxylate ion ( n ho ) . the db7 molecules form centrosymmetric dimers ( figure 3a ) , which are arranged in layers parallel to the bc plane . each of the two water molecules ( w1 and w2 ) forms an o ho hydrogen bond to one of the db7 carboxylate oxygens ( figure 4 ) ; the other water protons are disordered over two positions each with a site occupancy of 0.5 . w1 and w2 form infinite corrugated chains , which are interlinked by the carboxylate group of db7 ( figure 4 ) , leading to zigzag sheets ( figure 3b ) . the zigzag sheets , containing all of the strong hydrogen bonds , are interleaved in a zip motif along a. packing diagram of hy2 viewed along the crystallographic b axis ( a ) and c axis ( b ) . hydrogen bonds are denoted as green dotted lines and the centrosymmetric db7 dimer and zigzag sheet are highlighted . the main difference in the molecular conformation of db7 in hya and hy2 is a 52 rotation of the c18c19c20o2 dihedral ( figure 5a ) moving the carboxylate group from an intra- to intermolecular hydrogen bonding conformation . the conformation of db7 in hya is closely related to that in anhydrate form ii ( figure s2 of the supporting information ) . owing to the shape complementarity of the db7 conformers in both hydrates , inversion related dimers similar to those formed in hy2 are also seen in hya ( figures 3a and 5c ) . however , the hya structure also features intermolecularly hydrogen bonded r22(12 ) dimers involving strong n the hya structures of the fresh and aged crystals differ in containing 1.95 ( hya1.95 ) and 1.73 ( hya1.73 ) mol of water per mol of db7 , respectively . the loss of 0.22 mol of water on aging resulted in a volume change of 3.5% , with the c - axis change of 2.5% being the most significant . three water sites ( w13 ) water site w1 is disordered over two positions , which are so close in proximity that only one of the two positions can be occupied at any time ( figure 6a ) . the w1 sites refined to an occupancy of 46% ( w1a ) and 52% ( w1b ) for hya1.95 , and 68% ( w1a ) and 30% ( w1b ) for hya1.73 . the site occupancy from the refinement of w2 was similar to that of w1b , i.e. , 52% for hya1.95 and 30% for hya1.73 . thus , if w2 is present , w1b is occupied , and if not , w1a is likely to be occupied ( figure 6b ) . water site w3 is also disordered over two positions , which are too close to be occupied at any one time . in hya , one of the w3 water protons is located on a special position , a 2-fold axis , so a maximum of 0.5 mol water per mol db7 would be present at full occupancy . refinement of the w3 waters revealed nearly full occupancy , 45% for both hya1.95 and hya1.73 , across the symmetry related sites . as in hy2 , the hya water molecules form strong hydrogen bonds to db7 and one another . w1b can form two ow1ho interactions , involving carboxylate o1 and the w2 oxygen ( figure 6 ) . w2 participates in four hydrogen bonds , as a donor to the carboxylate o2 and the oxepine n and an acceptor to w1b and w3 . together with the db7 carboxylate oxygens , w1b and w2 form an extended ring motif ( figure 6 ) the third water molecule , w3 , is hydrogen bonded to carboxylate o2 and w2 ( when present ) , linking the hydrogen bonded rings along the c axis , the axis most affected in length by the hya water content . the 1d hydrogen bonding network of hya is a marked contrast to the 2d sheets of water and db7 molecules found in hy2 . in hya the 1d hydrogen bonded chains ( figure 5c ) are close packed in the second and third dimensions through centrosymmetric db7 dimers . ( a ) overlay of db7 hy2 ( red ) and hya1.95 ( blue ) conformations ( rmsd1 = 0.38 ) . ( c ) packing diagram of hya1.95 viewed along c. centrosymmetric dimer , water position w2 , and 1d building block are highlighted ; w1 and w3 are obscured by the carboxylate groups in this view . the distinct water sites are coded in different colors : w1a , rose ; w1b , red ; w2 , blue ; w3 , green . note that only one of the two differently colored w3 orientations can be occupied as the two positions are related by a 2-fold axis at a proton position . ( a ) showing the time - averaged water occupancy and ( b ) one of many possible instantaneous snapshots of the water positions in hya in the range of occupancies seen in the two crystals . both hydrates are zwitterionic , but the n3h in each structure is close to one of the carboxylate oxygen atoms . computationally , we can confirm the zwitterionic assignment by a method used to distinguish between salts and cocrystals . hypothetical neutral hydrates are constructed from chy2 ( section 3.4.2 ) and chya2.0 ( section 3.4.3 ) by moving the acidic proton to the nearby carboxylate oxygen and adjusting the bond lengths to give a cis - carboxylic acid for hya and the less common trans - acid conformation for hy2 . full optimization of both charge neutral structures results in the acidic proton moving back to reform the observed zwitterionic hydrate structures , showing that both hydrates are more stable in zwitterionic than neutral form with no appreciable barrier to proton migration ( tables s5 and s7 of the supporting information ) . the converse was also found : full optimization of the corresponding hypothetical zwitterionic versions of forms i iii results in the proton moving to give the observed locally neutral structures . the hydration and dehydration behavior of the db7 crystal forms was investigated between 0% and 95% rh at 25 c ( figure 7 ) . two of the anhydrates , forms i ( figure 7a ) and ii ( figure 7b ) , show almost no water uptake ( form i < 0.32% and form ii < 0.04% ) up to 95% rh . in contrast , anhydrate form iii undergoes a phase transformation with a mass increase corresponding to 2 mol of water per mol of db7 at rh values > 70% ( figure 7c ) . the product hydrate phase , confirmed to be hy2 by xrpd , shows good rh stability with a marginal weight loss between 95 and 25% rh . hysteresis was observed in the form iii / hy2 isotherm despite the lengthy equilibration times ( up to 48 h ) at each rh . the profile of the moisture sorption / desorption isotherms for form iii hy2 with steps accompanied by phase changes , is typical of a stoichiometric hydrate . in contrast , highly crystalline hya shows the typical ( de)sorption isotherms of a nonstoichiometric hydrate ( figure 7d , see also figure s3 of the supporting information ) as the mass gradually changes depending on the humidity during sorption and desorption . the lowest water content measured by tga ( figure 7d ) for hya equilibrated at 0% rh corresponded to 1.29 mol of water per mol of db7 . the maximum water vapor uptake into the hya structure was estimated to be approximately 1.85 mol of water per mol of db7 at 95% rh . gravimetric moisture sorption and desorption curves of db7 solid forms at 25 c : ( a ) form i , ( b ) form ii , ( c ) form iii / hy2 , and ( d ) hya . the gray circles represent data points that fulfill the preset equilibrium conditions ( see experimental section ) , whereas the crosses mark measurement values that did not reach equilibrium within the allowed time limit ( 48 h ) . the automated gravimetric moisture sorption / desorption analysis of hy2 and hya was complemented with long - term drying experiments at 0% rh ( storage over p2o5 ) . dehydration of hy2 was observed immediately at this condition , and form iii was determined to be a kinetic dehydration product , with further transformation to the more stable forms i and ii occurring within 2 weeks . hya could be similarly dehydrated at 0% rh ; however , loss of the last strongly bound 1.3 waters of crystallization from this hydrate leads to a structural collapse to mainly amorphous db7 . the amorphous form has limited stability at 0% rh as there was evidence of forms i and ii nucleating in small amounts during the low rh drying . the gravimetric moisture sorption / desorption studies ( figure 7 ) were correlated with structural changes to hy2 and hya using variable - humidity xrpd at 30 c ( figures 8 and 9 ) . in agreement with the results obtained in figure 7c , distinct changes occur in the xrpd pattern of the hy2 sample at rh values < 30% ( figure 7a ) . the disappearance of high intensity hy2 peaks ( e.g. , at 6.82 , 10.62 , 16.37 , 16.71 , and 17.57 2 ) and appearance of form iii peaks ( e.g. , 7.33 , 12.68 , and 14.81 2 ) indicate a major change in the crystal structure ( figure 8c f ) on loss of the waters of crystallization . the unusual increase in volume / z upon dehydration ( figure 8b ) can be attributed to the less efficient packing of db7 with itself and ( a ) moisture - dependent xrpd measurements showing the dehydration of hy2 to form iii . peak positions marked with * correspond to hy2 impurities ( dehydration not complete due to the shorter time intervals of 12 h and 10% rh steps in contrast to 48 h and 5% steps in figure 7c ) . ( b ) moisture - dependent changes in the cell volume per asymmetric unit for the transformation hy2 to form iii . ( c , d ) molecular packing of db7 in hy2 and ( e , f ) db7 in form iii . disorder of the methyl group in form iii is indicated with different shades of gray . in contrast to hy2 , there is no significant change in the crystal lattice of hya with varying rh ( figure 9b ) apart from anisotropic expansion of the network to accommodate a variable amount of water . the anisotropy of the lattice expansion was quantified by indexation and rietveld refinement of the hya xrpd patterns recorded at different rh values ( figure 9a ) . using the hya1.95 structure as a starting model , rietveld refinement found the c axis to be affected the most by changes in the water content , in agreement with the scxrd experiments , but over a wider range of compositions ( table s2 of the supporting information ) . structural refinement also revealed that the water occupancy of w3 stays close to 50% ( i.e. , full ) independent of the rh , suggesting that the w3 position is integral to the hya structure . however , the occupancy parameters for w1 and w2 collectively decreased at progressively lower rh values . the w1w2 distance was also found to depend on the hydration state . at moderate rhs ( 2080% ) a plateau is reached with the modeled w1w2 distance being too short for two distinct sites , suggesting either water mobility or static disorder ( figure 9c ) . a further distinct change in the w1w2 distance is seen below 25% rh , the moisture range where in addition to continued diffraction peak shifting , selected reflections , in particular ( 002 ) , are broadened . the lowest hya water content observed in the rh - dependent xrpd experiments of hya was 1.32 mol of water per mol of db7 ( measured with tga ) . ( b , c ) changes in the hya cell volume , length of the c - axis , fractional occupancies of w1 + w2 and w3 oxygens , and distance between w1 and w2 oxygens on lowering the rh ( desorption ) . water position w1 was modeled as one site only . 100% rh data points given in ( b ) and ( c ) were derived from slurry experiments in water . ( d f ) packing diagrams of hya highlighting the water oxygen positions at different hya hydration states derived from single crystal x - ray data ( d , e ) and rietveld refinement of powder x - ray data ( f ) . red lines between water oxygens indicate the disordered sites for w3 and close proximity of the w1 and w2 positions in ( f ) . w1a and the moisture - dependent xrpd experiments were extended to water activity = 1 ( 100% rh ) by measuring a thick hya slurry in water . the powder pattern of the 100% rh hya sample refined to 1.96 mol equiv of water per db7 , comparable to hya1.95 observed ( by scxrd ) for a freshly crystallized single crystal . while a change in water content and unit cell volume was to be expected on increasing the rh from 90 to 100% , the sharp uptake of 0.10 mol water per mol db7 and unit cell volume increase of 0.5% ( figure 9b ) was unexpected and presumably reflects the slow hydration kinetics of hya in the solid state ( table s2 of the supporting information ) . an ordered hy2 structural model was derived from the experimental crystal structure by replacing the two disordered water molecules related by a 21 screw axis with one molecule in each orientation ( section 12 of the supporting information ) . this lower symmetry hy2 structure ( figure s10b ) was lattice energy minimized to give the computational model of the hy2 structure , denoted chy2 . the geometry optimizations of four hypothetical hydrate and framework structures derived from chy2 ( table s5 of the supporting information ) showed that proton transfer to a neutral db7 molecule occurred when there was no water hydrogen bonding to the carboxylate oxygen involved in the n although the layer structure ( figure 2 ) suggests that it might be possible to remove both w1 and w2 , doing this computationally results in proton transfer and an unstable , low density isomorphic desolvate with no strong hydrogen bonds between db7 molecules . partial removal of the w1 molecule gave hypothetical zwitterionic sesquihydrate ( chy21.5(0.5w1+w2 ) ) and monohydrate ( chy21.0(w2 ) ) structures , which differed in lattice energy from hy2 by 55 and 107 kj mol , respectively ( table s5 of the supporting information , averaged over the two dispersion models ) . estimating whether hy2 could undergo a phase transition to an isostructural sesqui- or monohydrate requires comparison of the lattice energies of the model hydrates with that of ice ( section 16 of the supporting information ) . this gives an unfavorable phase transition energy of more than 30 kj mol for each water removed . thus , the computational thermodynamic estimates show that a stepwise dehydration of the hy2 structure , preserving