Knowledge about and Attitude towards Epilepsy among Menit Community, Southwest Ethiopia.
Nearly 80% of the people with epilepsy are found in developing countries, where epilepsy remains a major public health problem, not only because of its health implications but also for its social, cultural, psychological and economic effects. The objective of this study was to assess knowledge and attitude of Menit community in Benchi-Maji Zone as regards epilepsy. The study was conducted in Menit Goldia Province, one of the provinces of Bench-Maji Zone. Community based descriptive cross-sectional study was employed. Households were selected by using systematic sampling technique. Data was collected through pretested interviewer administered questionnaire. Data was cleaned, coded and entered in to EPI data version 3.1. The data was cleaned and transported into SPSS for analysis. Among the study participants, 808(97.1%) ever heard about epilepsy, 85.3% reported that epilepsy is a mental disease where as 40.6%, 49% and 49.4% respectively believed that it is hereditary, contagious, and God's curse. Among the respondents, 252(30%) think that epileptics should be isolated from the community, 387(46.1%) and 336(40%) do not want to shake hands with epileptics and keep their children away from epileptic patients respectively. In this study, 85.6% and86.8% of the respondents were not knowledgeable about and had negative attitude towards epilepsy respectively. Ethnicity and educational status were significantly associated with knowledge and attitude. The FGD participants noted that the disease is God's curse, and they lacked knowledge about the cause of epilepsy. The study participants' level of knowledge about and attitude towards epilepsy was not satisfactory. Thus, it needs attention from concerned bodies.


How parrots see their colours: novelty in the visual pigments of Platycercus elegans.
Intraspecific differences in retinal physiology have been demonstrated in several vertebrate taxa and are often subject to adaptive evolution. Nonetheless, such differences are currently unknown in birds, despite variations in habitat, behaviour and visual stimuli that might influence spectral sensitivity. The parrot Platycercus elegans is a species complex with extreme plumage colour differences between (and sometimes within) subspecies, making it an ideal candidate for intraspecific differences in spectral sensitivity. Here, the visual pigments of P. elegans were fully characterised through molecular sequencing of five visual opsin genes and measurement of their absorbance spectra using microspectrophotometry. Three of the genes, LWS, SW1 and SWS2, encode for proteins similar to those found in other birds; however, both the RH1 and RH2 pigments had polypeptides with carboxyl termini of different lengths and unusual properties that are unknown previously for any vertebrate visual pigment. Specifically, multiple RH2 transcripts and protein variants (short, medium and long) were identified for the first time that are generated by alternative splicing of downstream coding and non-coding exons. Our work provides the first complete characterisation of the visual pigments of a parrot, perhaps the most colourful order of birds, and moreover suggests more variability in avian eyes than hitherto considered.


Obstructor A organizes matrix assembly at the apical cell surface to promote enzymatic cuticle maturation in Drosophila.
Assembly and maturation of the apical extracellular matrix (aECM) is crucial for protecting organisms, but underlying molecular mechanisms remain poorly understood. Epidermal cells secrete proteins and enzymes that assemble at the apical cell surface to provide epithelial integrity and stability during developmental growth and upon tissue damage. We analyzed molecular mechanisms of aECM assembly and identified the conserved chitin-binding protein Obst-A (Obstructor A) as an essential regulator. We show in Drosophila that Obst-A is required to coordinate protein and chitin matrix packaging at the apical cell surface during development. Secreted by epidermal cells, the Obst-A protein is specifically enriched in the apical assembly zone where matrix components are packaged into their highly ordered architecture. In obst-A null mutant larvae, the assembly zone is strongly diminished, resulting in severe disturbance of matrix scaffold organization and impaired aECM integrity. Furthermore, enzymes that support aECM stability are mislocalized. As a biological consequence, cuticle architecture, integrity, and function are disturbed in obst-A mutants, finally resulting in immediate lethality upon wounding. Our studies identify a new core organizing center, the assembly zone that controls aECM assembly at the apical cell surface. We propose a genetically conserved molecular mechanism by which Obst-A forms a matrix scaffold to coordinate trafficking and localization of proteins and enzymes in the newly deposited aECM. This mechanism is essential for maturation and stabilization of the aECM in a growing and remodeling epithelial tissue as an outermost barrier.


