What is the role of HPV typing in the United States now and in the next five years in a vaccinated population?
To review the current state of HPV typing of the vaccinated population in the United States and potential for typing of this population over the next 5 years. An expert forum conducted on September 12-13, 2008, by the Society of Gynecologic Oncologists including 56 experts in cervical cancer and titled "Future strategies of cervical cancer prevention: what do we need to do now to prepare?" In principle, screening with HPV DNA testing for oncogenic genotypes followed by cytologic triage has attractive features that may serve well the screening needs of a post-vaccination era in the US. Particularly in light of the recent FDA approval of a HPV genotyping test, the group focused on how typing could be used to assist clinical decisions and whether its implementation would be cost-effective. Furthermore, it was agreed upon that HPV typing should not be used to determine who should be vaccinated against HPV. There was considerable discussion regarding the potential misuse and overuse of HPV typing in low risk women among healthcare providers. As HPV typing technologies gain traction in the United States, its appropriate use will depend on the evolving natural history of the vaccinated cohort, continued educational efforts of healthcare providers, and most importantly, creating an integrated approach to cervical cancer prevention that will lead to a greater decrease in the incidence of cervical disease in the US while allowing for cost equipoise. On September 12-13, 2008, the Society of Gynecologic Oncologists (SGO) convened a symposium of 56 cervical cancer experts titled "Future strategies of cervical cancer prevention: what do we need to do now to prepare?" to discuss evidence-based strategies in cervical cancer prevention and control, including HPV vaccination. This paper is the second in a series of manuscripts which highlight concepts, information, obstacles and approaches discussed during the Forum's sessions and focuses on the current state of HPV typing of the vaccinated population in the United States and typing of this population over the next 5 years.


A microfluidic interface for the culture and sampling of adiponectin from primary adipocytes.
Secreted from adipose tissue, adiponectin is a vital endocrine hormone that acts in glucose metabolism, thereby establishing its crucial role in diabetes, obesity, and other metabolic disease states. Insulin exposure to primary adipocytes cultured in static conditions has been shown to stimulate adiponectin secretion. However, conventional, static methodology for culturing and stimulating adipocytes falls short of truly mimicking physiological environments. Along with decreases in experimental costs and sample volume, and increased temporal resolution, microfluidic platforms permit small-volume flowing cell culture systems, which more accurately represent the constant flow conditions through vasculature in vivo. Here, we have integrated a customized primary tissue culture reservoir into a passively operated microfluidic device made of polydimethylsiloxane (PDMS). Fabrication of the reservoir was accomplished through unique PDMS "landscaping" above sampling channels, with a design strategy targeted to primary adipocytes to overcome issues of positive cell buoyancy. This reservoir allowed three-dimensional culture of primary murine adipocytes, accurate control over stimulants via constant perfusion, and sampling of adipokine secretion during various treatments. As the first report of primary adipocyte culture and sampling within microfluidic systems, this work sets the stage for future studies in adipokine secretion dynamics.


Nucleic acids as cofactors for factor XI and prekallikrein activation: Different roles for high-molecular-weight kininogen.
The plasma zymogens factor XI (fXI) and prekallikrein (PK) are activated by factor XIIa (fXIIa) during contact activation. Polyanions such as DNA and RNA may contribute to thrombosis and inflammation partly by enhancing PK and fXI activation. We examined PK and fXI activation in the presence of nucleic acids, and determine the effects of the cofactor high molecular weight kininogen (HK) on the reactions. In the absence of HK, DNA and RNA induced fXI autoactivation. Proteases known to activate fXI (fXIIa and thrombin) did not enhance this process appreciably. Nucleic acids had little effect on PK activation by fXIIa in the absence of HK. HK had significant but opposite effects on PK and fXI activation. HK enhanced fXIIa activation of PK in the presence of nucleic acids, but blocked fXI autoactivation. Thrombin and fXIIa could overcome the HK inhibitory effect on autoactivation, indicating these proteases are necessary for nucleic acid-induced fXI activation in an HK-rich environment such as plasma. In contrast to PK, which requires HK for optimal activation, fXI activation in the presence of nucleic acids depends on anion binding sites on the fXI molecule. The corresponding sites on PK are not necessary for PK activation. Our results indicate that HK functions as a cofactor for PK activation in the presence of nucleic acids in a manner consistent with classic models of contact activation. However, HK has, on balance, an inhibitory effect on nucleic acid-supported fXI activation and may function as a negative regulator of fXI activation.


Porphyria cutanea tarda: clinical and laboratory features.
Eleven patients with porphyria cutanea tarda were studied. Biochemical confirmation of the clinical diagnosis required only determination of the total urine porphyrin concentration in a sample of urine voided on rising in the morning. The patients were divided for convenience of discussion into four groups differing in age, sex and etiologic factors. Of the six patients in whom a liver biopsy was done one was shown to have micronodular cirrhosis. Except for a modest elevation in the serum glutamic oxaloacetic transaminase values when the patients were first seen, no evidence was found for liver disease apart from the presence of porphyria cutanea tarda. One patient recovered solely by abstaining from alcohol consumption. Five patients underwent phlebotomy; their iron stores had been found to be between 2 and 3 g. Decreasing urine porphyrin values correlated well with decreasing serum ferritin values during the course of phlebotomy. Porphyria cutanea tarda, which is due to a deficiency of uroporphyrinogen decarboxylase, is manifested in association with alcohol abuse, estrogen therapy, exposure to chlorinated hydrocarbons or increased tissue iron stores, or a combination of these factors. Although relatively uncommon, this condition raises important and unresolved issues regarding the hepatotoxicity of alcohol, estrogens, chlorinated hydrocarbons and iron.


Psychometric characteristics of the mutuality scale in heart failure patients and caregivers.
Higher patient-caregiver mutuality is associated with improved patient and caregiver outcomes, but no studies have tested the psychometric characteristics of the mutuality scale (MS) in heart failure (HF) patient and caregiver population. To test the validity and reliability of the MS. A cross-sectional design. The MS validity and reliability were tested with confirmatory factor analysis (CFA) and hypothesis testing, and with Cronbach's alpha and model-based internal consistency index, respectively. CFA supported the validity of the MS in the HF patient and caregiver versions. Hypothesis testing showed significant correlations between both versions of the MS and anxiety, depression, quality of life, and self-care. Also, MS caregiver version scores correlated significantly with caregiver preparedness. Cronbach's alphas and the model-based internal consistency index ranged between 0.72 and 0.94 in both versions. The Mutuality Scale showed supportive validity and reliability for HF patients and caregivers.


Sumatriptan decreases food intake and increases plasma growth hormone in healthy women.
We studied the effect of the 5-HT1B/ID receptor agonist sumatriptan (6 mg s.c.) on plasma growth hormone and prolactin and food intake in 15 healthy female subjects using a double-blind, placebo-controlled, cross-over design. Sumatriptan significantly elevated plasma growth hormone but did not alter plasma prolactin. Sumatriptan also significantly lowered total food intake in a buffet meal, particularly decreasing the intake of fat. Our results indicate that 5-HT1B/ID receptors may be involved in the regulation of food intake in humans. In addition, while activation of 5-HT1B/ID receptors stimulates growth hormone release in both men and women, sumatriptan lowers plasma prolactin only in men, suggesting sex differences in the 5-HT regulation of prolactin release.


Nivolumab-induced autoimmune diabetes mellitus presenting as diabetic ketoacidosis in a patient with metastatic lung cancer.
Advances in cancer immunotherapy have generated encouraging results in multiple malignancies refractory to standard chemotherapies. As the use of immune checkpoint inhibitors (ICI) proliferates, the incidence of autoimmune side effects associated with these agents, termed immune related adverse events (irAE), is expected to increase. The frequency of significant irAE in ICI treated patients is about 10-20% and early recognition is critical to prevent serious morbidity and even mortality. New onset autoimmune diabetes mellitus (DM) associated with immune checkpoint inhibitor treatment is extremely rare, occurring in less than 1% of patients. Autoimmune DM often presents as diabetic ketoacidosis, a medical emergency requiring immediate treatment. We describe the first reported case of a patient with lung cancer who developed autoimmune diabetes after nivolumab treatment and was found to have three diabetes related (islet) autoantibodies present before ICI treatment and seroconversion of another after ICI treatment and onset of autoimmune DM. A 34 year old African American woman with metastatic non-small cell lung cancer (NSCLC) was treated with nivolumab in the second line setting after disease progression following standard chemoradiation therapy. After receiving two doses of nivolumab, the patient developed abrupt onset of hyperglycemia and diabetic ketoacidosis. Autoimmune diabetes was diagnosed on the basis of undetectable C-peptide levels, seropositivity of three diabetes related (islet) autoantibodies and absolute insulin dependence. The patient eventually required use of continuous subcutaneous insulin infusion (insulin pump) due to erratic glycemic excursions and multiple readmissions for DKA. Human leucocyte antigen (HLA) genoyping revealed none of the high risk haplotypes associated with the development of type 1 diabetes. Interestingly, a frozen blood sample obtained prior to treatment with nivolumab tested positive for three of the four diabetes related (islet) autoantibodies despite no prior history of diabetes and no family history of diabetes. Notably, at the time of manuscript preparation, the patient is without evidence of NSCLC recurrence with no further treatment since the nivolumab therapy. New onset autoimmune diabetes mellitus associated with nivolumab has been described only in case reports and occurs at rates of < 1% in the large clinical trials which garnered FDA approval in the second line setting for NSCLC. As ICI use continues to expand across a wide variety of malignancies, clinicians must maintain a high index of suspicion for irAE, including autoimmune DM and other endocrinopathies. A multidisciplinary team and thorough education of the patient are recommended to optimize management of new onset adult autoimmune DM. Our patient may have been at greater risk for the development of ICI related autoimmune diabetes due to the presence of three diabetes related autoantibodies prior to therapy; however, about half of the reported cases of autoimmune DM after anti-PD-1 therapy occurred in patients with no detectable diabetes related autoantibodies. Further studies are needed to delineate genetic and immunologic biomarkers that may be useful in identifying patients at risk of developing ICI related autoimmune DM.