the hy2 db7 framework structure ( i.e. , a structurally related sesquihydrate or monohydrate as a transient , intermediate hydrate along the dehydration pathway ) , is highly unlikely . this is consistent with hy2 showing a single stepwise weight loss by moisture sorption analysis ( figure 7c ) and tga ( section 3.5.1 ) to water free db7 . the two hya structures show a similar nearly full occupancy of w1a + w1b ( 98% ) and w3 ( 45% ) , but differ in the w2 partial occupancy ( 52% or 30% ) . to form an ordered computational model of hya1.95 , the crystallographic 1.95 water molecules per db7 molecule had to be increased to 2 by modeling the 98% ( 46% w1a + 52% w1b ) , 52% and 45% occupancy ratios for w1 , w2 , and w3 as 100% , 50% , and 50% , respectively . all symmetry was removed from the c2/c , z = 1 structure of hya , resulting in a p1 , z = 4 cell in which the four w1 sites are independently occupied and half of the w2 and w3 sites are occupied ( section 13 of the supporting information ) . all possibilities leading to the described occupancy ratios were used as starting structures and the lowest energy structure after geometry optimization was used as chya2.0 hereafter . similarly , for the hya1.73 structure the occupancies were modeled as 100% , 25% , and 50% for w1 , w2 , and w3 giving a model chya1.75(2 ) ( table s7 of the supporting information ) , which agreed with the experimental structure . the confirmation that the ordered cell models chya2.0 and chya1.75 were reasonable representations of the disordered structures is given in section 15 of the supporting information . a set of hypothetical hya framework structures can be derived from the experimental structure , ranging from a hemipentahydrate ( chya2.5 , 2.5 mol of water per mol db7 at full w1 and w2 occupancy ) to the water free framework ( table s7 of the supporting information ) . the lattice energy of chya2.5 was higher ( less stable ) by about 2 kj mol than the sum of the lattice energies of chya2.0 and 0.5 mol of ice xi , implying that additional water to fully occupy the w2 site would destabilize the hya structure . computational dehydration of chya2.0 to chya1.75 , chya1.5 , and chya1.25 produces a range of possible structures , which are very close in energy ( table s7 of the supporting information ) . removal of water from hya was calculated to be energetically quite feasible and to require considerably less energy than removing comparable amounts of water from hy2 ( tables s5 and s7 ) . the calculated isomorphic hya desolvate ( chya0.0 ) also undergoes a proton transfer to the neutral molecule on optimization ( table s7 of the supporting information ) . in fact , proton transfer from db7 to db7 was observed when the carboxylate of the n hor22(12 ) dimer ( figure 5b ) was not also acting as an acceptor for an ow ho hydrogen bond . thus , the hydrogen bonded water molecules ( w1 , w2 , or w3 ) appear to be essential for zwitterion formation . since chya0.0 is estimated to be over 25 kj mol less stable than form ii , an isomorphic desolvate of hya containing zwitterionic or neutral db7 molecules is unlikely to form . this is consistent with a destructive dehydration mechanism on loss of all waters of crystallization . the suggestion of water mobility / disorder from the diffraction experiments and static lattice energy modeling led to the use of c cp / mas nmr spectroscopy to characterize hya as a function of water content . except for slight peak intensity variations , the ssnmr spectra of hya samples equilibrated between 22 and 90% rh were essentially identical ( figure 10a ; figure s5 of the supporting information ) . since partial occupancy of the hya water sites over this rh range would necessarily result in a distribution of hydrogen bonding arrangements ( figure 6 ) , the observation of a single c resonance for each carbon atom in db7 shows that the different hydrogen bonding environments are effectively averaged as the waters of crystallization move in and out of the water sites in the hya crystal structure . thus , ssnmr shows that the water of crystallization is highly mobile over a wide range of moderate and high rh . after equilibrating the sample at rh 11% ( figure 10b ) , peak splitting for carbon atoms c20 , c5 , and c17 and broadening of several c nmr resonances were seen across the ssnmr spectrum of hya . the additional peaks were not due to forms i iii or hy2 ( ref ( 47 ) and figure s6 of the supporting information ) phase impurities , as there is no match to their chemical shifts . instead , the emergence of low intensity peaks was ascribed to restricted water mobility in the contracted hya cell , which would also explain the diffraction peak broadening seen in figure 9a . support for this interpretation of the low rh ssnmr spectrum ( figure 10 ) comes from dft shielding calculations on 25 variably hydrated chya structures ( section 17 of the supporting information ) , which generated 80 c nmr spectra of db7 in different hya water environments . these showed that the chemical shifts of c20 , c5 , c18 , and c17 , because of their proximity to the water molecules , were particularly sensitive to hydration state and water positions , varying by 35 ppm ( table s11 of the supporting information ) . since the regions in the ssnmr spectrum shown computationally to be most sensitive to water in the hya structure coincide with those where peak splitting is observed at low rh ( figure 10 ) , we can account for the peak splitting as a manifestation of reduced water mobility on the ssnmr time scale . ( a , b ) representative experimental hya spectra differing in water content and ( c h ) six of 80 nmr - castep computed c cp / mas nmr spectra . ( c , d ) the two db7 environments observed in the ordered p1 chya2.0 model and ( e h ) the four environments observed in the ordered p1 chya1.25(13 ) model , which correspond to 0.75 w1 , 0.25 w2 , and 0.25 w3 . in contrast to hya , the ssnmr spectra of hy2 ( figure s6 of the supporting information ) show a distinct phase change to form iii in this case between 11 and 22% rh , in good agreement with the moisture sorption ( figure 7c ) and xrpd data ( figure 8a , b ) . hy2 and hya were exposed to deuterium oxide vapor ( 98% and 11% rh ) and characterized by raman spectroscopy at different time points ( figure 11 ) to investigate water dynamics in the two hydrates . the wavenumber region 34002800 cm is composed of peaks arising from (o h ) , (n h ) , and (c h ) stretching vibrations ; (o d ) stretching modes of d2o are seen between 2600 and 2300 cm . the emergence of o d peaks in the spectra of both hydrates on exposure to deuterium oxide vapor confirms that d2o displaces h2o in both crystal structures . thus , not only is the water of crystallization in hya mobile , but water exchange is also observed for the stoichiometric hydrate , hy2 . water diffusion in and out of the two crystal structures is rapid and quantitative , as derived from the fact that the raman spectra after exposure to d2o for 72 h were indistinguishable from fully deuterated hy2 and hya . db7 also has an ammonium proton that could exchange a hydrogen for a deuterium . to see whether the ammonium group had undergone exchange , the hydrate samples that had been exposed to d2o vapor were dehydrated and compared to the raman spectra of the neat forms . the comparison showed that within 72 h of d2o vapor exposure the ammonium group was not affected at all . raman spectra of db7 hy2 ( a ) and hya ( b , c ) as a function of time of exposure to d2o vapor , 98% rh ( a , b ) and 11% rh ( c ) . d stretching vibrations emerge over the course of 72 h. the tga curve of hy2 ( figure 12a ) shows a one - step mass loss of 8.67 0.05% , corresponding to exactly two mole equivalents of water . dehydration starts immediately under the dry conditions of a nitrogen purge . on removing the water of crystallization , the clear hy2 crystals turn opaque , but maintain their shape ( pseudomorphosis ) . slow heating dsc experiments of hy2 ( figure 12a ) using pinhole lids show four events : ( 1 ) dehydration of hy2 to form iii takes place below 45 c . ( 2 ) at 95 c there is an exothermic phase transformation of form iii mostly to form ii , with minor amounts of form i present . the measured enthalpy of transition is in agreement with the value derived using pure form iii as a starting material ( trshiii ii = 13.3 0.2 kj mol ) . ( 3 ) upon further heating , melting of the minor form i component is observed at an onset temperature of 176.0 0.4 c ( small peak ) . ( 4 ) at an onset temperature of 180.9 0.2 c , the form ii melts . by increasing the heating rate used in the hy2 dsc experiments to 5 c min or more ( figure 12a : 5 , 10 , and 50 c min ) , an additional thermal event is observed above 118 c corresponding to the inhomogeneous melting of form iii . melting is followed by the recrystallization of mainly form ii , then the melting of forms i and ii. by embedding the hy2 crystals in high viscosity silicone oil or using hermetically sealed dsc pans ( figure 12a ) , the incongruent melting of hy2 ( peritectic decomposition ) is observed at 89.8 1.3 c . compared to hy2 , higher temperatures are required to fully dehydrate hya . by tga , a hya sample pre - equilibrated at 75% rh ( stored over a saturated nacl solution ) showed a mass loss of 7.61 0.04% , corresponding to 1.74 mol equivalents of water ( figure 12b ) . toward the end of the dehydration process a loss of birefringence is observed in polarized light , indicating that the strongly bound water is released by a process that leads to a loss of crystallinity . the generation of mainly amorphous db7 with form i and ii impurities was independently confirmed by xrpd and raman spectroscopy . the loss of crystallinity upon dehydrating hya was also supported by dsc , based on the appearance of an exothermic event immediately following the dehydration endotherm . upon further heating , the few crystal nuclei produced at lower temperatures ( during dehydration ) grew into a mixture of forms i and ii , evidenced by the final two melting events . the lower the dehydration temperature , the less form ii was observed . dsc traces of hya collected at faster heating rates ( 10 c min ) show an overlap of dehydration and peritectic decomposition at 99 c . the peritectic decomposition of hya , measured in hermetically sealed dsc pans , is observed at 98.9 0.9 c . differential scanning calorimetry ( dsc ) and thermogravimetric analysis ( tga ) thermograms of ( a ) hy2 and ( b ) hya . tga curves were recorded in open pans at a heating rate of 5 c min . raman spectra were recorded for hy2 and hya every 2 to 5 c from 25 to 130 c , then subjected to pca to track the solid state transformations ( figure 13b , c ) by comparison to reference spectra of the db7 forms ( figure 13a ) . the raman spectra recorded throughout the hy2 dehydration appear as three distinct clusters , corresponding to hy2 , form iii , and form ii with form i impurities ; in between the clusters ( 50 , 100 , and 102 c , figure 13b ) are mixed phases . these results show that form iii is a distinct intermediate phase along the thermally induced dehydration pathway of hy2 . ( the close proximity of the dehydration product to phase pure form ii relative to phase pure form i in figure 13b shows that form ii is the main component , and the tightness of the final product cluster shows that the form i impurities did not transform to form ii . ) this dehydration pathway is in agreement with the dsc and tga results in figure 12a . the dehydration of hy2 does not go via an appreciably stable amorphous phase or phase pure form i because the triangles representing form i and amorphous db7 are not located on the dehydration pathway . the structural changes accompanying the dehydration of hya to a form i and ii mixture could be described with three pcs , but a continuous transformation pathway is evident from two of them , figure 13c . as the small structural variation within hya exhibiting variable water content between 25 and 62 c is apparent by pc2 , pc1 shows the transformation from the hya crystal lattice ( 70 c ) through the amorphous state ( 82 c ) to the form i / ii mixture ( 86130 c ) ( figure 13c ) . ( b , c ) raman principal component analysis ( pca ) plots for the first and second principal component ( pc1 and pc2 ) of db7 forms occurring during dehydration of hy2 ( b ) and hya ( c ) . each triangle corresponds to a raman spectrum : black , recorded during dehydration and used for constructing the pca models ; gray , reference spectra for forms i and ii and amorphous db7 . numbers correspond to the temperature at which each spectrum was recorded ; black ellipse corresponds to the 95% hotelling t2 , and areas of distinct db7 solid forms are encircled . the % -values on the axes indicate the data variance modeled by the respective pc . form iii was obtained for the first time following the salt disproportionation of db7 phosphate at ph 6 . only after we were unable to directly crystallize form iii from solution and systematically studied the dehydration mechanism of hy2 was it evident that hy2 was the kinetic phosphate salt disproportionation product and necessary intermediate phase for obtaining form iii . structural analysis has shown that the molecular conformation of db7 in hy2 with intramolecular hydrogen bonds seems to predispose it to dehydration to form iii , the only anhydrate , which also features intramolecular hydrogen bonding . to the best of our knowledge , following our observation of batch - to - batch variability in the disorder ratio of the dimethylpropionic acid side chain in form iii , we performed systematic drying studies