Multiwell Assay for the Analysis of Sugar Gut Permeability Markers: Discrimination of Sugar Alcohols with a Fluorescent Probe Array Based on Boronic Acid Appended Viologens.
With the aim of discerning between different sugar and sugar alcohols of biomedical relevance, such as gut permeability, arrays of 2-component probes were assembled with up to six boronic acid-appended viologens (BBVs): 4,4'-o-BBV, 3,3'-o-BBV, 3,4'-o-BBV, 4,4'-o,m-BBV, 4,7'-o-PBBV, and pBoB, each coupled to the fluorophore 8-hydroxypyrene, 1,3,6-trisulfonic acid trisodium salt (HPTS). These probes were screened for their ability to discriminate between lactulose, l-rhamnose, 3-O-methyl-d-glucose, and xylose. Binding studies of sugar alcohols mannitol, sorbitol, erythritol, adonitol, arabitol, galactitol, and xylitol revealed that diols containing threo-1,2-diol units have higher affinity for BBVs relative diols containing erythro-1,2 units. Those containing both threo-1,2- and 1,3-syn diol motifs showed high affinity for boronic acid binding. Fluorescence from the arrays were examined by principle component analysis (PCA) and linear discriminant analysis (LDA). Arrays with only three BBVs sufficed to discriminate between sugars (e.g., lactulose) and sugar alcohols (e.g., mannitol), establishing a differential probe. Compared with 4,4'-o-BBV, 2-fold reductions in lower limits of detection (LOD) and quantification (LOQ) were achieved for lactulose with 4,7-o-PBBV (LOD 41 μM, LOQ 72 μM). Using a combination of 4,4'-o-BBV, 4,7-o-PBBV, and pBoB, LDA statistically segregated lactulose/mannitol (L/M) ratios from 0.1 to 0.5, consistent with values encountered in small intestinal permeability tests. Another triad containing 3,3'-o-BBV, 4,4'-o-BBV, and 4,7-o-PBBV also discerned similar L/M ratios. This proof-of-concept demonstrates the potential for BBV arrays as an attractive alternate to HPLC to analyze mixtures of sugars and sugar alcohols in biomedical applications and sheds light on structural motifs that make this possible.


Breast Density Notification: An Australian Perspective.
Breast density, also known as mammographic density, refers to white and bright regions on a mammogram. Breast density can only be assessed by mammogram and is not related to how breasts look or feel. Therefore, women will only know their breast density if they are notified by the radiologist when they have a mammogram. Breast density affects a woman's breast cancer risk and the sensitivity of a screening mammogram to detect cancer. Currently, the position of BreastScreen Australia and the Royal Australian and New Zealand College of Radiologists is to not notify women if they have dense breasts. However, patient advocacy organisations are lobbying for policy change. Whether or not to notify women of their breast density is a complex issue and can be framed within the context of both public health ethics and clinical ethics. Central ethical themes associated with breast density notification are equitable care, patient autonomy in decision-making, trust in health professionals, duty of care by the physician, and uncertainties around evidence relating to measurement and clinical management pathways for women with dense breasts. Legal guidance on this issue must be gained from broad legal principles found in the law of negligence and the test of materiality. We conclude a rigid legal framework for breast density notification in Australia would not be appropriate. Instead, a policy framework should be developed through engagement with all stakeholders to understand and take account of multiple perspectives and the values at stake.


Lack of Utility of Anti-tTG IgG to Diagnose Celiac Disease When Anti-tTG IgA Is Negative.
Guidelines for diagnosing celiac disease (CD) recommend initial testing with a highly sensitive serologic test for anti-tissue transglutaminase immunoglobulin A antibodies (tTG IgA). When the probability of CD is high, IgA deficiency should be considered. The 2 approaches to address this include measuring "both tTG IgA and tTG IgG" or measuring "total IgA." We aim to assess the utility of an isolated positive tTG IgG result in diagnosing CD. We conducted a retrospective review of patients undergoing serologic testing for CD from January 1997 to June 2014. Patients with positive tTG IgG and negative tTG IgA were included. Moreover, all patients who had any other positive CD-specific serologic findings were excluded. Demographics, clinical presentation, tests, and biopsy results were recorded. The indication for checking celiac serology was gastrointestinal symptoms in 172 of 233 patients, iron deficiency anemia in 12, and high-risk screening in 48. Small bowel biopsy was performed in 178 patients (77%); 160 had normal results and 18 had histologic changes suggestive of enteropathy. Nine patients had increased intraepithelial lymphocytes, and 9 had partial villous atrophy. Only 6 cases of CD were, however, confirmed. The utility of isolated tTG IgG in diagnosis of CD was low at 3% (6/178). In this cohort of patients, the utility of isolated tTG IgG in diagnosing CD was low at 3%.