The diagnostic utility of a genetics evaluation in children with pervasive developmental disorders.
A genetics evaluation of children with pervasive developmental disorders (PDDs) identifies a diagnosis in 6% to 15% of cases. However, previous studies have not measured the incidence of genetic disorders among children with autistic-like features who do not necessarily meet the Diagnostic and Statistical Manual for Mental Disorders, Fourth Edition criteria for PDD. We identified 101 patients at our institution referred for PDD, autism, Asperger syndrome, or autistic features. Seventy-eight were males and 23 were females, giving a male-to-female ratio of 3.4:1. No diagnosis was identified on examination alone, although Rett syndrome was suspected in six females. Seventeen patients did not undergo any type of testing because of noncompliance. Of the remaining 84 patients analyzed, seven (8.3%) were found to have abnormalities on testing. Three chromosomal anomalies were found: one with 5p duplication, one with low-level mosaicism for trisomy 21, and one with an unbalanced 10;22 translocation. Three females were diagnosed with Rett syndrome after MECP2 analysis identified a disease-causing mutation. The remaining patient was found to have an elevated urine orotic acid, with a normal ammonia level, of unknown significance. On the basis of our series, the yield of a genetics evaluation in patients with features of PDD who do not necessarily meet the Diagnostic and Statistical Manual for Mental Disorders, Fourth Edition criteria is 8.3%. Approximately half of these were the result of a chromosomal abnormality. Three cases of Rett syndrome were identified for which autistic behaviors are a well-described feature. These findings suggest that a high-resolution karyotype provides the greatest diagnostic yield for patients with autistic-like features. MECP2 analysis should be considered for females who present with autistic behaviors.


Molecular Mechanisms of Takotsubo Syndrome.
Takotsubo syndrome (TTS) is a severe but reversible acute heart failure syndrome that occurs following high catecholaminergic stress. TTS patients are similar to those with acute coronary syndrome, with chest pain, dyspnoea and ST segment changes on electrocardiogram, but are characterised by apical akinesia of the left ventricle, with basal hyperkinesia in the absence of culprit coronary artery stenosis. The pathophysiology of TTS is not completely understood and there is a paucity of evidence to guide treatment. The mechanisms of TTS are thought to involve catecholaminergic myocardial stunning, microvascular dysfunction, increased inflammation and changes in cardiomyocyte metabolism. Here, we summarise the available literature to focus on the molecular basis for the pathophysiology of TTS to advance the understanding of the condition.


Setting health research priorities using the CHNRI method: VII. A review of the first 50 applications of the CHNRI method.
Several recent reviews of the methods used to set research priorities have identified the CHNRI method (acronym derived from the "Child Health and Nutrition Research Initiative") as an approach that clearly became popular and widely used over the past decade. In this paper we review the first 50 examples of application of the CHNRI method, published between 2007 and 2016, and summarize the most important messages that emerged from those experiences. We conducted a literature review to identify the first 50 examples of application of the CHNRI method in chronological order. We searched Google Scholar, PubMed and so-called grey literature. Initially, between 2007 and 2011, the CHNRI method was mainly used for setting research priorities to address global child health issues, although the first cases of application outside this field (eg, mental health, disabilities and zoonoses) were also recorded. Since 2012 the CHNRI method was used more widely, expanding into the topics such as adolescent health, dementia, national health policy and education. The majority of the exercises were focused on issues that were only relevant to low- and middle-income countries, and national-level applications are on the rise. The first CHNRI-based articles adhered to the five recommended priority-setting criteria, but by 2016 more than two-thirds of all conducted exercises departed from recommendations, modifying the CHNRI method to suit each particular exercise. This was done not only by changing the number of criteria used, but also by introducing some entirely new criteria (eg, "low cost", "sustainability", "acceptability", "feasibility", "relevance" and others). The popularity of the CHNRI method in setting health research priorities can be attributed to several key conceptual advances that have addressed common concerns. The method is systematic in nature, offering an acceptable framework for handling many research questions. It is also transparent and replicable, because it clearly defines the context and priority-setting criteria. It is democratic, as it relies on "crowd-sourcing". It is inclusive, fostering "ownership" of the results by ensuring that various groups invest in the process. It is very flexible and adjustable to many different contexts and needs. Finally, it is simple and relatively inexpensive to conduct, which we believe is one of the main reasons for its uptake by many groups globally, particularly those in low- and middle-income countries.


Differential patterns of evolution and distribution of the symbiotic behaviour in nostocacean cyanobacteria.
Many cyanobacteria commonly identified as belonging to the genus Nostoc are well-known cyanobionts (symbionts) of a wide variety of plants and fungi. They form symbioses with bryophytes, pteridophytes, gymnosperms and angiosperms that are considerably different in the type of reciprocal interaction between the host and the cyanobiont. The phylogenetic and taxonomic relationships among cyanobionts isolated from different hosts and Nostoc strains isolated from free-living conditions are still not well understood. We compared phylogeny and morphology of symbiotic cyanobacteria originating from different host plants (genera Gunnera, Azolla, Cycas, Dioon, Encephalartos, Macrozamia and Anthoceros) with free-living Nostoc isolates originating from different habitats. After preliminary clustering with ARDRA (amplified rDNA restriction analysis), phylogeny was reconstructed on the basis of 16S rRNA gene sequences and compared with morphological characterization, obtaining several supported clusters. Two main Nostoc clusters harboured almost all cyanobionts of Gunnera, Anthoceros and of several cycads, together with free-living strains of the species Nostoc muscorum, Nostoc calcicola, Nostoc edaphicum, Nostoc ellipsosporum and strains related to Nostoc commune. We suggest that the frequent occurrence of symbiotic strains within these clusters is explained by the intensive hormogonia production that was observed in many of the strains studied. However, no evidence for discrimination between symbiotic and free-living strains, either by molecular or morphological approaches, could be found. Sequences of Azolla cyanobiont filaments, taken directly from leaf cavities, clustered tightly with sequences from the planktic cyanobacterium Cylindrospermopsis raciborskii, from the benthic Anabaena cylindrica 133 and from Anabaena oscillarioides HINDAK 1984/43, with high bootstrap values. The phylogenetic analysis showed that two distinct patterns of evolution of symbiotic behaviour might exist for the nostocacean cyanobacteria, one leading to symbioses of Nostoc species with a wide variety of plants, the other leading to the association of a unique cyanobacterial type with the water fern Azolla.


Adiponectin corrects high-fat diet-induced disturbances in muscle metabolomic profile and whole-body glucose homeostasis.
We provide here a detailed and comprehensive analysis of skeletal muscle metabolomic profiles in response to adiponectin in adiponectin knockout (AdKO) mice after high-fat-diet (HFD) feeding. Hyperinsulinemic-euglycemic clamp studies showed that adiponectin administration corrected HFD-induced defects in post/basal insulin stimulated R(d) and insulin signaling in skeletal muscle. Lipidomic profiling of skeletal muscle from HFD-fed mice indicated elevated triacylglycerol and diacylglycerol species (16:0-18:1, 18:1, and 18:0-18:2) as well as acetyl coA, all of which were mitigated by adiponectin. HFD induced elevated levels of various ceramides, but these were not significantly altered by adiponectin. Adiponectin corrected the altered branched-chain amino acid metabolism caused by HFD and corrected increases across a range of glycerolipids, fatty acids, and various lysolipids. Adiponectin also reversed induction of the pentose phosphate pathway by HFD. Analysis of muscle mitochondrial structure indicated that adiponectin treatment corrected HFD-induced pathological changes. In summary, we show an unbiased comprehensive metabolomic profile of skeletal muscle from AdKO mice subjected to HFD with or without adiponectin and relate these to changes in whole-body glucose handling, insulin signaling, and mitochondrial structure and function. Our data revealed a key signature of relatively normalized muscle metabolism across multiple metabolic pathways with adiponectin supplementation under the HFD condition.


Genistein Induces Alterations of Epigenetic Modulatory Signatures in Human Cervical Cancer Cells.
Epidemiological studies indicate that diet rich in fruits and vegetables is associated with decreased cancer risk thereby indicating that dietary polyphenols can be potential chemo-preventive agents. The reversible nature of epigenetic modifications makes them a favorable target for cancer prevention. Polyphenols have been shown to reverse aberrant epigenetic patterns by targeting the regulatory enzymes, DNA methyltransferases (DNMTs) and histone deacetylases (HDACs). In vitro and in silico studies of DNMTs and HDACs were planned to examine genistein's role as a natural epigenetic modifier in human cervical cancer cells, HeLa. Expression of the tumour suppressor genes (TSGs) [MGMT, RARβ, p21, E-cadherin, DAPK1] as well the methylation status of their promoters were examined alongwith the activity levels of DNMT and HDAC enzymes after treatment with genistein. Expression of DNMTs and HDACs was also studied. In-silico studies were performed to determine the interaction of genistein with DNMTs and HDACs. Genistein treatment significantly reduced the expression and enzymatic activity of both DNMTs and HDACs in a time-dependent way. Molecular modeling data suggest that genistein can interact with various members of DNMT and HDAC families and support genistein mediated inhibition of their activity. Timedependent exposure of genistein reversed the promoter region methylation of the TSGs and re-established their expression. In this study, we find that genistein is able to reinstate the expression of the TSGs studied by inhibiting the action of DNMTs and HDACs. This shows that genistein could be an important arsenal in the development of epigenetic based cancer therapy.


Influence of High Intensity Focused Ultrasound on the Microstructure and c-di-GMP Signaling of Pseudomonas aeruginosa Biofilms.
Bacterial biofilms are typically more tolerant to antimicrobials compared to bacteria in the planktonic phase and therefore require alternative treatment approaches. Mechanical biofilm disruption from ultrasound may be such an alternative by circumventing rapid biofilm adaptation to antimicrobial agents. Although ultrasound facilitates biofilm dispersal and may enhance the effectiveness of antimicrobial agents, the resulting biological response of bacteria within the biofilms remains poorly understood. To address this question, we investigated the microstructural effects of Pseudomonas aeruginosa biofilms exposed to high intensity focused ultrasound (HIFU) at different acoustic pressures and the subsequent biological response. Confocal microscopy images indicated a clear microstructural response at peak negative pressures equal to or greater than 3.5 MPa. In this pressure amplitude range, HIFU partially reduced the biomass of cells and eroded exopolysaccharides from the biofilm. These pressures also elicited a biological response; we observed an increase in a biomarker for biofilm development (cyclic-di-GMP) proportional to ultrasound induced biofilm removal. Cyclic-di-GMP overproducing mutant strains were also more resilient to disruption from HIFU at these pressures. The biological response was further evidenced by an increase in the relative abundance of cyclic-di-GMP overproducing variants present in the biofilm after exposure to HIFU. Our results, therefore, suggest that both physical and biological effects of ultrasound on bacterial biofilms must be considered in future studies.