of hy2 to estimate the effects of rh and temperature on the level of disorder present in the form iii dehydration product and to seek conditions that would select for the ordered structures . the disorder ratio was quantified from the intensities of the dimethylpropionic acid methyl c ssnmr peaks ( figure 14a , table s3 of the supporting information ) . unfortunately , despite the wide range of drying conditions surveyed , we were not successful in preparing either an ordered form iii sample or one having the minor conformation dominating . ( a ) c cp / mas nmr spectra of db7 form iii produced by dehydrating hy2 under a variety of conditions ( defined in the supporting information , table s3 ) . highlighted are the c18 and c21 peaks characterizing the structural disorder of the dimethyl propionic acid side chain . ( b ) dsc thermograms of db7 form iii lots n2/0/rt/24 and n2/0/5/48 measured at a heating rate of 1 c min . lot n2/0/rt/24 was prepared by drying hy2 under n2 purge at room temperature for 24 h and lot n2/0/5/48 under n2 purge at 5 c for 48 h. subsequent dsc investigations ( figure 14b ) were conducted to examine whether the disorder ratio in form iii influences its solid form transformation to forms ii and/or i. although the outcomes were somewhat variable , form iii samples obtained by drying hy2 under a variety of conditions ( table s3 of the supporting information ) ranged from having only small amounts of form i to as much as 90% form i after the solid state transformation . importantly , the heat induced phase transformation does not seem to be related to the disorder in form iii . form iii samples n2/0/5/48 and n2/0/rt/24 , for example , show a similar level of disorder in their c ssnmr spectra , but differ substantially in their thermal behavior ( figure 14b ) . thus , while the dehydration reaction of hy2 always produces the two different conformations in form iii , small differences in the phase purity and sample history also influence the outcome of the solid form transformation to forms i and/or ii. the db7 hydrates and db7 anhydrates showed appreciable kinetic stability in the solid state based on the gravimetric moisture sorption / desorption analysis . however , these forms were not sufficiently stable in suspensions to obtain equilibrium solubility data of interest for quantitatively establishing their thermodynamic stability relationships . the stability order of the two stable anhydrate polymorphs forms i and ii and the two hydrate forms was therefore qualitatively estimated as a function of temperature from kinetic solubility measurements in acetonitrile / water ( 1:1 ) . this solvent mixture was chosen because the db7 and db7 solid forms were sufficiently soluble to meaningfully estimate their relative solubility using the crystal16 parallel reactor system . unfortunately , the fast transformation kinetics of form iii to form ii in stirred solvent prevented any measurement of the kinetic solubility of form iii . the acetonitrile / water solubility data for forms i and ii ( figure 15a ) agree with the monotropic relationship previously derived from solubility measurements in methyl isobutyl ketone and dsc experiments , as the kinetic solubility data plotted in vant hoff form showed no signs of convergence or crossover below the db7 melting temperature ( figure 15b ) . hya is less soluble than hy2 in acetonitrile / water ( water activity , aw , of 1:1 mixture is 0.9 at 25 c , figure 15a ) , with the vant hoff plots further suggesting that hya is the thermodynamically most stable db7 hydrate at this water activity below the hydrates melting points . the zwitterionic hydrates are much more soluble than the charge neutral neat forms at all measured temperatures , but their relative solubility is clearly decreasing with decreasing temperature ( figure 15b ) . ( a ) solubility ( mole fraction ) of db7 forms i , ii , hy2 , and hya in acetonitrile / water ( 1:1 , aw 0.9 ) as a function of temperature and ( b ) vant hoff plot of the molar solubility as a function of temperature . the critical water activity defining the crossover in thermodynamic stability of the db7 hydrate db7 anhydrate pairs could not be determined from the solubility measurements at a single water activity ; however , these results did reveal that both forms i and ii are more stable than hy2 and hya at water activities up to at least 0.9 over the investigated temperature range . above aw = 0.9 , the rt thermodynamic stability order of the db7/db7 forms was investigated through slurry experiments in methanol / water mixtures ( section 8 of the supporting information ) . db7 hy2 and hya were separately added to methanol / water mixtures ( aw = 0.9 , 0.95 , and 1.0 ) and stirred for 21 days . slurries in mixtures of aw = 0.9 and 0.95 resulted in form ii , while the pure water slurry yielded hya . in showing that form ii is more stable at aw = 0.95 and hya is more stable in pure water ( aw = 1.0 , ph 5.9 ) , the critical water activity at which the solubility order of form ii/hya is reversed was more precisely bracketed between 0.95 and 1.0 . the hy2 to hya transformation in water did not allow us to estimate the hy2 aw dependency . therefore , we used gravimetric sorption / desorption experiments to estimate the influence of moisture . the desorption curve is similar to that of phase pure hy2 ( figure 7c ) . the water is released at rh < 25% , and the product contains forms i and ii from the spiking and form iii as the dehydration product . upon subsequently increasing the rh of the mixed anhydrate sample , hydration of form iii to hy2 is observed as well as a transformation to form ii ( figure 16 , 2 ) . the latter , which is indicated by the decrease in mass despite increasing the humidity at rh values > 85% , is consistent with the greater stability ( lower solubility ) of form ii than hy2 observed in 1:1 acetonitrile / water ( aw 0.9 ) , figure 15 . gravimetric moisture sorption and desorption curve of db7 hy2 spiked with forms i and ii at 25 c . the gray circles are data points that fulfill the preset equilibrium conditions ( see experimental section ) , whereas the crosses mark measurement values that did not reach the equilibrium within the allowed time limit ( 48 h ) . the forms noted in parentheses were identified as minor components by xrpd . to investigate the influence of ph on the db7/db7 form stability , slurry experiments were conducted in aqueous buffer solutions at 25 c . solvent mediated transformations ( smt ) starting from hy2/form iii showed that hya is obtained if ph 6 and anhydrous form ii if ph 5 . in contrast , the phosphate salt of db7 was shown to disproportionate in aqueous media , causing either hy2 ( ph 67.8 ) or form ii ( ph 2.85 ) to nucleate . the fact that hy2 converted to hya in the higher ph smt experiments confirms that hy2 was a kinetic disproportionation product at 25 c . thus , hya is the thermodynamically most stable db7 form in water , but this stability relationship has been established only at ph 5.9 ( from water slurry experiments ) . db7 has a rich solid form landscape composed of three neat polymorphs , two hydrates , three solvates , and an amorphous phase . with the goal of exploring the thermodynamic stability relationships between the hydrates and anhydrates , as well as their interconversion pathways , we have established through a combination of diverse experimental techniques and computational chemistry that db7 form appearance and stability depend on temperature , water activity , and ph . form ii is monotropically more stable than forms i , iii , and hy2 and usually more stable than hya . hya is the most stable db7 crystal form in saturated water solutions , but only at ph 5.9 . characterizing the structure stability relationships of the db7 crystal forms was complicated not only by the difficulty in isolating highly crystalline , phase pure samples of the metastable forms ( section 3.1 ) , but also the slow rate at which hya and hy2/form iii materials reached their equilibrium moisture content . equilibrium water contents were attainable for hya using extended equilibration times at most rhs , although complete rehydration to full crystallographic water occupancy was never achieved even in aqueous slurries . in contrast , the reversible dehydration of hy2 to form iii mediated by water vapor was readily apparent from a quick measurement of the moisture sorption desorption isotherms of either form , yet extended equilibration times did little to minimize the hysteresis ( figure 7c ) caused by slow transformation kinetics . the slow interconversion of hy2 and form iii in the solid state at moderate rhs frustrated attempts to establish the critical water activity relating these metastable forms . two distinctly different moisture sorption behaviors were established for hy2 and hya through detailed characterization of highly crystalline , phase pure samples as a function of water vapor pressure . curiously , our initial attempts to rationalize the different hygroscopicity of the db7 hydrates in terms of structural features ( solvent pockets , channels ) common to stoichiometric and nonstoichiometric hydrates yielded anything but expected results . figure 17 shows the void space ( default probe radius = 1.2 ) created by removing the waters of crystallization from the hya1.95 , hya1.73 , hya1.32 , and hy2 structures . counterintuitively , the water accessible voids in nonstoichiometric hya appear to be isolated from one another ( figure 17a c ) , while water channels are clearly seen running zigzag along the crystallographic b - axis in the stoichiometric hydrate , hy2 ( figure 17d ) . solvent accessible voids containing waters of crystallization in ( a ) hya1.95 , ( b ) hya1.73 , ( c ) hya1.32 , and ( d ) hy2 , calculated using a 1.2 probe radius . cursory inspection of the water channels in hy2 and the proximity of solvent pockets in hya suggests that the direction of water ingress / egress is likely to be along the b - axis in each structure . however , the 100 k structures individually do not reveal how either hydrate framework would respond to perturbations in rh , particularly at crystallization processing and storage relevant temperatures . water vapor sorption isotherms ( figure 7 ) , however , show that the db7/water stoichiometry is retained across a wide rh range for hy2 and that a fraction of the water in hya is adventitious . yet they reveal little as to how water is held in each crystal structure or the phase transformations that accompany dehydration and rehydration . indeed , only through a combination of experiment and computation were we able to paint a molecular level picture of two very different rh stability profiles and ( de)hydration mechanisms for hy2 and hya . nonstoichiometric hya retains between 1.29 and 1.95 waters of crystallization , all of which reside in the same solvent pockets in the crystal structure ( figure 17a c ) and are exchangeable ( figure 11b , c ) . the conspicuous absence of continuous water channels in hya suggests that for nonstoichiometric hydration to occur , cooperative movement of the carboxylate and dibenzoxepine groups ( figures 9d f and 17b c ) must temporarily open up diffusion pathways , similar to that seen in -cyclodextrin ( -cd ) hydrate and ciprofloxacin . water diffusion in and out of discrete crystallographic sites gives rise to varying disorder and occupancy in the diffraction time - averaged hya crystal structures , but is sufficiently rapid , at least under moderate to high rh conditions , that single , averaged db7c resonances are seen in the ssnmr spectra ( figure 10a ) . below 11% rh , water mobility is observably decreased with the resonances of c nuclei that are in close proximity to the partially occupied water sites showing splitting ( figure 10b ) . the decrease in water mobility witnessed on the ssnmr time scale at low rh is presumably caused by anisotropic contraction of the hya lattice as water is continuously lost from the crystal . in fact , rietveld refinement of the hya xrpd patterns collected across a wide rh range confirmed that the hya lattice essentially breathes to accommodate varying amounts of water with smooth changes in the cell volume and shifting of the water sites / occupancies ( figure 9 ) . void space analysis of the hya framework structures derived from hya1.95 , hya1.73 , and hya1.32 showed that as the unit cell contracts at lower hydration states ( figure 17a c ) , the solvent accessible voids decrease from 10.2% in hya1.95 to 4.2% in hya1.32 . isostructural dehydration modeling , in showing the small energy penalty that is incurred to attain full water site occupancy ( hemipentahydrate stoichiometry ) and the appreciably larger one for decreasing the water content below 1.25 mol equiv in the hya crystal structure , helped to rationalize the db7/water stoichiometries that were experimentally accessible . in the end , elevated temperatures or long - time storage over p2o5 ( 0% rh ) was required to overcome the activation energy for removing the final 1.3 equiv of water from the contracted hya structure . however , with no clear path to an appreciably stable neat polymorph , dehydration led to structural collapse as first observed by dsc ( figure 12b ) , then later confirmed by raman spectroscopy ( figure 13c ) . in contrast to hya , hy2 retains its full complement of water over a large rh range at 25 c ( figure 7c ) . stoichiometric hydration may not have been anticipated based on the location of water molecules in open channels in the hy2 crystal structure , but it was suggested by computational dehydration modeling ( section 3.4.2 ) . in revealing a significant energy penalty