Development of an in vivo rabbit ulnar loading model.
Ulnar and tibial cyclic compression in rats and mice have become the preferred animal models for investigating the effects of mechanical loading on bone modeling/remodeling. Unlike rodents, rabbits provide a larger bone volume and normally exhibit intracortical Haversian remodeling, which may be advantageous for investigating mechanobiology and pharmaceutical interventions in cortical bone. Therefore, the objective of this study was to develop and validate an in vivo rabbit ulnar loading model. Ulnar tissue strains during loading of intact forelimbs were characterized and calibrated to applied loads using strain gauge measurements and specimen-specific finite element models. Periosteal bone formation in response to varying strain levels was measured by dynamic histomorphometry at the location of maximum strain in the ulnar diaphysis. Ulnae loaded at 3000 microstrain did not exhibit periosteal bone formation greater than the contralateral controls. Ulnae loaded at 3500, 4000, and 4500 microstrain exhibited a dose-dependent increase in periosteal mineralizing surface (MS/BS) compared with contralateral controls during the second week of loading. Ulnae loaded at 4500 microstrain exhibited the most robust response with significantly increased MS/BS at multiple time points extending at least 2weeks after loading was ceased. Ulnae loaded at 5250 microstrain exhibited significant woven bone formation. Rabbits required greater strain levels to produce lamellar and woven bone on periosteal surfaces compared with rats and mice, perhaps due to lower basal levels of MS/BS. In summary, bone adaptation during rabbit ulnar loading was tightly controlled and may provide a translatable model for human bone biology in preclinical investigations of metabolic bone disease and pharmacological treatments.


Clinical significance of anti-At(a).
Ata is a high-frequency red blood cell (RBC) antigen. Anti-At(a) has been reported in rare At(a-) black subjects. We report two cases of anti-At(a). A clinically significant anti-At(a) was found in a 26-year-old black woman with systemic lupus erythematosus. The patient had a transfusion reaction with chills and nausea during a RBC survival study, and 95% of the radiolabeled At(a+) RBCs were destroyed within 3 h. A concurrently performed monocyte monolayer assay was strongly reactive. Anti-At(a) thus can cause rapid hemolysis of transfused RBCs, but At(a-) donor units are extremely scarce in rare donor registries. A second patient at our hospital had anti-At(a) which did not affect her newborn. She also had autoimmune disease, insulin-dependent diabetes mellitus.


Discovery of an integrative network of microRNAs and transcriptomics changes for acute kidney injury.
The contribution of miRNA to the pathogenesis of acute kidney injury (AKI) is not well understood. Here we evaluated an integrative network of miRNAs and mRNA data to discover a possible master regulator of AKI. Microarray analyses of the kidneys of mice treated with cisplatin were used to extract putative miRNAs that cause renal injury. Of them, miR-122 was mostly downregulated by cisplatin, whereas miR-34a was upregulated. A network integrating dysregulated miRNAs and altered mRNA expression along with target prediction enabled us to identify Foxo3 as a core protein to activate p53. The miR-122 inhibited Foxo3 translation as assessed using an miR mimic, an inhibitor, and a Foxo3 3'-UTR reporter. In a mouse model, Foxo3 levels paralleled the degree of tubular injury. The role of decreased miR-122 in inducing Foxo3 during AKI was strengthened by the ability of the miR-122 mimic or inhibitor to replicate results. Increase in miR-34a also promoted the acetylation of Foxo3 by repressing Sirt1. Consistently, cisplatin facilitated the binding of Foxo3 and p53 for activation, which depended not only on decreased miR-122 but also on increased miR-34a. Other nephrotoxicants had similar effects. Among targets of p53, Phlda3 was robustly induced by cisplatin, causing tubular injury. Consistently, treatment with miR mimics and/or inhibitors, or with Foxo3 and Phlda3 siRNAs, modulated apoptosis. Thus, our results uncovered an miR integrative network regulating toxicant-induced AKI and identified Foxo3 as a bridge molecule to the p53 pathway.