Miscible Polyether/Poly(ether-acetal) Electrolyte Blends.
This study shows that it is possible to obtain homogeneous mixtures of two chemically distinct polymers with a lithium salt for electrolytic applications. This approach is motivated by the success of using mixtures of organic solvents in modern lithium-ion batteries. The properties of mixtures of a polyether, poly(ethylene oxide) (PEO), a poly(ether-acetal), poly(1,3,6-trioxocane) (P(2EO-MO)), and lithium bis(trifluoromethanesulfonyl)imide (LiTFSI) salt were studied by small-angle neutron scattering (SANS) and electrochemical characterization in symmetric cells. The SANS data are used to determine the miscibility window and quantify the effect of added salt on the thermodynamic interactions between the polymers. In the absence of salt, PEO/P(2EO-MO) blends are homogeneous and characterized by attractive interactions, i.e., a negative Flory-Huggins interaction parameter, χ. The addition of small amounts of salt results in a positive effective Flory-Huggins interaction parameter, χ eff, and macrophase separation. Surprisingly, miscible blends and negative χ eff parameters are obtained when the salt concentration is increased beyond a critical value. The electrochemical properties of PEO/P(2EO-MO)/LiTFSI blends at a given salt concentration were close to those obtained in PEO/LiTFSI electrolytes at the same salt concentration. This suggests that in the presence of PEO the electrochemical properties exhibited by P(2EO-MO) chains are similar to those of PEO chains. This work opens the door to a new direction for creating new and improved polymer electrolytes either by combining existing polymers and salt or by synthesizing new polymers with the specific aim of including them in miscible polymer blend electrolytes.


Interactions between sire family and production environment (temperate vs. tropical) on performance and thermoregulation responses in growing pigs.
The aim of this study was to evaluate the effect of 2 climatic environments (temperate [TEMP] vs. tropical humid [TROP]) on production and thermoregulation traits in growing pigs. A backcross design involving Large White (LW; heat sensitive) and Creole (CR; heat tolerant) pigs was studied. The same 10 F LW × CR boars were mated with related LW sows in each environment. A total of 1,298 backcross pigs ( = 634 pigs from 11 batches for the TEMP environment and = 664 pigs from 12 batches for the TROP environment) were phenotyped on BW (every 15 d from wk 11 to 23 of age), voluntary feed intake (ADFI, from wk 11 to 23), backfat thickness (BFT; at wk 19 and 23), skin temperature (ST; at wk 19 and 23), and rectal temperature (RT; at wk 19, 21, and 23). The feed conversion ratio was computed for the whole test period (11 to 23 wk). The calculation of the temperature-humidity index showed an average difference of 2.4°C between the TEMP and TROP environments. The ADG and ADFI were higher in the TEMP environment than in the TROP environment (834 vs. 754 g/d and 2.20 vs. 1.80 kg/d, respectively; < 0.001). Body temperatures were higher in the TROP environment than in the TEMP environment (35.9 vs. 34.8°C for ST and 39.5 vs. 39.3°C for RT, respectively; < 0.001). Most of the studied traits (i.e., BW, BFT, ADG, ADFI, and RT) were affected by sire family × environment interactions ( < 0.05), resulting in "robust" and "sensitive" families. Our results show a family dependency in the relationships between heat resistance and robustness, suggesting the possibility of finding genotypes with high production and low heat sensitivity. Further research is needed to confirm the genetic × environment interaction and to detect QTL related to heat tolerance.


[Preventing anterior cruciate ligament injury in children is effective].
An anterior cruciate ligament (ACL) injury in children is a devastating injury. After an ACL injury 1 in 3 children obtain a second injury of the ipsilateral or contralateral ACL. Children who suffer an ACL injury also have a ten times higher risk of osteoarthritis. Preventative training programmes can decrease the risk of acute knee injuries in young sportspeople; however, implementation of these prevention programmes is challenging, so it is important to inform associations, clubs, youth trainers and parents about the added value of these programmes. Children with ACL injuries must receive specialized guidance during rehabilitation, regardless of whether they have been treated conservatively or surgically. Because of the risk of a second ACL injury,we recommend that children should not to return to pivoting sports until at least 12 months after surgery for ACL.


Dual functions of wingless in the Drosophila leg imaginal disc.
The Drosophila gene wingless is a member of the Wnt gene family, a group of genes that are involved in embryonic development and the regulation of cell proliferation. wingless encodes a secreted glycoprotein that plays a role in embryogenesis as well as in the development of adult structures. In the primordia of the adult limbs, the imaginal discs, wingless is expressed in an anterior ventral sector and is required for specification of ventral fate. Ectopic expression of low levels of Wingless in the leg discs leads to partial ventralization and outgrowths of the proximodistal axis. Wingless has thus been proposed to specify ventral fate in a concentration dependent manner (i.e., as a morphogen) and to organize the proximodistal axis. We have extended the analysis of Wingless function in the leg primordium through targeted ectopic expression. We find that Wingless has two functions in the leg disc. In the specification of ventral fate, our data indicate that Wingless does not function as a morphogen but instead appears to collaborate with other factors. In addition to its role in ventral fate specification, Wingless inhibits the commitment of dorsal cells toward a determined state and influences the regulation of proliferation. We propose a model in which Wingless achieves separate functions via spatially regulated mechanisms and discuss the significance of these functions during axial patterning and organization.


A study on risk factors of breast cancer among patients attending the tertiary care hospital, in udupi district.
Cancer has become one of the ten leading causes of death in India. Breast cancer is the most common diagnosed malignancy in India, it ranks second to cervical cancer. An increasing trend in incidence is reported from various registries of national cancer registry project and now India is a country with largest estimated number of breast cancer deaths worldwide. To study the factors associated with breast cancer. To study the association between breast cancer and selected exposure variables and to identify risk factors for breast cancer. A hospital based Case control study was conducted at Shirdi Sai Baba Cancer Hospital and Research Center, Manipal, Udupi District. Total 188 participants were included in the study, 94 cases and 94 controls. All the study participants were between 25 to 69 years of age group. The cases and controls were matched by ± 2 years age range. Non vegetarian diet was one of the important risk factors (OR 2.80, CI 1.15-6.81). More than 7 to 12 years of education (OR 4.84 CI 1.51-15.46) had 4.84 times risk of breast cancer as compared with illiterate women. The study suggests that non vegetarian diet is the important risk factor for Breast Cancer and the risk of Breast Cancer is more in educated women as compared with the illiterate women. This is a Hospital based study so generalisability of the findings could be limited.


Microbial N2O consumption in and above marine N2O production hotspots.
The ocean is a net source of N2O, a potent greenhouse gas and ozone-depleting agent. However, the removal of N2O via microbial N2O consumption is poorly constrained and rate measurements have been restricted to anoxic waters. Here we expand N2O consumption measurements from anoxic zones to the sharp oxygen gradient above them, and experimentally determine kinetic parameters in both oxic and anoxic seawater for the first time. We find that the substrate affinity, O2 tolerance, and community composition of N2O-consuming microbes in oxic waters differ from those in the underlying anoxic layers. Kinetic parameters determined here are used to model in situ N2O production and consumption rates. Estimated in situ rates differ from measured rates, confirming the necessity to consider kinetics when predicting N2O cycling. Microbes from the oxic layer consume N2O under anoxic conditions at a much faster rate than microbes from anoxic zones. These experimental results are in keeping with model results which indicate that N2O consumption likely takes place above the oxygen deficient zone (ODZ). Thus, the dynamic layer with steep O2 and N2O gradients right above the ODZ is a previously ignored potential gatekeeper of N2O and should be accounted for in the marine N2O budget.


Regulation of cell-type-specific interleukin-2 receptor alpha-chain gene expression: potential role of physical interactions between Elf-1, HMG-I(Y), and NF-kappa B family proteins.
The interleukin 2 receptor alpha-chain (IL-2R alpha) gene is rapidly and potently induced in T cells in response to mitogenic stimuli. Previously, an inducible enhancer between nucleotides -299 and -228 that contains NF-kappa B and CArG motifs was identified. We now report the characterization of a second essential positive regulatory element located between nucleotides -137 and -64 that binds Elf-1 and HMG-I(Y). This element had maximal activity in lymphoid cells, paralleling the cell type specificity of Elf-1 expression. Transcription from the IL-2R alpha promoter was inhibited when either the Elf-1 or the HMG-I(Y) binding site was mutated. Coexpression of both proteins activated transcription of the -137 to -64 element in COS-7 cells. Elf-1 physically associated with HMG-I and with NF-kappa B p50 and c-Rel in vitro, suggesting that protein-protein interactions might functionally coordinate the actions of the upstream and downstream positive regulatory elements. This is the first report of a physical interaction between an Ets family member and NF-kappa B family proteins. These findings provide significant new insights into the protein-protein and protein-DNA interactions that regulate cell-type-specific and inducible IL-2R alpha gene expression and also have implications for other genes regulated by Elf-1 and NF-kappa B family proteins.


Changing spectrum of infective endocarditis in children: a 30 years experiences from a tertiary care center in Taiwan.
The epidemiology of infective endocarditis (IE) changes with the medical advances. This study aimed to evaluate the trends in a pediatric cohort. From hospital database (1983-2011), patients <18 years who fulfilled the modified Duke criteria of IE were identified. We enrolled 112 patients (M/F 57/55) with 116 IE episodes. About 86 patients (74.1%) had preexisting cardiac lesions and 23 patients (19.6%) were immunocompromised hosts. Prior dental procedure was noted in 12 (10.3%) patients, including 4 with simple ventricular septal defect. The overall mortality was 10.7%. The risk factors included vegetations in both ventricles (odds ratio = 7.81, P = .019) and prior use of broad-spectrum antibiotics (odds ratio = 3.75, P = .055). Approximately one-third of the patients (29.3%) required surgical intervention. We identified an increasing trend in the proportion of hospital-acquired IE (from 12% during 1983-1991 to 39% during 2002-2011), and the spectrum of offending pathogens showed a trend for fewer Streptococcus species, more Staphylococcus aureus and increased pathogen diversity. The leading pathogens were Gram-negative bacilli in hospital-acquired IE and Streptococcus species in community-acquired IE. Hospital-acquired IE was associated with younger age, a higher proportion of immunocompromised patients, a history of central line indwelling and higher mortality. In contrast, more surgical intervention and embolic events occurred in community-acquired IE patients. The mortality of pediatric IE remains high. Dental procedures were noted in one-tenth of the patients. Although increased S. aureus-caused episodes and pathogen diversity were noted, Streptococcus species remain the most common pathogen.