incurred by removing water molecules from this hydrate , lattice energy calculations for hy2 and hypothetical lower hydrate and framework structures derived from hy2 suggested that even partial dehydration to a lower isostructural hydrate would be highly unlikely . the inability to dehydrate hy2 ( without inducing a phase change ) can not be taken to mean that water exchange in this channel hydrate is not possible , however . in fact , water mobility was found to be similar in hy2 and hya by raman using h2o / d2o exchange ( figure 11a , for details see section 9 of the supporting information ) . that water is able to rapidly enter and leave the open channels in hy2 naturally raises the question : why is the stoichiometry of this hydrate conserved over such a wide rh range ? the answer lies in how the hy2 crystal structure responds to the loss of water . in this case , the hy2 framework , apparently incapable of making the type of adjustment that hya makes to accommodate appreciably lower amounts of water , experiences a significant increase in free volume on dehydration . the decreased packing efficiency , along with the loss of strong intermolecular waterdb7 hydrogen bonds , is considerably destabilizing . as a result , hy2 retains its full complement of water under most rh conditions at ambient temperature . with even modest increases in temperature ( figure 13 ) or decreases in rh ( figure 7c ) , however , hy2 gives way exclusively to a neat form ( form iii ) . the path to form iii , a neat polymorph that is more efficiently packed than the isomorphic dehydrate , is conceivably paved by the db7 conformations that allow intramolecular hydrogen bonding seen in the parent hy2 structure to be conserved ( after proton transfer ) . some of the propionic acid side chains in db7 undergo a significant conformational change on loss of water from hy2 , leading to a disordered form iii structure . this change probably accounts for the large rh range of coexistence , i.e. , hysteresis in the moisture sorption efficient and reversible nonstoichiometric hydration may be rationalized , and in favorable cases ( e.g. , cromolyn sodium , amg 222 tosylate , and pfizer s glycogen phosphorylase inhibitor ) , anticipated for open channel structures ; however , the hydrates of db7 highlight the dangers of inferring stoichiometric vs nonstoichiometric hydration from general structural features . the moisture sorption properties of hya may be unconventional , but they are not unprecedented . other carefully studied systems , such as topotecan hcl , paroxetine hcl form ii , gsk s apoa-1 up - regulator , ciprofloxacin , thiamine hcl , and -cd , show a similar ability to equilibrate with the humidity of their surroundings despite lacking continuous channels . however , hy2 is unusual in its ability to retain stoichiometric amounts of water in open water channels over such a large rh range . clearly , with neither hydrate behaving according to the conventional classifications based on either structure or absorption isotherms , jumping to conclusions about the hydration behavior from a few hasty experiments would have been very misleading . having a molecular level understanding of how water vapor is sorbed in hygroscopic materials and the risks that water uptake poses to the physical and chemical stability of a pharmaceutical product is invaluable , as some sources of water uptake are more manageable than others , careful evaluation of the stoichiometry , stability relationships , and transformation pathways of pharmaceutical hydrates should inform the solid form screening and selection process . nonstoichiometric hydration , for example , is an intrinsic and generally undesirable property of a solid form that , once established , might prompt further solid form screening and ultimately would have to be managed in the event that an alternate , nonhygroscopic form is not found . however , water uptake in the amorphous components of poorly crystalline materials is oftentimes correctable with improvements in crystallinity that are generally realized during crystallization process optimization . hygroscopicity is , of course , but one of many considerations taken in the selection of a crystal form for a commercial drug product . perhaps more important to delivering a drug safely and efficaciously are the solubility and stability , both thermodynamic and kinetic , of the solid - state form . as metastable forms of the 5-ht2a and h1 inverse agonist , hy2 and hya offer the advantage of generally higher solubility , which might be desirable for ensuring rapid oral delivery of an insomnia medication . however , from a manufacturability and control perspective , the added risk of form conversions established in our assessment of the structure stability relationships would have to be carefully weighed in the selection of either of these high energy forms for a commercial drug product . in this case , the difficulties we encountered with producing the db7 hydrates at small scale and avoiding conversions to the more stable neat forms prompted further evaluation of salt and cocrystal options to determine whether these were better suited for enabling a drug product . the computational modeling of static structures derived from the time and spatially averaged crystal structures supports the interpretation of the experimental results and accounts for the variability in disorder and water mobility shown by ssnmr and raman h2o / d2o exchange . however , current state - of - the - art calculations still struggle to obtain sufficient accuracy for the small energy differences involved and can not include the important effects of water activity or temperature , let alone proton - transfer energies . ideally , we would wish to simulate the nonstoichiometric hya at ambient temperature to confirm how the water moves and determine the correlated motions within the db7 framework ( cf . apoa-1 up - regulator ) . nonetheless , by combining such a range of experimental and appropriate computational work to get a coherent molecular understanding of this complex hydrate / anhydrate system , we show both the potential of computational chemistry for corroborating and assisting in an experimental program , as well as the need for further developments . a polymorphic system , 3-(4-dibenzo[b , f]oxepin-11-yl - piperazin-1-yl)-2,2-dimethylpropanoic acid , has two zwitterionic hydrate phases with disordered water , which differ remarkably in stability and hydration / dehydration mechanisms . the full multidisciplinary investigation of the behavior of the hydrate and anhydrate phases was complicated by the difficulty in isolating highly crystalline , phase pure samples , the sluggish equilibration of the hydrates at different rh , the metastability of the various forms at different conditions , and the variable hydration state of hya . however , structural based models for both hydrates and the metastable form iii rationalize this behavior . this system exemplifies the dangers in assuming behaviors based on cursory observation of stoichiometric and nonstoichiometric hydration . temperature , water activity , and ph determine the stability ranges of the db7 solid forms , considerably complicating the processing , storage , and handling of this unconventional pharmaceutical hydrate system . Output:	elucidating the crystal structures , transformations , and thermodynamics of the two zwitterionic hydrates ( hy2 and hya ) of 3-(4-dibenzo[b , f][1,4]oxepin-11-yl - piperazin-1-yl)-2,2-dimethylpropanoic acid ( db7 ) rationalizes the complex interplay of temperature , water activity , and ph on the solid form stability and transformation pathways to three neutral anhydrate polymorphs ( forms i , ii , and iii ) . hya contains 1.29 to 1.95 molecules of water per db7 zwitterion ( db7z ) . removal of the essential water stabilizing hya causes it to collapse to an amorphous phase , frequently concomitantly nucleating the stable anhydrate forms i and ii. hy2 is a stoichiometric dihydrate and the only known precursor to form iii , a high energy disordered anhydrate , with the level of disorder depending on the drying conditions . x - ray crystallography , solid state nmr , and h / d exchange experiments on highly crystalline phase pure samples obtained by exquisite control over crystallization , filtration , and drying conditions , along with computational modeling , provided a molecular level understanding of this system . the slow rates of many transformations and sensitivity of equilibria to exact conditions , arising from its varying static and dynamic disorder and water mobility in different phases , meant that characterizing db7 hydration in terms of simplified hydrate classifications was inappropriate for developing this pharmaceutical .
PubmedSumm2098	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: nonpenetrating glaucoma surgeries ( npgs ) , namely , deep sclerectomy and viscocanalostomy with or without implantation of surgical adjuvants , have been developed as an alternative to conventional filtering procedures [ 16 ] . these procedures aim to reduce the intraocular pressure ( iop ) by enhancing the natural aqueous outflow channels ; they reduce the outflow resistance at the inner wall of the schlemm 's canal and the juxtacanalicular trabecular meshwork [ 4 , 7 , 8 ] . the anterior chamber is not opened in the surgical field , thus reducing the likelihood of complications related to conventional penetrating glaucoma surgery ( e.g. , hypotony and choroidal detachment , among others ) [ 3 , 4 , 9 ] . although there is some inconsistency in comparing the efficacy of npgs to trabeculectomy , there seems to be sufficient evidence that npgs provides iop reduction into the high teens [ 1 , 1012 ] . however , due to the nonpenetrating nature of these procedures , the presence of peripheral anterior synechiae ( pas ) at the surgical area remains a challenge because the outflow through the descemet 's membrane is limited in presence of pas . pas can be treated during npgs by a technique previously reported by our group . by transecting the pas during the surgery , spontaneous improvement of aqueous outflow through the trabeculo - descemet 's membrane has been observed . however , to date , no studies have assessed the clinical results of nonpenetrating glaucoma surgery and goniosynechiolysis ab interno in pas - associated glaucoma . this study was initiated and conducted to report the safety and efficacy of deep sclerectomy with goniosynechiolysis ab interno combined with phacoemulsification in the treatment of eyes with cataract and chronic glaucoma associated with pas . the protocol was consistent with the principles of the declaration of helsinki . because of the retrospective nature of the study and because none of the individual - related data were passed to third parties , no approval by the ethical review committee was required . we retrospectively reviewed medical records of patients who had been subject to npgs at our department between january 2000 and december 2006 . twenty eyes of 16 patients ( 8 female and 8 male , mean age 70.0 y , range 5982 ) were identified who had been scheduled for a combined surgery that included npgs , namely , deep sclerectomy with introduction of an adjuvant implant or ocular viscoelastic device ( sk - gel , t - flux or healon gv ) , phacoemulsification and posterior chamber intraocular lens ( iol ) implantation ( one - site surgery ) . choice of implant used was solely based on availability in the operating theatre . in all cases , the indication for the surgery was chronic glaucoma with peripheral anterior synechia ( goniosynechiae ) at the superior iridocorneal angle covering the trabecular meshwork and cataract . the indication for surgery was iop above 20 mmhg under maximally tolerated medical therapy , or above 18 mmhg in cases of severe glaucomatous optic disc cupping ( cdr above 0.8 ) or progression of the disease defined by mean deviation increase of greater than 2 db in the visual field testing . the following data were collected and analyzed : age , gender , pre- and postoperative iop , pre- and postoperative glaucoma medication , type of glaucoma implant , and complications during and after the surgery . after minimal cauterization , a superficial scleral flap was created that reached far into the clear cornea . the deep scleral flap had a triangular shape of approximately 3 3 3 mm . an incision of 2.8 mm width was made into the anterior chamber , and a tunnel was formed under the superficial flap . a deep scleral flap was prepared that was associated with an unroofing of schlemm 's canal and presentation of a descemet 's window , through which one could observe the anterior chamber and peripheral iris . afterwards , the juxtacanalicular trabecular meshwork was peeled . the presence of peripheral anterior synechiae ( goniosynechiae ) in the surgical area usually prevents sufficient aqueous humor outflow , potentially leading to surgical failure . under full visual control through the previously prepared descemet 's window ( figure 1 ) , the anterior chamber was filled with air via paracentesis , followed by introduction of a fine iris spatula into the anterior chamber towards the surgical area . the inserted spatula is utilized to release the peripheral goniosynechia via a gentle swinging movement under direct visualisation through the trabeculo - descemet 's membrane . air movement towards the iridocorneal angle , now open , confirmed successful goniosynechiolysis ab interno . the surgery was completed with the introduction of an implant ( sk - gel , t - flux , or healon gv ) and watertight suturing of the superficial flap and