Structure of subtilosin A, a cyclic antimicrobial peptide from Bacillus subtilis with unusual sulfur to alpha-carbon cross-links: formation and reduction of alpha-thio-alpha-amino acid derivatives.
The complete primary and three-dimensional solution structures of subtilosin A (1), a bacteriocin from Bacillus subtilis, were determined by multidimensional NMR studies on peptide produced using isotopically labeled [(13)C,(15)N]medium derived from Anabaena sp. grown on sodium [(13)C]bicarbonate and [(15)N]nitrate. Additional samples of 1 were also generated by separate incorporations of [U-(13)C,(15)N]-L-phenylalanine and [U-(13)C,(15)N]-L-threonine using otherwise unlabeled media. The results demonstrate that in addition to having a cyclized peptide backbone (amide between N and C termini), three cross-links are formed between the sulfurs of Cys13, Cys7, and Cys4 and the alpha-positions of Phe22, Thr28, and Phe31, respectively. The stereochemistry of all residues in 1 except for the three modified ones was confirmed to be L by complete desulfurization with nickel boride, acid hydrolysis to the constituent amino acids, and conversion of these to the corresponding pentafluoropropanamide isopropyl esters for chiral GC MS analysis. The stereochemistry at the modified residues was determined by subjecting each of the eight possible stereoisomers of 1 to eight rounds of ARIA structure calculations, starting with the same NMR peak files and assignments. The stereoisomer with the l stereochemistry at Phe22 (alpha-R) and d stereochemistry at Thr28 (alpha-S) and Phe31 (alpha-S) (LDD isomer) fit the NMR data, giving the lowest energy family of structures with the best rmsd. Thus, biochemical formation of the unusual thio links proceeds with net retention of configuration at Phe22, and inversion at Thr28 and Phe31. Model amino acid derivatives bearing a sulfide moiety at the alpha-carbon were synthesized by reaction of the corresponding alpha-alkoxy compounds with benzyl thiol and SnCl(4). Separation of their pure stereoisomers and desulfurization with nickel boride demonstrated that the reduction of such compounds proceeds with epimerization, in contrast to the previously reported retention of stereochemistry for analogous reaction of steroidal sulfides. However, desulfurization of subtilosin A to cyclic peptide 14, which is inactive as an antimicrobial agent, occurs with inversion of stereochemistry at the alpha-carbons of Phe22 and Thr28 and with 4:1 retention at Phe31. This indicates that the desulfurization reaction proceeds via an N-acyl imine and that the structure of the surrounding peptide controls the geometry of reduction. Posttranslational linkage of a thiol to the alpha-carbon of an amino acid residue is unprecedented in ribosomally synthesized peptides or proteins, and very rare in secondary metabolites. Subtilosin A (1) represents a new class of bacteriocins.


Assessment of Coronary Flow Velocity Reserve in the Left Main Trunk Using Phase-contrast MR Imaging at 3T: Comparison with 15O-labeled Water Positron Emission Tomography.
The aim of this study was to verify coronary flow velocity reserve (CFVR) on the left main trunk (LMT) in comparison with myocardial flow reserve (MFR) by 15O-labeled water positron emission tomography (PET) (MFR-PET) in both the healthy adults and the patients with coronary artery disease (CAD), and to evaluate the feasibility of CFVR to detect CAD. Eighteen healthy adults and 13 patients with CAD were evaluated. CFVR in LMT was estimated by 3T magnetic resonance imaging (MRI) with phase contrast technique. MFR-PET in the LMT territory including anterior descending artery and circumflex artery was calculated as the ratio of myocardial blood flow (MBF)-PET at stress to MBF-PET at rest. There was a significant positive relationship between CFVR and MFR-PET (R = 0.45, P < 0.0001). Inter-observer calculations of CFVR showed good correlation (R2 = 0.93, P < 0.0001). The CFVR in patients with CAD was significantly lower than that in healthy adults (1.90 ± 0.61 vs. 2.77 ± 1.03, respectively, P = 0.01), which were similar to the results of MFR-PET (2.23 ± 0.84 vs. 3.96 ± 1.04, respectively, P < 0.0001). For the detection of patients with CAD, the area under the curve was 0.78 (P = 0.01). The sensitivity was 0.77 and specificity was 0.72 when a cut-off of 2.15 was used. CFVR by 3T was validated with MFR-PET. CFVR could detect the patients with CAD. This method is a simple and reliable index without radiation or contrast material.


How species respond to multiple extinction threats.
It is well established that different species vary in their vulnerability to extinction risk and that species biology can underpin much of this variation. By contrast, very little is known about how the same species responds to different threat processes. The purpose of this paper is therefore twofold: to examine the extent to which a species' vulnerability to different types of threat might covary and to explore the biological traits that are associated with threat-specific responses. We use an objective and quantitative measure of local extinction risk to show that vulnerability to local population decline in primates varies substantially among species and between threat types. Our results show that a species' response to one threat type does not predict its response to others. Multivariate analyses also suggest that different mechanisms of decline are associated with each type of threat, since different biological traits are correlated with each threat-specific response. Primate species at risk from forestry tend to exhibit low ecological flexibility, while those species vulnerable to agriculture tend to live in the canopy and eat low-fruit diets; in further contrast, primates at risk from hunting tend to exhibit large body size. Our analyses therefore indicate that a species' vulnerability to local extinction can be highly variable and is likely to depend on both threat type and biology.


Design and validation of a tool for neurite tracing and analysis in fluorescence microscopy images.
For the investigation of the molecular mechanisms involved in neurite outgrowth and differentiation, accurate and reproducible segmentation and quantification of neuronal processes are a prerequisite. To facilitate this task, we developed a semiautomatic neurite tracing technique. This article describes the design and validation of the technique. The technique was compared to fully manual delineation. Four observers repeatedly traced selected neurites in 20 fluorescence microscopy images of cells in culture, using both methods. Accuracy and reproducibility were determined by comparing the tracings to high-resolution reference tracings, using two error measures. Labor intensiveness was measured in numbers of mouse clicks required. The significance of the results was determined by a Student t-test and by analysis of variance. Both methods slightly underestimated the true neurite length, but the differences were not unanimously significant. The average deviation from the true neurite centerline was a factor 2.6 smaller with the developed technique compared to fully manual tracing. Intraobserver variability in the respective measures was reduced by a factor 6.0 and 23.2. Interobserver variability was reduced by a factor 2.4 and 8.8, respectively, and labor intensiveness by a factor 3.3. Providing similar accuracy in measuring neurite length, significantly improved accuracy in neurite centerline extraction, and significantly improved reproducibility and reduced labor intensiveness, the developed technique may replace fully manual tracing methods.


Endogenous opiate peptides may limit norepinephrine release during hemorrhage.
The involvement of the sympathetic nervous system in the cardiovascular response to hemorrhage and subsequent opiate receptor blockade was studied in conscious rabbits. Plasma catecholamines were measured by high-pressure liquid chromatography to indirectly assess sympathetic activity. Arterial blood samples were drawn at four times during the experiment: 1) before hemorrhage; 2) after a 15% blood loss; 3) after mean arterial blood pressure decreased to less than 40 mm Hg; and 4) 2 min after an i.v. injection of naloxone (3 mg/kg) or saline. Rapid removal of 15% of the total blood volume (approximately equal to 8 ml/kg) increased heart rate and plasma norepinephrine. Plasma epinephrine and blood pressure remained at control levels. Further hemorrhage (approximately equal to 16 ml/kg) produced a sudden decrease in blood pressure and a large increase in plasma epinephrine. Plasma norepinephrine was not significantly different from the previous sample. Subsequent injection of naloxone significantly increased plasma norepinephrine and blood pressure compared to the saline-treated group. Plasma epinephrine was similar in the two groups. These studies suggest that naloxone may exert its pressor effect during hemorrhagic hypotension in the conscious rabbit by blocking a naturally occurring, opiate peptide-mediated inhibition of norepinephrine release. The results are consistent with a peptidergic limit on sympathetic activity being responsible for the decrease in blood pressure seen during acute hemorrhage.


The relationship between cardiac output and dynamic cerebral autoregulation in humans.
Cerebral autoregulation adjusts cerebrovascular resistance in the face of changing perfusion pressures to maintain relatively constant flow. Results from several studies suggest that cardiac output may also play a role. We tested the hypothesis that cerebral blood flow would autoregulate independent of changes in cardiac output. Transient systemic hypotension was induced by thigh-cuff deflation in 19 healthy volunteers (7 women) in both supine and seated positions. Mean arterial pressure (Finapres), cerebral blood flow (transcranial Doppler) in the anterior (ACA) and middle cerebral artery (MCA), beat-by-beat cardiac output (echocardiography), and end-tidal Pco(2) were measured. Autoregulation was assessed using the autoregulatory index (ARI) defined by Tiecks et al. (Tiecks FP, Lam AM, Aaslid R, Newell DW. Stroke 26: 1014-1019, 1995). Cerebral autoregulation was better in the supine position in both the ACA [supine ARI: 5.0 ± 0.21 (mean ± SE), seated ARI: 3.9 ± 0.4, P = 0.01] and MCA (supine ARI: 5.0 ± 0.2, seated ARI: 3.8 ± 0.3, P = 0.004). In contrast, cardiac output responses were not different between positions and did not correlate with cerebral blood flow ARIs. In addition, women had better autoregulation in the ACA (P = 0.046), but not the MCA, despite having the same cardiac output response. These data demonstrate cardiac output does not appear to affect the dynamic cerebral autoregulatory response to sudden hypotension in healthy controls, regardless of posture. These results also highlight the importance of considering sex when studying cerebral autoregulation.


Wide-field computational imaging of pathology slides using lens-free on-chip microscopy.
Optical examination of microscale features in pathology slides is one of the gold standards to diagnose disease. However, the use of conventional light microscopes is partially limited owing to their relatively high cost, bulkiness of lens-based optics, small field of view (FOV), and requirements for lateral scanning and three-dimensional (3D) focus adjustment. We illustrate the performance of a computational lens-free, holographic on-chip microscope that uses the transport-of-intensity equation, multi-height iterative phase retrieval, and rotational field transformations to perform wide-FOV imaging of pathology samples with comparable image quality to a traditional transmission lens-based microscope. The holographically reconstructed image can be digitally focused at any depth within the object FOV (after image capture) without the need for mechanical focus adjustment and is also digitally corrected for artifacts arising from uncontrolled tilting and height variations between the sample and sensor planes. Using this lens-free on-chip microscope, we successfully imaged invasive carcinoma cells within human breast sections, Papanicolaou smears revealing a high-grade squamous intraepithelial lesion, and sickle cell anemia blood smears over a FOV of 20.5 mm(2). The resulting wide-field lens-free images had sufficient image resolution and contrast for clinical evaluation, as demonstrated by a pathologist's blinded diagnosis of breast cancer tissue samples, achieving an overall accuracy of ~99%. By providing high-resolution images of large-area pathology samples with 3D digital focus adjustment, lens-free on-chip microscopy can be useful in resource-limited and point-of-care settings.


Cephalic version by moxibustion for breech presentation.
Moxibustion (a type of Chinese medicine which involves burning a herb close to the skin) to the acupuncture point Bladder 67 (BL67) (Chinese name Zhiyin), located at the tip of the fifth toe, has been proposed as a way of correcting breech presentation. To examine the effectiveness and safety of moxibustion on changing the presentation of an unborn baby in the breech position, the need for external cephalic version (ECV), mode of birth, and perinatal morbidity and mortality for breech presentation. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register (26 March 2012), MEDLINE (1966 to 1 August 2011), EMBASE (1980 to August 2011), CINAHL (1982 to 1 August 2011), MIDIRS (1982 to 1 August 2011) and AMED (1985 to 1 August 2011) and searched bibliographies of relevant papers. The inclusion criteria were published and unpublished randomised controlled trials comparing moxibustion (either alone or in combination with acupuncture or postural techniques) with a control group (no moxibustion), or other methods (e.g. external cephalic version, acupuncture, postural techniques) in women with a singleton breech presentation. Two review authors independently assessed eligibility and trial quality and extracted data. The outcome measures were baby's presentation at birth, need for external cephalic version, mode of birth, perinatal morbidity and mortality, maternal complications and maternal satisfaction, and adverse events. Six new trials have been added to this updated review. One trial has been moved to studies awaiting classification while further data are being requested. This updated review now includes a total of eight trials (involving 1346 women). Meta-analyses were undertaken (where possible) for the main and secondary outcomes. Moxibustion was not found to reduce the number of non-cephalic presentations at birth compared with no treatment (P = 0.45). Moxibustion resulted in decreased use of oxytocin before or during labour for women who had vaginal deliveries compared with no treatment (risk ratio (RR) 0.28, 95% confidence interval (CI) 0.13 to 0.60). Moxibustion was found to result in fewer non-cephalic presentations at birth compared with acupuncture (RR 0.25, 95% CI 0.09 to 0.72). When combined with acupuncture, moxibustion resulted in fewer non-cephalic presentations at birth (RR 0.73, 95% CI 0.57 to 0.94), and fewer births by caesarean section (RR 0.79, 95% CI 0.64 to 0.98) compared with no treatment. When combined with a postural technique, moxibustion was found to result in fewer non-cephalic presentations at birth compared with the postural technique alone (RR 0.26, 95% CI 0.12 to 0.56). This review found limited evidence to support the use of moxibustion for correcting breech presentation. There is some evidence to suggest that the use of moxibustion may reduce the need for oxytocin. When combined with acupuncture, moxibustion may result in fewer births by caesarean section; and when combined with postural management techniques may reduce the number of non-cephalic presentations at birth, however, there is a need for well-designed randomised controlled trials to evaluate moxibustion for breech presentation which report on clinically relevant outcomes as well as the safety of the intervention.


Evidence for secretory pathway localization of a voltage-dependent anion channel isoform.
Voltage-dependent anion channels (VDACs) are pore-forming proteins (porins) that form the major pathway for movement of adenine nucleotides through the outer mitochondrial membrane. Electrophysiological studies indicate that VDAC-like channel activity is also prevalent in the cell membranes of many mammalian cells. However, the multitopological localization of porins outside the mitochondrion has remained an extremely controversial issue. Herein, we show that usage of two alternative first exons of the murine VDAC-1 gene leads to expression of two porins differing within their N termini. One porin (plasmalemmal VDAC-1) harboring a hydrophobic leader peptide is primarily targeted through the Golgi apparatus to the cell membrane. In contrast, the second isoform lacking the N-terminal leader (mitochondrial VDAC-1) is translocated more efficiently into the outer mitochondrial membrane. Thus, our data provide unique genetic evidence in favor of a multitopological localization of a mitochondrial porin.


Isolate Specificity and Polygenic Inheritance of Resistance in Barley to the Heterologous Rust Pathogen Puccinia graminis f. sp. avenae.
Barley is a near-nonhost to numerous heterologous (nonadapted) rust pathogens because a small proportion of genotypes are somewhat susceptible. We assessed 66 barley accessions and three mapping populations (Vada × SusPtrit, Cebada Capa × SusPtrit, and SusPtrit × Golden Promise) for response to three Swedish oat stem rust (Puccinia graminis f. sp. avenae) fungal isolates and determined that barley is a near-nonhost to P. graminis f. sp. avenae and that resistance was polygenically inherited. The parental genotypes Vada and Golden Promise were immune to all three isolates, whereas Cebada Capa was immune to two isolates and moderately resistant to the third. Phenotypic data from the Vada × SusPtrit mapping population and the barley accessions tested also demonstrated isolate-specific resistance. In particular, the SusPtrit parent and several other accessions allowed sporulation by isolate Ingeberga but were resistant to isolate Evertsholm. Nine chromosomal regions carried quantitative trait loci (QTL) (Rpgaq1 to Rpgaq9) of varying effect, most of which colocated to previously identified QTL for resistance to other heterologous rust pathogens. Rpgaq1 on chromosome 1H (Vada and Golden Promise) was effective toward all isolates tested. Microscopic examination indicated that resistance was prehaustorial in Vada whereas, in SusPtrit, both pre- and posthaustorial mechanisms play a role.


The Body Acoustic: Ultrasonic Neuromodulation for Translational Medicine.
For the greater part of the last century, ultrasound (US) has seen widespread use in applications ranging from materials science to medicine. The history of US in medicine has also seen promising success in clinical diagnostics and regenerative medicine. Recent studies have shown that US is able to manipulate the nervous system, leading toward potential treatment for various neuropathological conditions, a phenomenon known as ultrasonic neuromodulation (NM). Ultrasonic NM is a promising alternative to pharmaceuticals and surgery, due to high spatiotemporal resolution combined with the potentially noninvasive means of application. Current advances have made progress in establishing effective dosage limits, waveform parameters, and stimulus regimes in order to achieve desired effects in a variety of tissue and cell types. However, to date there has been limited systematic analysis of the complex variables involved in creating a therapeutic US stimulation regime specifically tailored to the nervous system. Without a fundamental understanding of the effects of US on neural tissue, including the surrounding bone, musculature, and vasculature, the safety and efficacy of US as an NM tool is yet to be determined. Advances in imaging technology and focusing hardware highlight new avenues for potential clinical applications for therapeutic ultrasonic stimulation. US may be an alternative to electrical and magnetic means of NM for targets in the central nervous system as well as in the peripheral and autonomic nervous systems. This review provides a historical perspective on the past, present, and future of US as a translational therapeutic.


Expert opinion on metal chains and other indestructible objects as proper enrichment for intensively-farmed pigs.
EC Directive 2001/93 requires that all pigs have access to proper investigation and manipulation materials. Intensively farmed pigs in Europe are frequently provided with a short metal chain with or without an indestructible object attached to the chain. To date authorities are regarding this as proper enrichment, perhaps with (in)direct reference to the RICHPIG model as a justification. However, it has become increasingly clear that the chains do not provide proper enrichment, and that adding an indestructible object to the end of the chain may even reduce rather than improve pig welfare. To test this hypothesis an expert survey was conducted containing 26 more or less compound questions. On a scale from 0 to 10 experts specified their level of agreement with the hypothesis, the prevalence and welfare scores of nine indestructible enrichment materials. In total 36 experts, mostly pig-welfare scientists, responded (response rate: 39%). Indestructible objects are less prevalent in countries that provide straw (like Sweden and the UK) and outside the EU (US). They are more prevalent in the Netherlands, Belgium, France and Finland, while the prevalence seems to be low in Spain. Balls, wood and pipes were provided most frequently: hard wood especially in the UK (as specified in farm assurance); indestructible balls and pipes in Germany and the Netherlands. The experts' score for agreement with the hypothesis was only 4.6 on average (scale 0-10; n = 25). Enrichment materials, ranked from high to low welfare score, were grouped in 5 significance levels as indicated by different superscripts based on Wilcoxon signed rank tests: Branched chains (5.1a), Chain on the floor (4.4b), Hard wood (3.7bc), Pipe (3.5c), Bare chain (3.3c), Short chain (3.1d), Small ball (2.8d), Big ball (2.5d), and Chain hanging too high (1.3e). Branched chains scored significantly better than all other indestructible materials and its welfare score (5.1 on average) was close to the pre-defined level of acceptability (5.5 on a scale from 0, worst, to 10, best). The welfare benefits of adding balls, pipes or hard wood to the metal chain were marginal, and well below what the experts considered acceptable enrichment. The branched-chains design, by contrast, appears to be the most viable alternative. It involves providing a longer chain, i.e. with the free end reaching to floor level, adding 'branches', i.e. several short chains ending at the nose height of the pigs, and providing more chains per pen (i.e. 1 branched chain per 5 pigs). Branched chains should be implemented widely and in the short term as a first step towards, and benchmark for, providing proper enrichment to intensively-farmed pigs.


Skeletal muscle necrosis and regeneration after injection of BaH1, a hemorrhagic metalloproteinase isolated from the venom of the snake Bothrops asper (Terciopelo).
The effects of BaH1, a hemorrhagic metalloproteinase isolated from Bothrops asper venom, on mouse gastrocnemius muscle was investigated. The toxin induced severe hemorrhage within minutes after injection. Groups of necrotic muscle fibers were observed after the sixth hour, with evident disorganization of myofibrillar material. At the ultrastructural level myofibrils in these cells lost their regular arrangement, Z lines were absent, and sarcomeres were disoriented, although there was no evidence of myofilament clumping or hypercontraction. Plasma membrane was interrupted in many portions. Mitochondrial alterations included swelling and the formation of flocculent densities and dense intracristal plates. At 12, 24, and 48 hr necrotic cells had amorphous masses of myofilaments and abundant phagocytic cells were observed both within necrotic fibers and in the interstitial space. Fraction D-1, which contains the three hemorrhagic metalloproteinases BaH1, BH2, and BH3, did not cause direct muscle damage when incubated with gastrocnemius muscle in vitro. Upon intramuscular injection in mice this fraction induced a small but significant increment in muscle lactic acid levels. Observations carried out 7 and 14 days after BaH1 injection revealed some regenerating muscle fibers with centrally located nuclei. However, other areas had few regenerating fibers of reduced diameter, surrounded by abundant fibroblasts, fibrosis, calcification, and remnants of necrotic muscle cells. When BaH1 was injected together with B. asper myotoxin III, a myotoxic phospholipase A2 that induces myonecrosis but does not affect blood vessels, a poor muscle regeneration was observed. In contrast, if B. asper myotoxin III was injected alone, regeneration proceeded normally and successfully.