conjunctiva . further surgery was deemed necessary in the event that target iop could be achieved with maximally tolerated medical therapy . complete success was defined as iop 21 mmhg without medication and qualified success with an iop 21 mmhg with medication . all data was collected and stored by the same person ( pr ) , using microsoft excel . the statistical analysis was performed with the program bias for windows ( version 8.6.0 , frankfurt , germany ) . in addition to descriptive statistical analysis , the paired t - test was applied for comparison of preoperative iop versus iop at 12 months after the surgery . the significance level was set at 0.05 . to account for the correlation of the right and left eyes we performed a linear mixed model with unstructured covariance structure using sas version 9.3 . all surgeries were performed by the same experienced glaucoma surgeon ( gs ) . with the described surgical technique , a selective treatment of peripheral anterior synechia was possible during npgs by direct visualisation through a trabeculo - descemet 's window in 19 of 20 eyes ( 95% ) . in one case , the surgery was converted to phacotrabeculectomy with iridectomy due to intraoperative rupture of descemet 's window . sk - gel ( corneal , paris , france ) was implanted in 10 eyes ( 50% ) , t - flux ( ioltech , la rochelle , france ) in 6 eyes ( 30% ) , and healon gv ( amo , ettlingen , germany ) in 3 eyes ( 15% ) . preoperative gonioscopy had revealed interrupted areas of peripheral anterior synechiae ( goniosynechiae ) in all cases , and the angle was open ( shaffer ii , iii , or iv ) in more than 180. none of the eyes developed a pupillary block postoperatively . during the follow - up period no yag laser goniopunctures were performed . no further glaucoma surgery was necessary in any of the cases during the follow - up period . the mean preoperative iop was 20.3 5.2 mmhg ( median 20 , range 1230 ) on 2.4 1.0 medications ( median 2 , range 05 ) . the course of iop was as follows ( figures 2 and 3 ) : at day 1 , mean iop was 11.7 6.1 mmhg ( median 11 , range 221 ) ; at week 1 , mean iop was 13.2 6.7 mmhg ( median 13.5 , range 224 ) ; at month 1 , mean iop was 12.7 3.5 mmhg ( median 12 , range 621 ) ; and at month 3 , mean iop was 14.2 4.2 mmhg ( median 14 , range 621 ) . one year postoperatively , the mean iop was 15.3 3.3 mmhg ( median 16 , range 1020 ) with 0.6 1.0 medication , which was significantly different from preoperative iop ( p = 0.004 ) . within the linear mixed model only time had an influence with the mean iop being lowered by 5.96 mmhg , 95% confidence interval [ 2.83 ; 9.09 ] , p = 0.001 . this linear mixed model also showed that the results were not influenced by the inclusion of both eyes in 4 patients . a postoperative iop of 21 mmhg ( complete success ) was achieved in 17/19 eyes ( 89.5% ) at 3 months and in 12/19 eyes ( 63.2% ) at 12 months postoperatively . in the remaining eyes ( 10.5% at 3 months and 36.8% at 12 months ) , additional medication , at mean quantities of 0.3 and 0.6 , respectively , led to an iop 21 mmhg ( qualified success ) . using a more stringent criterion of iop < 18 mmhg , success was achieved in 15/19 eyes ( 78.9% ) at three months and in 14/19 eyes ( 73.7% ) at one year after surgery . one year after the surgery , the mean iop reduction was 5.0 6.4 mmhg ( median 4 , range 18 to + 8) . the mean number of medications decreased from 2.4 ( median 2 ) before the surgery to 0.63 ( median 0 ) one year postoperatively . peripheral anterior synechia ( goniosynechia ) , if present directly at the area of flap created in npgs , may compromise the surgical outcome . peripheral anterior synechia can lead to a potential failure of npgs if they occur at the sites where npgs would typically reduce the outflow resistance , namely , at the trabecular meshwork adjacent to the sclerocorneal flap . in a prospective study , moreno - montas et al . releasing these synechiae using the technique previously described by our group not only may be primarily beneficial by clearing the outflow tract but also may be advantageous if nd : yag laser goniopuncture is needed postoperatively ( in the case of insufficient iop regulation after primary surgery ) . absence of pas enables the laser surgeon to aim directly for perforation of descemet 's membrane with a nd : yag laser without any obstruction , thus leading to a lower risk of inadvertent laser damage to the peripheral iris , which in itself would induce further goniosynechiae after the nd : yag goniopuncture . in our experience , releasing the pas by this maneuver led to an immediate improvement of aqueous humor outflow through the trabecular meshwork . although we can not quantify this improvement , we believe that treatment of pas , when present at the surgical field , enhances the trans - descemetic outflow in npgs . in our series the surgery was completed with the introduction of an implant ( sk - gel , t - flux , or healon gv ) . the sk - gel and the healon gv are implants that dissolve over time in about 4 to 6 months after surgery , as they are made of hyaluronic acids . on the other hand the t - flux is a nonabsorbable implant made of poly megma ( hydrophilic acrylic ) . the overall results and success rates might differ upon absorption or nonabsorption of the implant . our study sample is too small to answer to this question and this is certainly a limitation . furthermore , the follow - up time is limited ; thus we are not able to provide long - term results of our study sample . the presence of goniosynechia might be related to a previous intraocular inflammation . in our study sample the patients did not report previous inflammatory disease and no active inflammation was detectable at the time of presentation . a preoperative gonioscopy had revealed presence of very few goniosynechia ; however the cases have been classified as open angle glaucoma , as inflammation was neither reported nor present , and almost all parts of the angle were clearly open . the functional relevance of the goniosynechia at the surgical field was detected during the surgery . as a matter of fact report a mean reduction of iop of around 5.5 mmhg with reduction of medications from 2.0 preoperatively to 0.3 after deep sclerectomy and phacoemulsification . the type of implant used ( sk - gel versus t - flux ) does not seem to have an impact on the success of the surgery . in a meta - analysis , cheng et al . conclude that the efficacy of nonpenetrating glaucoma surgery as measured by iop reduction is equivalent in cases with and without introduction of an implant during the surgery . our results are similar to those reported by schreyger et al . ; we noted a mean iop reduction of 5.0 6.4 mmhg and a reduction in the number of medications from 2.4 before surgery to 0.6 after surgery . it is also noteworthy that the operations in both studies were performed by the same surgeon ( gs ) . we , thus , conclude that the iop reduction seen in eyes with chronic glaucoma and pas treated with npgs with goniosynechiolysis ab interno in combination with phacoemulsification is similar to the iop reduction seen in eyes with poag without pas treated with npgs and phacoemulsification . the exact position of the peripheral anterior synechiae can be determined prior to npgs by gonioscopy , and the surgeon may choose to create a flap at another position ( e.g. , lateral ) where such adhesion may not be present . in our experience , however , this situation is rare as goniosynechiae are often present at several positions around the iridocorneal angle . as a result , changing the access location is not always an option . moreover , most surgeons perform this very delicate surgery from a preferred access area ( usually superior ) and changing this preferred access site may lead to a worse outcome . some surgeons make microincisions in descemet 's membrane ( descemet 's fenestration ) if the outflow test reveals insufficient aqueous flow during the surgery . however , in our experience , synechiolysis ab interno as performed in our series of patients unequivocally enhances outflow , which can be directly visualized during the surgery . descemet 's fenestration may be considered if goniosynechiolysis does not lead to the desired outflow rate . in the case that the spatula inadvertently damages and perforates the trabeculo - descemet 's membrane during goniosynechiolysis , the iris should be observed carefully . if there is an iris prolapse , a peripheral iridectomy should be performed ; if there are only microperforations without iris prolapse , injection of air into the anterior chamber can be considered because air maintains the anterior chamber and prevents iris incarceration into the defect of descemetic membrane . in our experience surgery can and should be converted to trabeculectomy in case of descemet 's membrane rupture and iris prolapse . the efficacy and success rates of npgs , in terms of iop reduction , reports a higher success rate ( iop 20 mmhg without medication ) when combining deep sclerectomy with phacoemulsification versus deep sclerectomy alone ( 90% versus 62% ) , whereas marek et al . thus , we conjecture that the combination of phacoemulsification and deep sclerectomy surgery most likely does not compromise the success rate with regard to iop reduction . in fact , it is possible that cataract surgery exerts a positive influence on the postoperative iop . however , it seems unlikely that the cataract surgery alone would have led to the iop reduction seen with the combined surgery . we do not have data on a control group ; however we believe that reduction of the preoperative iop from 20.3 5.2 mmhg with 2.4 1.0 medications to 15.3 3.3 mmhg with 0.6 1.0 medications after the surgery appears to be impossibly attributed to the phacoemulsification alone . phacoemulsification and intraocular lens implantation were performed at the same surgical area as the deep sclerectomy ( superior , one - site surgery ) . in a literature review ( january 2012 ) , we were unable to find any comparative studies on one - site versus two - site combined npgs and phacoemulsification . therefore , it remains unclear whether the cataract surgery incision site had any influence on safety or efficacy of the procedure in our study . similarly , there were no medline - listed publications on the efficacy of trabeculectomy in presence of peripheral anterior synechia . although we do not expect the presence of pas to compromise the efficacy results of trabeculectomy , no published study data are available supporting this expectation . our positive experience with goniosynechiolysis ab interno , as presented in this paper , has encouraged us to perform nonpenetrating glaucoma surgery even in the presence of peripheral anterior synechiae . after development and publication of the goniosynechiolysis method our study delivers first data on the efficacy and safety of this procedure . a comparative study of nonpenetrating glaucoma surgery with goniosynechiolysis ab interno versus conventional trabeculectomy in presence of peripheral anterior synechia would shed light on the question of comparative efficacy and safety of these methods . with goniosynechiolysis ab interno , the surgeon is able to perform nonpenetrating glaucoma surgery even in presence of peripheral anterior synechia . deep sclerectomy with goniosynechiolysis ab interno in combination with phacoemulsification was safe and effective in treatment of eyes with cataract and chronic glaucoma associated with peripheral anterior synechia . Output:	purpose . to report one - year results of phacoemulsification combined with deep sclerectomy and goniosynechiolysis ab interno for chronic glaucoma associated with peripheral anterior synechiae ( pas ) . methods . we retrospectively analyzed medical charts of 16 patients ( 20 eyes ) treated by one - site combined phacoemulsification and deep sclerectomy with goniosynechiolysis ab interno . pas were transected by a spatula introduced into the anterior chamber through a paracentesis . to account for the correlation of right and left eyes a linear mixed model with unstructured covariance structure was calculated . results . the mean preoperative intraocular pressure ( iop ) was 20.3 5.2 mmhg with 2.4 1.0 medications . one year postoperatively , the mean iop was 15.3 3.3 mmhg ( p = 0.004 , paired t - test ) with 0.6 1.0 medications . a postoperative iop of 21 mmhg without medication was achieved in 17 of 19 eyes ( 89.5% ) and in 12/19 eyes ( 63.2% ) at 3 and 12 months after surgery , respectively . in the remaining eyes ( 10.5% at 3 months and 36.8% at 12 months ) , additional medication led to an iop 21 mmhg or the target pressure . no case required further glaucoma surgery . in one eye , conversion of the surgery to trabeculectomy was necessary due to descemet 's window rupture . conclusions . with goniosynechiolysis ab interno , effective and safe nonpenetrating glaucoma surgery is possible in presence of pas .