Developing a pictorial Epworth Sleepiness Scale.
The Epworth Sleepiness Scale (ESS) was designed to be self-completed by the patient. However, it may not be understood by all, and unrecognised problems with literacy can impair the process. The ESS has been translated into a pictorial version for use in those with normal or diminished literacy skills. An evaluation of the patients' ability to self-complete the ESS was undertaken in sleep and non-sleep respiratory clinics. Errors or problems encountered were recorded on a standard questionnaire. With the aid of a medical artist, pictorial representations of the eight ESS questions were developed and the new pictorial ESS was offered to patients alongside the traditional ESS. The two scales were compared for agreement with a kappa statistic, and patients were asked to record a preference for either the written or the pictorial scale. Evaluation of the traditional ESS showed that 33.8% (27/80) of ESS-naive patients made errors and 22.5% (18/80) needed help completing the questionnaire. The translated pictorial ESS showed good agreement with the traditional ESS on most questions; median kappa score 0.63, IQR 0.04. Fifty-five per cent reported a preference for the pictorial scale compared with the standard written ESS. Despite the fact that errors were frequently made on the traditional ESS, 96.8% of participants in the second study reported both scales to be easy to complete. More people (75.6%) reported the pictorial ESS to be very easy, in comparison with (64.6%) the worded ESS questionnaire. Errors are common when patients self-complete the traditional written ESS. Pictures with words have been shown to enhance the understanding and translation of medical information, and a pictorial translation of the ESS produces scores comparable with the traditional ESS and may be a suitable alternative for those with normal or diminished literacy.


Risk factors for compartment syndrome in traumatic brachial artery injuries: an institutional experience in 139 patients.
The brachial artery is the most common vascular injury encountered in upper extremity trauma. If not treated promptly, it can result in compartment syndrome (CS) and long-term disability. Here, we report an institutional experience of traumatic brachial artery injuries and establish risk factors for the development of upper extremity CS in this setting. A retrospective review of 139 patients with traumatic brachial artery injury from 1985 to 2001 at a single institution. Patients were divided into two cohorts, those with evidence of CS and those without CS (NCS), for comparison. One hundred thirty-nine patients presented with traumatic brachial artery injuries (mean age, 28.4 years). Twenty-nine patients (20.9%) were diagnosed with upper extremity CS, and 28 of these patients underwent fasciotomy on recognition of their CS. Seven patients (6.4%) in the NCS cohort underwent fasciotomy as a result of absent distal pulses on initial examination. Mean follow-up was 51.6 days. Two patients required revision of their arterial repair, and one patient underwent amputation. The risk of CS was increased in the presence of combined arterial injuries (p = 0.03), combined nerve injuries (p = 0.04), motor deficits (p < 0.0001), fractures, and increased intraoperative blood loss (p = 0.001). Multivariate logistic regression performed on these variables revealed that elevated intraoperative blood loss, combined arterial injury, and open fracture were independent risk factors for the development of CS (OR 1.12, 5.79, and 2.68, respectively). Prompt evaluation and management of traumatic brachial artery injuries is important to prevent CS, which can lead to functional deficits. In the setting of combined arterial injury, open fracture, and significant intraoperative blood loss, prophylactic fasciotomy should be considered.


The naked truth: the face and body sensitive N170 response is enhanced for nude bodies.
Recent event-related potential studies have shown that the occipitotemporal N170 component--best known for its sensitivity to faces--is also sensitive to perception of human bodies. Considering that in the timescale of evolution clothing is a relatively new invention that hides the bodily features relevant for sexual selection and arousal, we investigated whether the early N170 brain response would be enhanced to nude over clothed bodies. In two experiments, we measured N170 responses to nude bodies, bodies wearing swimsuits, clothed bodies, faces, and control stimuli (cars). We found that the N170 amplitude was larger to opposite and same-sex nude vs. clothed bodies. Moreover, the N170 amplitude increased linearly as the amount of clothing decreased from full clothing via swimsuits to nude bodies. Strikingly, the N170 response to nude bodies was even greater than that to faces, and the N170 amplitude to bodies was independent of whether the face of the bodies was visible or not. All human stimuli evoked greater N170 responses than did the control stimulus. Autonomic measurements and self-evaluations showed that nude bodies were affectively more arousing compared to the other stimulus categories. We conclude that the early visual processing of human bodies is sensitive to the visibility of the sex-related features of human bodies and that the visual processing of other people's nude bodies is enhanced in the brain. This enhancement is likely to reflect affective arousal elicited by nude bodies. Such facilitated visual processing of other people's nude bodies is possibly beneficial in identifying potential mating partners and competitors, and for triggering sexual behavior.


Coupling of beta-cell desensitization by hyperglycemia to excessive stimulation and circulating insulin in glucose-infused rats.
Nondiabetic rats were infused with glucose for 48 h to maintain moderate or marked hyperglycemia (mean blood glucose 13.2 +/- 0.7 or 22.8 +/- 0.3 mM, respectively). The two levels of hyperglycemia increased plasma insulin levels severalfold but decreased the insulin response to 27 mM glucose by 19 and 95%, respectively, versus saline infusion. Diazoxide (5 mg.kg-1.h-1), when continuously infused during the hyperglycemia protocols, completely inhibited the glucose-induced rise in plasma insulin levels. Diazoxide transformed beta-cell insensitivity to stimulation: glucose-induced insulin release was thus increased 318% after moderate hyperglycemia and 707% after marked hyperglycemia. These stimulatory effects of diazoxide were reversed by exogenous insulin infusion (8 or 2 U/24 h) in a dose-dependent manner. It is concluded that excessive beta-cell stimulation rather than glucotoxicity underlies hyperglycemia-induced beta-cell insensitivity. Effects of hyperinsulinemia can form part of the mechanisms whereby excessive stimulation affects beta-cell secretion.


World Health Organization Grade III Supratentorial Extraventricular Ependymomas in Adults: Case Series and Review of Treatment Modalities.
Supratentorial ependymomas and their anaplastic variants are relatively uncommon central nervous system neoplasms that afflict both adults and children. Discuss the clinical and pathological features of patients with anaplastic ependymomas involving an extraventricular supratentorial location and review modalities and options of treatment for those rare tumors. Whereas the treatment algorithm in the pediatric population is well established, however, treatment in the adult population is less defined. Treatment options are exposed through the author's cases and review of the literature. In our case series of two adult patients with supratentorial ependymomas World Health Organization (WHO) Grade III (anaplastic variant), patients presented in both cases in the emergency room after having a generalized tonic-clonic seizure at home the first case, and mild hemiparesis the second case. Patients underwent surgical treatment, and a gross total resection was achieved in both cases. The histopathological examination revealed a diagnosis of anaplastic ependymoma (WHO Grade III). Both patients had additional radiotherapy, and in the first case, adjuvant platinum-based chemotherapy was administered due to leptomeningeal gliomatosis. In our experience, gross total resection was achieved in all patients with supratentorial extraventricular ependymomas WHO Grade III with additional radiotherapy and platinum-based chemotherapy. Patients require initial close serial imaging follow-up. The role of chemotherapy is still uncertain but may be necessary in younger patients and in tumors that behave more like the pediatric ependymomas.


Embryonic cardiomyocytes can orchestrate various cell protective mechanisms to survive mitochondrial stress.
Whereas adult cardiomyocytes are highly susceptible to stress, cardiomyocytes in the prenatal heart appear to be rather resistant. To investigate how embryonic cardiomyocytes respond to metabolic stress in vivo, we utilized tissue mosaicism for mitochondrial dysfunction in 13.5dpc mouse hearts. The latter is based on inactivation of the X-linked gene encoding Holocytochrome c synthase (Hccs), which is essential for mitochondrial respiration. In heterozygous heart conditional Hccs knockout females (cHccs(+/-)) random X chromosomal inactivation results in a mosaic of healthy and HCCS deficient cells in the myocardium. Microarray RNA expression analyses identified genes involved in unfolded protein response (UPR) and programmed cell death as differentially expressed in cHccs(+/-) versus control embryonic hearts. Activation of the UPR is localized to HCCS deficient cardiomyocytes but does not involve ER stress pathways, suggesting that it is caused by defective mitochondria. Consistently, mitochondrial chaperones, such as HSP10 and HSP60, but not ER chaperones are induced in defective cells. Mitochondrial dysfunction can result in oxidative stress, but no evidence for excessive ROS (reactive oxygen species) production was observed in cHccs(+/-) hearts. Instead, the antioxidative proteins SOD2 and PRDX3 are induced, suggesting that ROS detoxification prevents oxidative damage in HCCS deficient cardiomyocytes. Mitochondrial dysfunction and unrestricted UPR can induce cell death, and we detected the initiation of upstream events of both intrinsic as well as extrinsic apoptosis in cHccs(+/-) hearts. Cell death is not executed, however, suggesting the activation of antiapoptotic mechanisms. Whereas most apoptosis inhibitors are either unchanged or downregulated in HCCS deficient cardiomyocytes, Bcl-2 and ARC (apoptosis repressor with caspase recruitment domain) are induced. Given that ARC can inhibit both apoptotic pathways as well as necrosis and attenuates UPR, we generated cHccs(+/-) embryos on an Arc knockout background (cHccs(+/-),Arc(-/-)). Surprisingly, the absence of ARC does not induce cell death in embryonic or postnatal HCCS deficient cardiomyocytes and adult cHccs(+/-),Arc(-/-) mice exhibit normal cardiac morphology and function. Taken together, our data demonstrate an impressive plasticity of embryonic cardiomyocytes to respond to metabolic stress, the loss of which might be involved in the high susceptibility of postnatal cardiomyocytes to cell death.


RANK links thymic regulatory T cells to fetal loss and gestational diabetes in pregnancy.
Successful pregnancies rely on adaptations within the mother1, including marked changes within the immune system2. It has long been known that the thymus, the central lymphoid organ, changes markedly during pregnancy3. However, the molecular basis and importance of this process remain largely obscure. Here we show that the osteoclast differentiation receptor RANK4,5 couples female sex hormones to the rewiring of the thymus during pregnancy. Genetic deletion of Rank (also known as Tnfrsf11a) in thymic epithelial cells results in impaired thymic involution and blunted expansion of natural regulatory T (Treg) cells in pregnant female mice. Sex hormones, in particular progesterone, drive the development of thymic Treg cells through RANK in a manner that depends on AIRE+ medullary thymic epithelial cells. The depletion of Rank in the mouse thymic epithelium results in reduced accumulation of natural Treg cells in the placenta, and an increase in the number of miscarriages. Thymic deletion of Rank also results in impaired accumulation of Treg cells in visceral adipose tissue, and is associated with enlarged adipocyte size, tissue inflammation, enhanced maternal glucose intolerance, fetal macrosomia, and a long-lasting transgenerational alteration in glucose homeostasis, which are all key hallmarks of gestational diabetes. Transplantation of Treg cells rescued fetal loss, maternal glucose intolerance and fetal macrosomia. In human pregnancies, we found that gestational diabetes also correlates with a reduced number of Treg cells in the placenta. Our findings show that RANK promotes the hormone-mediated development of thymic Treg cells during pregnancy, and expand the functional role of maternal Treg cells to the development of gestational diabetes and the transgenerational metabolic rewiring of glucose homeostasis.


Joint-preserving surgical options for management of chondral injuries of the hip.
Management of injuries to the articular cartilage is complex and challenging; it becomes especially problematic in weight-bearing joints such as the hip. Several causes of articular cartilage damage have been described, including trauma, labral tears, and femoroacetabular impingement, among others. Because articular cartilage has little capacity for healing, nonsurgical management options are limited. Surgical options include total hip arthroplasty, microfracture, articular cartilage repair, autologous chondrocyte implantation, mosaicplasty, and osteochondral allograft transplantation. Advances in hip arthroscopy have broadened the spectrum of tools available for diagnosis and management of chondral damage. However, the literature is still not sufficiently robust to draw firm conclusions regarding best practices for chondral defects. Additional research is needed to expand our knowledge of and develop guidelines for management of chondral injuries of the hip.


Effects of Apelin on the fibrosis of retinal tissues and Müller cells in diabetes retinopathy through the JAK2/STAT3 signalling pathway.
Retinal fibrosis was a key characteristic of diabetes retinopathy (DR). Apelin was found to be a candidate for tissue fibrosis. Nevertheless, the role of Apelin in the Müller cells in DR remains unclear. This study identified the function and mechanism of Apelin in Müller cells and the fibrosis of retinal tissue. Western blot was carried out to detect the Apelin, GFAP, Collagen I, α-SMA, JAK2 and STAT3 protein levels. Masson staining was performed to display the histopathological changes in retinal tissue of diabetic mellitus (DM) rats. The immunofluorescence staining was conducted to evaluate the Apelin levels in the retinal tissue. The levels of GFAP, Collagen I and α-SMA in the retinal tissue of DM rats was visualised by the immunohistochemistry staining. The results showed that Apelin, GFAP, Collagen I andα-SMA expression was prominently elevated in the retinal tissue of DM rats and high glucose (HG)-exposed Müller cells. The results of Masson staining showed that the epiretinal fibrotic membrane was observed in DM rats. Apelin knockdown declined the GFAP, Collagen I andα-SMA levels. Besides, the protein levels of p-JAK2 and p-STAT3 were elevated in the HG-treated Müller cells, while Apelin knockdown declined them. FLLL32 treatment neutralised the role of Apelin. In conclusion, Apelin facilitated the fibrogenic activity of Müller cells through activating the JAK2/STAT3 signalling pathway, and thus inducing the retinal fibrosis in DR.


Towards a better monitoring of seed ageing under ex situ seed conservation.
Long-term conservation of 7.4 million ex situ seed accessions held in agricultural genebanks and botanic gardens worldwide is a challenging mission for human food security and ecosystem services. Recent advances in seed biology and genomics may have opened new opportunities for effective management of seed germplasm under long-term storage. Here, we review the current development of tools for assessing seed ageing and research advances in seed biology and genomics, with a focus on exploring their potential as better tools for monitoring of seed ageing. Seed ageing is found to be associated with the changes reflected in reactive oxygen species and mitochondria-triggered programmed cell deaths, expression of antioxidative genes and DNA and protein repair genes, chromosome telomere lengths, epigenetic regulation of related genes (microRNA and methylation) and altered organelle and nuclear genomes. Among these changes, the signals from mitochondrial and nuclear genomes may show the most promise for use in the development of tools to predict seed ageing. Non-destructive and non-invasive analyses of stored seeds through calorimetry or imaging techniques are also promising. It is clear that research into developing advanced tools for monitoring seed ageing to supplement traditional germination tests will be fruitful for effective conservation of ex situ seed germplasm.


TLR-3 polymorphism is an independent prognostic marker for stage II colorectal cancer.
Clinicopathologic stage is still the main parameter to evaluate the prognosis of newly diagnosed colorectal cancer (CRC) patients. Although molecular markers have been suggested for follow up of treated CRC patients, their complete clinical application is still under evaluation. To evaluate the association of immune-related genes with CRC prognosis and survival, a total of 19 single nucleotide polymorphisms (SNPs) were genotyped in 614 German patients within the Kiel cohort (POPGEN). A promoter variant (rs1800872) in the Interleukin-10 (IL-10) gene was associated with an increased lymph node metastasis involvement [odds ratio (OR) = 2.1, 95% confidence interval (CI) = 1.03-4.2, for carriers of the TT genotype]. More importantly, among 582 followed up patients the SNP rs3775291 in the toll-like receptor 3 (TLR-3) gene was associated with CRC specific survival (150 events). Patients carrying the TT genotype had a 93% increased risk of death compared with the CC carriers [hazard ratio (HR) = 1.93, 95% CI 1.14-3.28]. The observed effect of the TLR-3 variant was restricted to stage II patients (HR = 4.14, 95% CI 1.24-13.84) and to patients who did not receive adjuvant therapy (HR = 3.2, 95% CI 1.4-7.7). Our results may provide additional candidates for risk assessment in stage II CRC patients for treatment decision. Further validation of the presented findings is warranted.


Gene Duplication Accelerates the Pace of Protein Gain and Loss from Plant Organelles.
Organelle biogenesis and function is dependent on the concerted action of both organellar-encoded (if present) and nuclear-encoded proteins. Differences between homologous organelles across the Plant Kingdom arise, in part, as a result of differences in the cohort of nuclear-encoded proteins that are targeted to them. However, neither the rate at which differences in protein targeting accumulate nor the evolutionary consequences of these changes are known. Using phylogenomic approaches coupled to ancestral state estimation, we show that the plant organellar proteome has diversified in proportion with molecular sequence evolution such that the proteomes of plant chloroplasts and mitochondria lose or gain on average 3.6 proteins per million years. We further demonstrate that changes in organellar protein targeting are associated with an increase in the rate of molecular sequence evolution and that such changes predominantly occur in genes with regulatory rather than metabolic functions. Finally, we show that gain and loss of protein target signals occurs at a higher rate following gene duplication, revealing that gene and genome duplication are a key facilitator of plant organelle evolution.


Word pair classification during imagined speech using direct brain recordings.
People that cannot communicate due to neurological disorders would benefit from an internal speech decoder. Here, we showed the ability to classify individual words during imagined speech from electrocorticographic signals. In a word imagery task, we used high gamma (70-150 Hz) time features with a support vector machine model to classify individual words from a pair of words. To account for temporal irregularities during speech production, we introduced a non-linear time alignment into the SVM kernel. Classification accuracy reached 88% in a two-class classification framework (50% chance level), and average classification accuracy across fifteen word-pairs was significant across five subjects (mean = 58%; p < 0.05). We also compared classification accuracy between imagined speech, overt speech and listening. As predicted, higher classification accuracy was obtained in the listening and overt speech conditions (mean = 89% and 86%, respectively; p < 0.0001), where speech stimuli were directly presented. The results provide evidence for a neural representation for imagined words in the temporal lobe, frontal lobe and sensorimotor cortex, consistent with previous findings in speech perception and production. These data represent a proof of concept study for basic decoding of speech imagery, and delineate a number of key challenges to usage of speech imagery neural representations for clinical applications.


Self-rated health as a mediator between physical health conditions and depressive symptoms in older Chinese and Korean Americans.
In the present study, we examined self-rated health as a mediator between physical health conditions (chronic diseases and functional disability) and depressive symptoms in older Chinese and Korean Americans. Using harmonized data (N = 5,063) from the Population Study of Chinese Elderly (PINE) and the Study of Older Korean Americans (SOKA), we tested direct and indirect effect models. In both groups, chronic diseases and functional disability were closely associated with negative ratings of health and symptoms of depression. Analyses with the PROCESS macro showed that the effect of chronic diseases and functional disability on depressive symptoms was mediated by self-rated health in both groups; the indirect effect was greater in the Korean American sample than in the Chinese American sample. These findings contribute to the understanding of the psychological mechanisms that underlie the mind-body connection and highlight the potential importance of subjective health assessment as a useful tool for health promotion.


Effect of unsteady natural convection on the diameter of drawn polymer optical fiber.
This paper presents experimental results showing the effect various natural convection heating regimes have on the diameter of drawn polymer optical fiber. The airflow, adjacent to the polymer, can be either laminar, oscillatory, or chaotic, depending on the imposed thermal boundary conditions at the furnace and iris walls. When subject to oscillatory and chaotic natural convection, the drawn fiber varies in diameter 2.5 to 10 times more than that measured under laminar heating conditions. Particle image velocimetry shows that unsteady natural convection occurs with the interplay between two asymmetric counter-rotating convective cells. This represents a significant instability mechanism, one that has not been previously identified.