PubmedSumm2099	*TASK* the task is to summarize an input biomedical literature in six sentences *INPUT* the input is a biomedical literature *OUTPUT* the output is the summary of an input biomedical literature in six sentences *DOCUMENTATION* *EXAMPLES* Input: several groups of animals enter into symbiotic relationships with algae , the zoochlorellae of hydra being a well - known example ( habetha et al . 2003 ; kawaida et al . sacoglossan slugs are unique , however , in that they perform photosynthesis and fix carbon in a light - dependent manner using plastids that they sequester from the algae upon which they feed ( greene 1970 ; marn and ros 1989 ; hndeler et al . 2009 ) . five species among the sacoglossan slugs elysia chlorotica , e. timida , e. crispata , e. clarki , and plakobranchus ocellatus perform what is called long - term retention ( ltr ) of sequestered plastids ( kleptoplasts ) . that is , when adult animals are given the opportunity to graze upon their preferred algal food source , they can survive subsequent starvation for up to several months , during which time they maintain active plastids with functional photosystems ( mujer et al . 1 ) . though often described as solar - powered slugs , it is not yet clear how , exactly , the slugs benefit from the kleptoplasts , as recent findings show that plastid - bearing e. timida and p. ocellatus survive starvation for months in the dark just as well as they do in the light ( christa et al . ltr species are distinguished from short - term retention ( str ) species , the ingested plastids of which lose their photosynthetic ability rapidly over the first 2 weeks of starvation and are more rapidly digested than in ltr species ( hndeler et al . both ltr and str sacoglossans feed by tapping the plastid - rich cytosol of siphonaceous algae , which have large cells , centimeters or more in length . the sequestered kleptoplasts continue to perform photosynthesis for up to several months within specialized cells of the digestive gland of the slug . the sequestered kleptoplasts continue to perform photosynthesis for up to several months within specialized cells of the digestive gland of the slug . because photosystems are known to have a relatively high rate of protein turnover in higher plants and algae studied so far ( aro et al . 2000 ; komenda et al . 2012 ) , it was long speculated that the nuclear genomes of ltr species acquired genes of algal origin via lateral gene transfer ( lgt ) : genes that encode products such as light harvesting complex proteins or psbo ( pierce et al . 2007 ; rumpho et al . 2008 ) might help to maintain plastids in an active state by servicing the photosystems . a problem with the lgt hypothesis was that ability to perform ltr arose in multiple sacoglossan lineages independently , complicating the number and nature of putative transfers ( wgele et al . moreover , direct tests of the lgt hypothesis using deep sequencing on e. timida and p. ocellatus ( wgele et al . 2011 ) and later on e. chlorotica ( rumpho et al . 2011 ) showed that plastid - bearing ltr sacoglossans do not express any genes of algal origin . genome sequence data for e. chlorotica eggs furthermore showed that the slugs do not harbor algal dna ( bhattacharya et al . accordingly , lgt can not be the mechanism underlying kleptoplast survival . in search of an explanation for kleptoplast longevity in ltr sacoglossans , we revisit square one . there is a distinct trend among ltr slugs to specialize and often feed on a single algal species . the preferred algal species , however , are very different for different slug species and come from very distant corners of plastid diversity . e. chlorotica ingests plastids from vaucheria litorea , a xanthophyte alga housing a plastid of secondary endosymbiotic ( red algal ) origin ( rumpho et al . 2001 ; archibald 2009 ; gould 2012 ) , while e. timida ingests plastids from the ulvophyte green alga acetabularia acetabulum ( fig . 1 ; marn and ros 1989 ) . these two slugs together with p. ocellatus the slugs with greatest kleptoplast longevity feed and survive from just the one species of alga upon which they have specialized . the closely related e. crispata and e. clarki sequester plastids from ulvophytes , namely halimeda , bryopsis , batophora , caulerpa , penicillus , and codium ( clark and busacca 1978 ; curtis et al . plakobranchus ocellatus steals plastids from various algae , too , but during starvation , then strikingly retains only those of halimeda ( christa et al . we have observed the same to occur in starvation experiments on e. clarki , during which plastids of halimeda were detectable two weeks after the onset of starvation , while plastids of bryposis were not , suggesting that the latter had been digested while the former had been retained . this was determined using a barcoding approach that , for p. ocellatus , recently provided similar results ( christa et al . yet , if not all ingested plastids in ltr slugs are retained , could kleptoplast longevity in slugs be partly attributable to properties of the plastids themselves ? the plastids sequestered by the different ltr species belong to algae from quite distantly related lineages , but could they have something in common that has been so far overlooked ? the plastid genome of v. litorea , whose plastids remain photosynthetically active for up to 10 months in the slugs ( green et al . 2000 ) , was already fully sequenced in the course of studies on e. chlorotica ( rumpho et al . no chloroplast genome sequence , however , was available for a. acetabulum , the sole food source of e. timida . the a. acetabulum plastid genome has been found to contain large repetitive elements ( tymms and schweiger 1985 ) and estimated to reach a size of around 2,000 kb ( manhart et al . 1989 ) . through centrifugation , we generated a dna fraction that was enriched for a. acetabulum plastid dna and by shotgun sequencing of this chloroplast - enriched fraction , we obtained 138,285 kb of vector - trimmed raw data , from which we identified and assembled 63 contigs encoding proteins homologous to known plastid proteins of the utc clade ( ulvophyceae , trebouxiophyceae , and chlorophyceae ) of green algae . s1 , supplementary material online ) and an at content of 69% , which is comparable to that of the ulvophyceans pseudendoclonium akinetum ( 68.5% ) and bryopsis hypnoides ( 66.9% ; pombert et al . 2005 ; l et al . all contigs together represent a total length of approximately 350 kb , but we estimate the complete genome to be substantially larger , possibly as big as the 2,000 kb estimate of manhart et al . intron and intergenic region lengths , for example often many kilobases long by far exceed those identified from plastid genomes of related ulvophycean algae , whose genomes are less than 200 kb long ( pombert et al . next to the rpoc2 locus for example sits a 7,785 bp long open reading frame , potentially encoding a protein of 303 kda with no significant similarity ( e - value cutoff 10 ) to any known proteins ( fig . 2 ) . among the 51 protein coding genes with homology to common plastid genes , and for which we have full - length sequences ( table 1 ) , were ftsh and tufa , two proteins that we suggest in the following to be of particular interest with regard to understanding plastid longevity . these two genes are also encoded by the plastid genome of v. litorea ( fig . shown are contigs in comparison to the corresponding parts of the fully sequenced plastid genome of the phylogenetically related pseudendoclonium akinetum encoding identical genes . the comparison illustrates the expansion of most intergenic and intron regions , and the increase in introns and open reading frames ( orfs 300 bp shown in gray ) in acetabularia . although the rna polymerase rpoc2 in p. akinetum is encoded by a single reading frame , in a. acetabulum it is highly fragmented across many dozen kilobase pairs . many contigs assembled encode only a single gene ( e.g. , accd or rps7 ) . numbers beneath gene names represent the amino acid positions in the homolog of p. akinetum . tatc , for example , is only retained in plastid genomes of the red lineage , while psba and atpa , for instance , are encoded by the plastid genomes of all 51 organisms analyzed . the majority of algae and water , but not land - dwelling streptophytes ( embryophyta ) , encode ftsh and tufa on their plastid genomes . the cladogram is based on a multigene phylogeny of 17 genes ( table 1 ) that are shared by all plastid genomes shown . top left corner shows details on the three algae ( in bold ) , whose plastids are being sequestered by slugs . table 1list of the 51 full - length plastid encoded genes of acetabularia acetabulum identifiedgeneorf length ( bp)contig length ( bp)at contentpresence / absenceaccessionp . h.accd88513,80769.4hg18425atpa1,5098,86865.9hg18426atpb1,4402,57165.9hg18427atpe_13995,69466.2hg18428atpe_23995,97257.1naatpf5184,70573.3hg18429atph24959.8hg18430chlb1,5752,40767.3hg18431chli1,1073,27266.3hg18432chll86416,52670.1hg18433chln1,46168.8hg18434psbm10576.2hg18450clpp5914,06566.3hg18435cysa6932,50472.0hg18436cyst8046,56473.5hg18437ftsh13,48819,26362.3hg18438infa20711,47572.0hg18439rpl554073.5hg18462rpl1436969.1hg18456rps841474.2hg18469peta9243,12071.5hg18440petb6484,80867.1hg18441petd46265.6hg18442petg1023,22666.7hg18443psab2,0823,30559.1hg18444psac2467,36162.6hg18445psaj12629,13375.4hg18446psbj12961.2hg18448ycf435474.6hg18474psba1,0359,70059.5hg794360psbb1,4195,60762.4hg18447psbk1327,44570.5hg18449ycf1210275.5hg18472psbn1356,26472.6hg18451psbt965,85171.9hg18452psbz1894,96272.5hg18453rbcl_11,4583,33661.5hg18454rbcl_21,0892,65861.5narpl125287,37168.4hg18455rps947769.6hg18470rpl164294,23664.6hg18457rpl193394,65476.7hg18458rpl276818,09266.0hg18459rpl2328876.7hg18461rps1927969.2hg18466rpl203515,35678.4hg18460rps113819,84563.5hg18463rps143038,62672.6hg18464rps182403,23674.2hg18465rps46095,61973.7hg18467rps747116,82767.5hg18468tufa1,2304,58365.9hg18471ycf35162,75067.4hg18473note.the second column shows the gene length , the third column the contig length . for contigs encoding more than one gene , final columns indicate presence / absence of the genes from the plastid genomes of the related pseudendoclonium akinetum , oltmannsiellopsis viridis , and bryopsis hypnoides . sequencing contigs of the a. acetabulum plastid genome . shown are contigs in comparison to the corresponding parts of the fully sequenced plastid genome of the phylogenetically related pseudendoclonium akinetum encoding identical genes . the comparison illustrates the expansion of most intergenic and intron regions , and the increase in introns and open reading frames ( orfs 300 bp shown in gray ) in acetabularia . although the rna polymerase rpoc2 in p. akinetum is encoded by a single reading frame , in a. acetabulum it is highly fragmented across many dozen kilobase pairs . many contigs assembled encode only a single gene ( e.g. , accd or rps7 ) . numbers beneath gene names represent the amino acid positions in the homolog of p. akinetum . tatc , for example , is only retained in plastid genomes of the red lineage , while psba and atpa , for instance , are encoded by the plastid genomes of all 51 organisms analyzed . the majority of algae and water , but not land - dwelling streptophytes ( embryophyta ) , encode ftsh and tufa on their plastid genomes . the cladogram is based on a multigene phylogeny of 17 genes ( table 1 ) that are shared by all plastid genomes shown . top left corner shows details on the three algae ( in bold ) , whose plastids are being sequestered by slugs . list of the 51 full - length plastid encoded genes of acetabularia acetabulum identified note.the second column shows the gene length , the third column the contig length . for contigs encoding more than one gene , final columns indicate presence / absence of the genes from the plastid genomes of the related pseudendoclonium akinetum , oltmannsiellopsis viridis , and bryopsis hypnoides . when we compared plastid genome data of algae and plants , it became apparent that in particular land plant plastids lack several genes commonly encoded by plastid genomes of a large variety of different algae , from the red , as well as the green lineage ( rhodophytes and chlorophytes , respectively ; fig . 3 ) . among those genes was the protease encoding ftsh a protein essential for photosystem ii maintenance and tufa encoding the translation elongation factor tu ( watson and surzycki 1982 ) . early studies showed that sequestered plastids of v. litorea actively continue to transcribe and translate plastid - encoded psba ( mujer et al . 1996 ) , encoding the d1 protein of photosystem ii , and transcripts of the v. litorea plastid - encoded ftsh and tufa were further found among rna of e. chlorotica that had been starved for 2 months ( pierce et al . we found evidence for the presence of all three transcripts in e. timida slugs that had been starved for 1 month ( fig . moreover , translation of ftsh transcript would be impaired in the absence of the crucial elongation factor tu encoded by tufa . a replenishable supply of these gene products might be key to long - term plastid activity in slugs . we isolated total rna from slugs that had been starved for 1 month ( 31 days ) and performed nonquantitative reverse - transcriptase pcrs to screen for the presence of mrna of the three genes tufa , psba , and ftsh in starving e. timida slugs ( e. t. ) . rna of a. acetabulum ( a. a. ) served as an additional positive control . + rt/rt indicates the presence or absence of the reverse - transcriptase enzyme in the reaction . we isolated total rna from slugs that had been starved for 1 month ( 31 days ) and performed nonquantitative reverse - transcriptase pcrs to screen for the presence of mrna of the three genes tufa , psba , and ftsh in starving e. timida slugs ( e. t. ) . + rt/rt indicates the presence or absence of the reverse - transcriptase enzyme in the reaction . photosystem turnover in sequestered plastids of a. acetabulum and v. litorea , two preferred plastid sources for ltr slugs , has not been directly studied so far . constant photodamage in land plant plastids demands a high level of protein import from the cytosol to replace affected components of the photosystems ( aro et al . photosystem ii ( psii ) , in particular its d1 protein , is affected the most by photodamage , and the latter is also a major culprit regarding subsequent damage , as a degenerate d1 leads to an accumulation of reactive oxygen species ( ros ) ( nishiyama et al . downstream , ros not only affect a large variety of other biochemical pathways ( girotti 2001 ; apel and hirt 2004 ) but further inhibit the repair of the photosystem itself ( nishiyama et al . essential for the repair of a damaged psii is the removal of the faulty d1 protein , a process that is mainly mediated by the ftsh protease complex in plants and cyanobacteria ( nixon et al . ftsh proteins act as quality control proteases , as either hetero- or homo - oligomers ( janska et al . twelve ftsh genes are encoded in the arabidopsis thaliana nuclear genome , nine of which are targeted to the plastid ( sakamoto et al . although all chloroplast - targeted ftsh proteases in the land plant can apparently assemble into functional hetero - oligomers always consisting of a - type and b - type subunits in mitochondria , the functional ftsh complex can consist of only a homo - oligomer ( janska et al . furthermore , only one subunit needs to contain the functional proteolytic m41 domain ( zhang et al . 