Physiological and molecular responses of the spiny dogfish shark (Squalus acanthias) to high environmental ammonia: scavenging for nitrogen.
In teleosts, a branchial metabolon links ammonia excretion to Na(+) uptake via Rh glycoproteins and other transporters. Ureotelic elasmobranchs are thought to have low branchial ammonia permeability, and little is known about Rh function in this ancient group. We cloned Rh cDNAs (Rhag, Rhbg and Rhp2) and evaluated gill ammonia handling in Squalus acanthias. Control ammonia excretion was <5% of urea-N excretion. Sharks exposed to high environmental ammonia (HEA; 1 mmol(-1) NH4HCO3) for 48 h exhibited active ammonia uptake against partial pressure and electrochemical gradients for 36 h before net excretion was re-established. Plasma total ammonia rose to seawater levels by 2 h, but dropped significantly below them by 24-48 h. Control ΔP(NH3) (the partial pressure gradient of NH3) across the gills became even more negative (outwardly directed) during HEA. Transepithelial potential increased by 30 mV, negating a parallel rise in the Nernst potential, such that the outwardly directed NH4(+) electrochemical gradient remained unchanged. Urea-N excretion was enhanced by 90% from 12 to 48 h, more than compensating for ammonia-N uptake. Expression of Rhp2 (gills, kidney) and Rhbg (kidney) did not change, but branchial Rhbg and erythrocytic Rhag declined during HEA. mRNA expression of branchial Na(+)/K(+)-ATPase (NKA) increased at 24 h and that of H(+)-ATPase decreased at 48 h, while expression of the potential metabolon components Na(+)/H(+) exchanger2 (NHE2) and carbonic anhydrase IV (CA-IV) remained unchanged. We propose that the gill of this nitrogen-limited predator is poised not only to minimize nitrogen loss by low efflux permeability to urea and ammonia but also to scavenge ammonia-N from the environment during HEA to enhance urea-N synthesis.


Secular trend in the maturation of permanent teeth in a sample of Turkish children over the past 30 years.
This study was performed to evaluate the influence of secular trends on dental maturation among Turkish children over the past 30 years. Orthopantomograms of 757 (385 boys, 372 girls) Turkish children born in the 1980s, 1990s, and 2000s were evaluated. Three groups were formed based on decade with five subgroups by age from 9 to 13 years old for each gender. The number of samples in each age group and gender were matched. The mandibular left seven permanent teeth were evaluated based on formation stage to determine the overall dental maturity score. The groups were compared based on decade and gender. The Bonferroni-corrected Mann-Whitney U test and Kruskal-Wallis tests were used for statistical evaluation. Among 11-, 12-, and 13-year-olds born in the 2000s, girls exhibited significantly more mature dentition than did boys (p<0.01, p<0.05, and p<0.05, respectively). Twelve-year-old girls born in the 1990s and 2000s exhibited significantly more mature dentition than did girls born in the 1980s (p<0.01). Girls generally exhibited more mature dentition than boys. No significant positive secular trends in dental maturity were observed from the 1980s through the 2000s. Dental maturation among Turkish children was not affected by a secular trend.


Narrative reconstruction therapy for prolonged grief disorder-rationale and case study.
Prolonged grief disorder (PGD) is a potentially disabling condition affecting approximately 10% of bereaved people. It has been suggested that the impaired integration of the loss memory, as expressed in recurrent memories of the loss and disorganization of memory, is involved in the development of PGD. Narrative reconstruction (NR), originally designed for the treatment of posttraumatic stress disorder (PTSD) in an integrative therapy module, and consisting of exposure to the loss memory, detailed written reconstruction of the loss memory narrative and an elaboration of the personal significance of that memory for the bereaved, has been shown to be effective in the treatment of intrusion symptoms. In light of findings that cognitive behavior therapy (CBT), including cognitive restructuring and exposure, is effective in the treatment of PGD, we suggest the implementation of a somewhat novel therapy module, NR, for the treatment of intrusive phenomena in bereaved patients. The rationale for the implementation of NR for PGD and a case study of the treatment of a woman suffering from PGD after the death of her father are presented. Therapy took place in a university outpatient training clinic. Evaluations conducted before and after treatment and at a 3-month follow-up demonstrated the effectiveness of NR in reducing symptoms of PGD and depression. The analysis of spontaneous narratives recorded before and after treatment showed an increased organization of the narratives. This case report demonstrates an adaptation of NR for the treatment of PGD. The results provide preliminary support for the effectiveness of NR for PGD. The significance of the study and its limitations are discussed.


Triple-Decker Motif for Red-Shifted Fluorescent Protein Mutants.
Among fluorescent proteins (FPs) used as genetically encoded fluorescent tags, the red-emitting FPs are of particular importance as suitable markers for deep tissue imaging. Using electronic structure calculations, we predict a new structural motif for achieving red-shifted absorption and emission in FPs from the GFP family. By introducing four point mutations, we arrive to the structure with the conventional anionic GFP chromophore sandwiched between two tyrosine residues. Contrary to the existing red FPs in which the red shift is due to extended conjugation of the chromophore, in the triple-decker motif, the chromophore is unmodified and the red shift is due to π-stacking interactions. The absorption/emission energies of the triple-decker FP are 2.25/2.16 eV, respectively, which amounts to shifts of ∼40 (absorption) and ∼25 nm (emission) relative to the parent species, the I form of wtGFP. Using a different structural motif based on a smaller chromophore may help to improve optical output of red FPs by reducing losses due to radiationless relaxation and photobleaching.


To bet or not to bet? The error negativity or error-related negativity associated with risk-taking choices.
The functional significance of error-related negativity (Ne/ERN), which occurs at approximately the same time as erroneous responses, has been investigated extensively using reaction time (RT) tasks. The error detection theory assumes that the Ne/ERN reflects the mismatch detected by comparing representations of the intended and the actually performed actions. The conflict monitoring theory asserts that the Ne/ERN reflects the detection of response conflict between intended and actually performed actions during response selection. In this study, we employed a gambling task in which participants were required to choose whether they would take part in betting in each trial and they were presented with gain or loss feedback in both the "to bet" and the "not to bet" trials. The response-locked ERP magnitudes were more negative for "to bet" than for "not to bet" choices for both large and small stakes and were more negative for choices involving large rather than small stakes. Dipole source analysis localized the ERP responses to the anterior cingulate cortex (ACC). These findings suggest that the ACC signals the riskiness of choices and may function as an early warning system that alerts the brain to prepare for the potential negative consequence associated with a risky action.


Prevalence of Microbiological and Chemical Contaminants in Private Drinking Water Wells in Maryland, USA.
Although many U.S. homes rely on private wells, few studies have investigated the quality of these water sources. This cross-sectional study evaluated private well water quality in Maryland, and explored possible environmental sources that could impact water quality. Well water samples (n = 118) were collected in four Maryland counties and were analyzed for microbiological and chemical contaminants. Data from the U.S. Census of Agriculture were used to evaluate associations between the presence of animal feeding operations and well water quality at the zip code level using logistic regression. Overall, 43.2% of tested wells did not meet at least one federal health-based drinking water standard. Total coliforms, fecal coliforms, enterococci, and Escherichia coli were detected in 25.4%, 15.3%, 5.1%, and 3.4% of tested wells, respectively. Approximately 26%, 3.4%, and <1% of wells did not meet standards for pH, nitrate-N, and total dissolved solids, respectively. There were no statistically significant associations between the presence of cattle, dairy, broiler, turkey, or aquaculture operations and the detection of fecal indicator bacteria in tested wells. In conclusion, nearly half of tested wells did not meet federal health-based drinking water standards, and additional research is needed to evaluate factors that impact well water quality. However, homeowner education on well water testing and well maintenance could be important for public health.


Nonuniform spatial patterns of respiratory activity within biofilms during disinfection.
Fluorescent stains in conjunction with cryoembedding and image analysis were applied to demonstrate spatial gradients in respiratory activity within bacterial biofilms during disinfection with monochloramine. Biofilms of Klebsiella pneumoniae and Pseudomonas aeruginosa grown together on stainless steel surfaces in continuous-flow annular reactors were treated with 2 mg of monochloramine per liter (influent concentration) for 2 h. Relatively little biofilm removal occurred as evidenced by total cell direct counts. Plate counts (of both species summed) indicated an average 1.3-log decrease after exposure to 2 mg of monochloramine per liter. The fluorogenic redox indicator 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and the DNA stain 4',6-diamidino-2-phenylindole (DAPI) were used to differentiate respiring and nonrespiring cells in biofilms. Epifluorescence micrographs of frozen biofilm cross sections clearly revealed gradients of respiratory activity within biofilms in response to monochloramine treatment. These gradients in specific respiratory activity were quantified by calculating the ratio of CTC and DAPI intensities measured by image analysis. Cells near the biofilm-bulk fluid interface lost respiratory activity first. After 2 h of biocide treatment, greater respiratory activity persisted deep in the biofilm than near the biofilm-bulk fluid interface.


Two functional motifs define the interaction, internalization and toxicity of the cell-penetrating antifungal peptide PAF26 on fungal cells.
The synthetic, cell penetrating hexapeptide PAF26 (RKKWFW) is antifungal at low micromolar concentrations and has been proposed as a model for cationic, cell-penetrating antifungal peptides. Its short amino acid sequence facilitates the analysis of its structure-activity relationships using the fungal models Neurospora crassa and Saccharomyces cerevisiae, and human and plant pathogens Aspergillus fumigatus and Penicillium digitatum, respectively. Previously, PAF26 at low fungicidal concentrations was shown to be endocytically internalized, accumulated in vacuoles and then actively transported into the cytoplasm where it exerts its antifungal activity. In the present study, two PAF26 derivatives, PAF95 (AAAWFW) and PAF96 (RKKAAA), were designed to characterize the roles of the N-terminal cationic and the C-terminal hydrophobic motifs in PAF26's mode-of-action. PAF95 and PAF96 exhibited substantially reduced antifungal activity against all the fungi analyzed. PAF96 localized to fungal cell envelopes and was not internalized by the fungi. In contrast, PAF95 was taken up into vacuoles of N. crassa, wherein it accumulated and was trapped without toxic effects. Also, the PAF26 resistant Δarg1 strain of S. cerevisiae exhibited increased PAF26 accumulation in vacuoles. Live-cell imaging of GFP-labelled nuclei in A. fumigatus showed that transport of PAF26 from the vacuole to the cytoplasm was followed by nuclear breakdown and dissolution. This work demonstrates that the amphipathic PAF26 possesses two distinct motifs that allow three stages in its antifungal action to be defined: (i) its interaction with the cell envelope; (ii) its internalization and transport to vacuoles mediated by the aromatic hydrophobic domain; and (iii) its transport from vacuoles to the cytoplasm. Significantly, cationic residues in PAF26 are important not only for the electrostatic attraction and interaction with the fungal cell but also for transport from the vacuole to the cytoplasm, which coincides with cell death. Peptide containment within vacuoles preserves fungal cells from peptide toxicity.