2010 ) . in variegated arabidopsis mutants , the loss of only a single ftsh gene results in high levels of accumulated ros in the plastids , causing severe damage to the entire plant ( kato et al . 3 ) , the protease needs to be imported from the cytosol . in plastids sequestered by the slugs feeding only on one algal species , it is encoded by the plastid itself . this could explain why ltr species that feed on more than one algal species do not retain the ulvophyte bryopsis the correlation between plastid - encoded ftsh and ltr deserves further study in algal grazing experiments . that the plastids from some algal lineages , including ulvophytes , are more robust than land plant plastids was noted 40 years ago ( giles and sarafis 1972 ) . it was suggested that such robustness , sometimes bordering on apparent plastid autonomy , might be linked to the prolonged survival of plastids in what are now called ltr slugs ( trench et al . although the reason(s ) underlying the robustness of plastids that sacoglossans sequester remained obscure , the role of light stress and photodamage to psii , in particular , has always figured prominently in the issue of sacoglossan plastid longevity ( reviewed in rumpho et al . and cruz et al . ) . ( 2010 ) recently showed , for acetabularia plastids sequestered within in e. timida , that psii recovers remarkably well subsequent to bleaching ; they furthermore noted that e. timida kleptoplasts retain a. acetabulum photo - damage repair mechanisms . here we are suggesting that kleptoplast robustness and photodamage repair at psii are causally related , and that this conceivably could be attributable to only one or a few factors , with ftsh playing a pivotal role . our reasoning here is guided by the observation that protein synthesis in land plant plastids has two temporally distinct roles : 1 ) biogenesis of the photosynthetic apparatus followed by 2 ) the maintenance phase during which the repair of photodamaged psii in particular plays the most prominent role ( nixon et al . sacoglossans have no need for thylakoid biogenesis , as they acquire mature plastids , leaving the maintenance role as a possible factor . if the kleptoplasts are robust , which they are , then either the slugs actively render them robust , or their robustness is an intrinsic property , or a combination of the two . the observation that most sacoglossans simply digest all ingested plastids , regardless of their source ( hndeler et al . 2013 ) , indicates that ltr species specifically provide an environment where plastids can persist . the observation that some ltr species such as plakobranchus ingest plastids from several sources but retain only those from halimeda long - term as kleptoplasts ( christa et al . 2013 ) suggests that the ability to persist is a property intrinsic to the plastids . plakobranchus could also selectively digest some plastids faster than others , implying the existence of digestive recognition mechanisms for individual food particles . but selective digestion , even if it exists , would still not explain why some kleptoplasts can survive for so long within slugs , again pointing to plastid intrinsic properties . because ftsh connects psii repair to a plastid intrinsic property being plastid - encoded in plastids that undergo ltr , it emerges as a prime candidate for a causal factor behind plastid longevity . a clear prediction of this hypothesis is that acetabularia plastids should be particularly robust to high light intensities and recover faster from light stress , both in algae but especially in slugs , in comparison to plastids that lack ftsh genes , such as those of bryopsis . a further implication of these findings is that they might open new avenues of pursuit for the engineering of higher plant plastids with increased tolerance to light stress . there is a distinct trend among ltr slugs to specialize and often feed on a single algal species . the preferred algal species , however , are very different for different slug species and come from very distant corners of plastid diversity . e. chlorotica ingests plastids from vaucheria litorea , a xanthophyte alga housing a plastid of secondary endosymbiotic ( red algal ) origin ( rumpho et al . 2001 ; archibald 2009 ; gould 2012 ) , while e. timida ingests plastids from the ulvophyte green alga acetabularia acetabulum ( fig . 1 ; marn and ros 1989 ) . these two slugs together with p. ocellatus the slugs with greatest kleptoplast longevity feed and survive from just the one species of alga upon which they have specialized . the closely related e. crispata and e. clarki sequester plastids from ulvophytes , namely halimeda , bryopsis , batophora , caulerpa , penicillus , and codium ( clark and busacca 1978 ; curtis et al . plakobranchus ocellatus steals plastids from various algae , too , but during starvation , then strikingly retains only those of halimeda ( christa et al . we have observed the same to occur in starvation experiments on e. clarki , during which plastids of halimeda were detectable two weeks after the onset of starvation , while plastids of bryposis were not , suggesting that the latter had been digested while the former had been retained . this was determined using a barcoding approach that , for p. ocellatus , recently provided similar results ( christa et al . yet , if not all ingested plastids in ltr slugs are retained , could kleptoplast longevity in slugs be partly attributable to properties of the plastids themselves ? the plastids sequestered by the different ltr species belong to algae from quite distantly related lineages , but could they have something in common that has been so far overlooked ? the plastid genome of v. litorea , whose plastids remain photosynthetically active for up to 10 months in the slugs ( green et al . 2000 ) , was already fully sequenced in the course of studies on e. chlorotica ( rumpho et al . no chloroplast genome sequence , however , was available for a. acetabulum , the sole food source of e. timida . the a. acetabulum plastid genome has been found to contain large repetitive elements ( tymms and schweiger 1985 ) and estimated to reach a size of around 2,000 kb ( manhart et al . 1989 ) . through centrifugation , we generated a dna fraction that was enriched for a. acetabulum plastid dna and by shotgun sequencing of this chloroplast - enriched fraction , we obtained 138,285 kb of vector - trimmed raw data , from which we identified and assembled 63 contigs encoding proteins homologous to known plastid proteins of the utc clade ( ulvophyceae , trebouxiophyceae , and chlorophyceae ) of green algae . s1 , supplementary material online ) and an at content of 69% , which is comparable to that of the ulvophyceans pseudendoclonium akinetum ( 68.5% ) and bryopsis hypnoides ( 66.9% ; pombert et al . 2005 ; l et al . all contigs together represent a total length of approximately 350 kb , but we estimate the complete genome to be substantially larger , possibly as big as the 2,000 kb estimate of manhart et al . intron and intergenic region lengths , for example often many kilobases long by far exceed those identified from plastid genomes of related ulvophycean algae , whose genomes are less than 200 kb long ( pombert et al . next to the rpoc2 locus for example sits a 7,785 bp long open reading frame , potentially encoding a protein of 303 kda with no significant similarity ( e - value cutoff 10 ) to any known proteins ( fig . 2 ) . among the 51 protein coding genes with homology to common plastid genes , and for which we have full - length sequences ( table 1 ) , were ftsh and tufa , two proteins that we suggest in the following to be of particular interest with regard to understanding plastid longevity . these two genes are also encoded by the plastid genome of v. litorea ( fig . shown are contigs in comparison to the corresponding parts of the fully sequenced plastid genome of the phylogenetically related pseudendoclonium akinetum encoding identical genes . the comparison illustrates the expansion of most intergenic and intron regions , and the increase in introns and open reading frames ( orfs 300 bp shown in gray ) in acetabularia . although the rna polymerase rpoc2 in p. akinetum is encoded by a single reading frame , in a. acetabulum it is highly fragmented across many dozen kilobase pairs . many contigs assembled encode only a single gene ( e.g. , accd or rps7 ) . numbers beneath gene names represent the amino acid positions in the homolog of p. akinetum . tatc , for example , is only retained in plastid genomes of the red lineage , while psba and atpa , for instance , are encoded by the plastid genomes of all 51 organisms analyzed . the majority of algae and water , but not land - dwelling streptophytes ( embryophyta ) , encode ftsh and tufa on their plastid genomes . the cladogram is based on a multigene phylogeny of 17 genes ( table 1 ) that are shared by all plastid genomes shown . top left corner shows details on the three algae ( in bold ) , whose plastids are being sequestered by slugs . table 1list of the 51 full - length plastid encoded genes of acetabularia acetabulum identifiedgeneorf length ( bp)contig length ( bp)at contentpresence / absenceaccessionp . h.accd88513,80769.4hg18425atpa1,5098,86865.9hg18426atpb1,4402,57165.9hg18427atpe_13995,69466.2hg18428atpe_23995,97257.1naatpf5184,70573.3hg18429atph24959.8hg18430chlb1,5752,40767.3hg18431chli1,1073,27266.3hg18432chll86416,52670.1hg18433chln1,46168.8hg18434psbm10576.2hg18450clpp5914,06566.3hg18435cysa6932,50472.0hg18436cyst8046,56473.5hg18437ftsh13,48819,26362.3hg18438infa20711,47572.0hg18439rpl554073.5hg18462rpl1436969.1hg18456rps841474.2hg18469peta9243,12071.5hg18440petb6484,80867.1hg18441petd46265.6hg18442petg1023,22666.7hg18443psab2,0823,30559.1hg18444psac2467,36162.6hg18445psaj12629,13375.4hg18446psbj12961.2hg18448ycf435474.6hg18474psba1,0359,70059.5hg794360psbb1,4195,60762.4hg18447psbk1327,44570.5hg18449ycf1210275.5hg18472psbn1356,26472.6hg18451psbt965,85171.9hg18452psbz1894,96272.5hg18453rbcl_11,4583,33661.5hg18454rbcl_21,0892,65861.5narpl125287,37168.4hg18455rps947769.6hg18470rpl164294,23664.6hg18457rpl193394,65476.7hg18458rpl276818,09266.0hg18459rpl2328876.7hg18461rps1927969.2hg18466rpl203515,35678.4hg18460rps113819,84563.5hg18463rps143038,62672.6hg18464rps182403,23674.2hg18465rps46095,61973.7hg18467rps747116,82767.5hg18468tufa1,2304,58365.9hg18471ycf35162,75067.4hg18473note.the second column shows the gene length , the third column the contig length . for contigs encoding more than one gene , final columns indicate presence / absence of the genes from the plastid genomes of the related pseudendoclonium akinetum , oltmannsiellopsis viridis , and bryopsis hypnoides . are contigs in comparison to the corresponding parts of the fully sequenced plastid genome of the phylogenetically related pseudendoclonium akinetum encoding identical genes . the comparison illustrates the expansion of most intergenic and intron regions , and the increase in introns and open reading frames ( orfs 300 bp shown in gray ) in acetabularia . although the rna polymerase rpoc2 in p. akinetum is encoded by a single reading frame , in a. acetabulum it is highly fragmented across many dozen kilobase pairs . many contigs assembled encode only a single gene ( e.g. , accd or rps7 ) . numbers beneath gene names represent the amino acid positions in the homolog of p. akinetum . tatc , for example , is only retained in plastid genomes of the red lineage , while psba and atpa , for instance , are encoded by the plastid genomes of all 51 organisms analyzed . the majority of algae and water , but not land - dwelling streptophytes ( embryophyta ) , encode ftsh and tufa on their plastid genomes . the cladogram is based on a multigene phylogeny of 17 genes ( table 1 ) that are shared by all plastid genomes shown . top left corner shows details on the three algae ( in bold ) , whose plastids are being sequestered by slugs . list of the 51 full - length plastid encoded genes of acetabularia acetabulum identified note.the second column shows the gene length , the third column the contig length . for contigs encoding more than one gene , final columns indicate presence / absence of the genes from the plastid genomes of the related pseudendoclonium akinetum , oltmannsiellopsis viridis , and bryopsis hypnoides . when we compared plastid genome data of algae and plants , it became apparent that in particular land plant plastids lack several genes commonly encoded by plastid genomes of a large variety of different algae , from the red , as well as the green lineage ( rhodophytes and chlorophytes , respectively ; fig . 3 ) . among those genes was the protease encoding ftsh a protein essential for photosystem ii maintenance and tufa encoding the translation elongation factor tu ( watson and surzycki 1982 ) . early studies showed that sequestered plastids of v. litorea actively continue to transcribe and translate plastid - encoded psba ( mujer et al . 1996 ) , encoding the d1 protein of photosystem ii , and transcripts of the v. litorea plastid - encoded ftsh and tufa were further found among rna of e. chlorotica that had been starved for 2 months ( pierce et al . we found evidence for the presence of all three transcripts in e. timida slugs that had been starved for 1 month ( fig . 4 ) . moreover , translation of ftsh transcript would be impaired in the absence of the crucial elongation factor tu encoded by tufa . a replenishable supply of these gene products might be key to long - term plastid activity in slugs . we isolated total rna from slugs that had been starved for 1 month ( 31 days ) and performed nonquantitative reverse - transcriptase pcrs to screen for the presence of mrna of the three genes tufa , psba , and ftsh in starving e. timida slugs ( e. t. ) . rna of a. acetabulum ( a. a. ) served as an additional positive control . + rt/rt indicates the presence or absence of the reverse - transcriptase enzyme in the reaction . we isolated total rna from slugs that had been starved for 1 month ( 31 days ) and performed nonquantitative reverse - transcriptase pcrs to screen for the presence of mrna of the three genes tufa , psba , and ftsh in starving e. timida slugs ( e. t. ) . rna of a. acetabulum ( a. a. ) served as an additional positive control . + rt/rt indicates the presence or absence of the reverse - transcriptase enzyme in the reaction . photosystem turnover in sequestered plastids of a. acetabulum and v. litorea , two preferred plastid sources for ltr slugs , has not been directly studied so far . constant photodamage in land plant plastids demands a high level of protein import from the cytosol to replace affected components of the photosystems ( aro et al . photosystem ii ( psii ) , in particular its d1 protein , is affected the most by photodamage , and the latter is also a major culprit regarding subsequent damage , as a degenerate d1 leads to an accumulation of reactive oxygen species ( ros ) ( nishiyama et al . downstream , ros not only affect a large variety of other biochemical pathways ( girotti 2001 ; apel and hirt 2004 ) but further inhibit the repair of the photosystem itself ( nishiyama et al . 2006 ) . essential for the repair of a damaged psii is the removal of the faulty d1 protein , a process that is mainly mediated by the ftsh protease complex in plants and cyanobacteria ( nixon et al . ftsh proteins act as quality control proteases , as either hetero- or homo - oligomers ( janska et al . twelve ftsh genes are encoded in the arabidopsis thaliana nuclear genome , nine of which are targeted to the plastid ( sakamoto et al . although all chloroplast - targeted ftsh proteases in the land plant can apparently assemble into functional hetero - oligomers always consisting of a - type and b - type subunits in mitochondria , the functional ftsh complex can consist of only a homo - oligomer ( janska et al . furthermore , only one subunit needs to contain the functional proteolytic m41 domain ( zhang et al . 2010 ) . in variegated arabidopsis mutants , the loss of only a single ftsh gene results in high levels of accumulated ros in the plastids , causing severe damage to the entire plant ( kato et al . 3 ) , the protease needs to be imported from the cytosol . in plastids sequestered by the slugs feeding only on one algal species , this could explain why ltr species that feed on more than one algal species do not retain the ulvophyte bryopsis the correlation between plastid - encoded ftsh and ltr deserves further study in algal grazing experiments . that the plastids from some algal lineages , including ulvophytes , are more robust than land plant plastids was noted 40 years ago ( giles and sarafis 1972 ) . it was suggested that such robustness , sometimes bordering on apparent plastid autonomy , might be linked to the prolonged survival of plastids in what are now called ltr slugs ( trench et al . although the reason(s ) underlying the robustness of plastids that sacoglossans sequester remained obscure , the role of light stress and photodamage to psii , in particular , has always figured prominently in the issue of sacoglossan plastid longevity ( reviewed in rumpho et al . and cruz et al . ) . ( 2010 ) recently showed , for acetabularia plastids sequestered within in e. timida , that psii recovers remarkably well subsequent to bleaching ; they furthermore noted that here we are suggesting that kleptoplast robustness and photodamage repair at psii are causally related , and that this conceivably could be attributable to only one or a few factors , with ftsh playing a pivotal role . our reasoning here is guided by the observation that protein synthesis in land plant plastids has two temporally distinct roles : 1 ) biogenesis of the photosynthetic apparatus followed by 2 ) the maintenance phase during which the repair of photodamaged psii in particular plays the most prominent role ( nixon et al . sacoglossans have no need for thylakoid biogenesis , as they acquire mature plastids , leaving the maintenance role as a possible factor . if the kleptoplasts are robust , which they are , then either the slugs actively render them robust , or their robustness is an intrinsic property , or a combination of the two . the observation that most sacoglossans simply digest all ingested plastids , regardless of their source ( hndeler et al . 2013 ) , indicates that ltr species specifically provide an environment where plastids can persist . the observation that some ltr species such as plakobranchus ingest plastids from several sources but retain only those from halimeda long - term as kleptoplasts ( christa et al . 2013 ) suggests that the ability to persist is a property intrinsic to the plastids . plakobranchus could also selectively digest some plastids faster than others , implying the existence of digestive recognition mechanisms for individual food particles . but selective digestion , even if it exists , would still not explain why some kleptoplasts can survive for so long within slugs , again pointing to plastid intrinsic properties . because ftsh connects psii repair to a plastid intrinsic property being plastid - encoded in plastids that undergo ltr , it emerges as a prime candidate for a causal factor behind plastid longevity . a clear prediction of this hypothesis is that acetabularia plastids should be particularly robust to high light intensities and recover faster from light stress , both in algae but especially in slugs , in comparison to plastids that lack ftsh genes , such as those of bryopsis . a further implication of these findings is that they might open new avenues of pursuit for the engineering of higher plant plastids with increased tolerance to light stress . genes for ftsh and tufa are absent from the plastid genomes of higher plants ( martin et al . 1998 ) , but present in the genomes of most algal plastids , including those that are currently known for being sequestered by ltr sacoglossan slugs ( fig . thus , we posit and it remains to be tested that the gene content of a. acetabulum and v. litorea plastid genomes is directly involved in ltr of kleptoplasts . by bringing along their own replenishable supply of ftsh , these plastids might be better able to service photosystem ii by removing the damaged d1 protein in kleptoplasts . hereby , they are better equipped for an extended life in a foreign cytosol than plastids that are dependent upon ftsh that is nuclear encoded and must be imported . by similar reasoning , a replenishable supply of tufa might aid sustained plastid translation , though our current focus is on ftsh . it is possible that the slugs do not directly provide any supporting functions at all in terms of proteins targeted to the organelle to help the kleptoplasts stay photosynthetically active for months in the cytosol of digestive gland cells . even if d1 replacement does not occur in kleptoplasts , kleptoplast - encoded ftsh warrants further investigation regarding the nature of plastid longevity , not only in kleptoplasts of sacoglossan slugs . the a. acetabulum di1 strain we use to maintain lab cultures of e. timida was originally obtained from prof . menzel ( bonn , germany ) and grown at a 12 h/12 h light / dark rhythm , illuminated with 25 m quanta ms in 3.7% sea water ( tropic marin ) . the plastids of a. acetabulum were isolated as previously described ( tymms and schweiger 1985 ) but with penicillin streptomycin ( 10 ml / l ) and chloramphenicol ( 200 mg / l ) added 48 h prior to the plastid isolation to reduce bacterial contamination . after disruption and filtration of the algal homogenate , the flowthrough was incubated for 30 min with lysozyme ( 2 mg / ml ) and subsequently treated with 1 mg / ml dnase ( roche ) for 1.5 h. as a final step , plastid dna was extracted using plant dnazol ( invitrogen ) . the 138,285 kb of vector - trimmed raw data ( 289,644 reads in total with a mean average read length of 477 bp ) were manually assembled in packages using sequencher v5.1 ( gene codes ) . assembly parameters for the first run were a minimum match percentage of 98 and a minimum overlap of 50 , followed by a second assembly with a minimum match percentage of 90 and identical minimum overlap . there are 1,768 contigs ( 300 bp long with 10 reads / contig ) , and for 91 of them ( average coverage of 56 ) , the best blast hits are plastid - encoded genes ( e - value better than 10 ) . only three contigs with coverage greater than 10-fold ( average coverage of 14 ) hit genes of potentially eukaryotic nuclear origin by the criterion of sequence similarity , but those three hits are sequences of low complexity . although there were several bacterial and mitochondrial sequences among our > 10 contigs , there is very little , if any , demonstrably algal nuclear contamination within our sequenced > 10 contigs ( supplementary fig . s1 , supplementary material online ) , for which reason it seems likely that our plastid - related sequences represent bona fide acetabularia plastid dna , not nuclear pseudogenes thereof ( nupts ) . contigs of 1,000 bp length were screened for genes with homology to the plastid genomes of the utc clade , and the coding sequences of identified genes ( table 1 ) deposited at european nucleotide archive ( ena ) ( hg518425 - 74 ; hg794360 ) and reads submitted to the sequence read archive ( srr1038494 ) . to generate the cladogram ( fig . 3 ) , protein sequences of plastid genomes were first downloaded from ncbi ( september 2013 ) . 2000 ) and markov cluster algorithm ( mcl ) ( enright et al . 2002 ) . based on the 50 genes assembled for a. acetabulum ( table 1 ) , 17 universal clusters ( genes ) these clusters were aligned by multiple alignment using fast fourier transform ( mafft ) ( katoh et al . 2002 ) and concatenated , followed by tree construction using phyml ( guindon et al . rna was extracted from four animals that had starved for 31 days using trizol ( life technologies ) according to the manufacturer s instructions , but with an additional dnase treatment ( thermoscientific ) . reverse transcriptase pcrs were carried out using iscript select cdna synthesis kit from biorad and the phusion high - fidelity dna polymerase ( new england biolabs ) . primers used were : ftshf 5-ctgcagaaaaggtttggaggc-3 , ftshr 5-gtccgaggggagttgacttg-3 ; psbaf 5-tgcatggcctgtaatcggaa-3 , psbar 5-cggttgataacgtcagccca-3 ; tufaf 5-gcaaaacaagttggcgttcc-3 , tufar 5-ggctaataaagcagacccgga-3. Output:	plastids sequestered by sacoglossan sea slugs have long been a puzzle . some sacoglossans feed on siphonaceous algae and can retain the plastids in the cytosol of their digestive gland cells . there , the stolen plastids ( kleptoplasts ) can remain photosynthetically active in some cases for months . kleptoplast longevity itself challenges current paradigms concerning photosystem turnover , because kleptoplast photosystems remain active in the absence of nuclear algal genes . in higher plants , nuclear genes are essential for plastid maintenance , in particular , for the constant repair of the d1 protein of photosystem ii . lateral gene transfer was long suspected to underpin slug kleptoplast longevity , but recent transcriptomic and genomic analyses show that no algal nuclear genes are expressed from the slug nucleus . kleptoplast genomes themselves , however , appear expressed in the sequestered state . here we present sequence data for the chloroplast genome of acetabularia acetabulum , the food source of the sacoglossan elysia timida , which can maintain acetabularia kleptoplasts in an active state for months . the data reveal what might be the key to sacoglossan kleptoplast longevity : plastids that remain photosynthetically active within slugs for periods of months share the property of encoding ftsh , a d1 quality control protease that is essential for photosystem ii repair . in land plants , ftsh is always nuclear encoded , it was transferred to the nucleus from the plastid genome when charophyta and embryophyta split . a replenishable supply of ftsh could , in principle , rescue kleptoplasts from d1 photodamage , thereby influencing plastid longevity in sacoglossan slugs .

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