The interrelationship of the concentration of hydrogen ions, bicarbonate ions, carbon dioxide and calcium ions in the regulation of renal gluconeogenesis in the rat.
1. The interrelationship of acidosis and Ca(2+) on the stimulation of gluconeogenesis by rat kidney-cortex slices was studied. 2. Ca(2+) stimulated gluconeogenesis from glutamine, glutamate, 2-oxoglutarate, succinate, malate, pyruvate, lactate and fructose, but not from galactose. 3. The [Ca(2+)] needed for optimum gluconeogenesis was about 2mm, but at this concentration, acidosis, produced in vitro by a decrease of [HCO(3) (-)] in the medium at constant pCO(2) or by an increase in pCO(2) at constant [HCO(3) (-)], did not stimulate gluconeogenesis. 4. In the absence of Ca(2+), acidosis (low [HCO(3) (-)]) stimulated gluconeogenesis from glutamine, glutamate, 2-oxoglutarate, succinate, malate, pyruvate and lactate but not from fructose or galactose. With succinate as substrate, the stimulatory effect of acidosis (low [HCO(3) (-)]) disappeared at Ca(2+) concentrations above 1.0mm. 5. The [HCO(3) (-)] was the most important determinant of the acidosis effect since a decrease of pH caused by an increase in pCO(2) did not uniformly stimulate gluconeogenesis, whereas a decrease in [HCO(3) (-)] without a change in pH consistently stimulated glucose formation in a way similar to the stimulation produced by acidosis (low [HCO(3) (-)]) in the absence of Ca(2+). 6. Acidosis in vitro inhibited the rate of decrease of activity of phosphoenolpyruvate carboxylase in slices, and Ca(2+) caused an increase in the activity of fructose 1-phosphate aldolase. 7. Respiratory acidosis in vitro caused an increase in the activity of phosphoenolpyruvate carboxylase in kidney cortex and an increase in gluconeogenesis from glutamine. 8. Possible points of interaction between Ca(2+), H(+) and HCO(3) (-) with the gluconeogenic sequence are discussed.


CpG DNA increases primary malignant B cell expression of costimulatory molecules and target antigens.
Multiple factors, including expression of costimulatory molecules, antigen-presenting molecules, and target antigens, likely impact the efficacy of antibody therapy and other approaches to the immunotherapy of B cell malignancy. Unmethylated CpG-dinucleotides in select base contexts ("CpG motifs") that resemble sequences found in bacterial DNA are potent immunostimulatory agents capable of inducing a complex immune response, including a strong B cell stimulus. We examined the effect of a potent human CpG oligonucleotide (CpG ODN 2006) on different types of primary human malignant B cells and reactive follicular hyperplasia. CpG oligodeoxynucleotide (CpG ODN), but not control (non-CpG ODN), increased the expression of costimulatory molecules (CD40, CD80, CD86, CD54) on malignant B cells without altering the phenotype of B cells obtained from reactive follicular hyperplasia. CpG ODN also enhanced expression of class I and class II MHC in most samples. CD20 expression was increased in response to CpG ODN, most notably in B-CLL and marginal zone lymphoma. An inverse correlation was found between baseline expression of CD20 and CD40 and their expression after exposure to CpG ODN, thus the most significant increase in expression of these molecules was found in those samples that had the lowest baseline levels. In conclusion, CpG ODN can lead to increasing expression of molecules involved in costimulation, antigen presentation, and as targets for antibody-based therapy and deserve further evaluation as potential immunotherapeutic agents for B cell malignancy.


Effects of foveal priming and extrafoveal preview on object identification.
The results of three different experiments suggested that the relation between an object in the fovea on fixation n and an object subsequently brought into the fovea on fixation n + 1 affects the time to identify the second object. In Experiment 1 we extended previous work by demonstrating that a previously seen related priming object speeded the time to name a target object even when a saccade intervened between the two objects. In Experiment 2 we replicated this result and further showed that the benefit on naming time was due to facilitation from the related object rather than inhibition from the unrelated object. In addition, naming of the target object was much slower in both experiments when there was not a peripheral preview of the target object on fixation n. However, because the effect of the foveal priming object was greater when the target was not present than when it was present, priming did not appear to make extraction of the extrafoveal information more efficient. In Experiment 3, fixation times were recorded while subjects looked at four objects in order to identify them. Fixation time on an object was shorter when a related object was fixated immediately before it, even though the four objects did not form a scene. The size of the facilitation was roughly comparable to that in several analogous experiments where scenes were used. The results suggest that the effects of a predictive scene context on object identification may be explainable in terms of an object-to-object or "intralevel" priming mechanism.


Real-time optical holographic tracking of multiple objects.
A coherent optical correlation technique for real-time simultaneous tracking of several different objects making independent movements is described, and experimental results are presented. An evaluation of this system compared with digital computing systems is made. The real-time processing capability is obtained through the use of a liquid crystal television spatial light modulator and a dichromated gelatin multifocus hololens. A coded reference beam is utilized in the separation of the output correlation plane associated with each input target so that independent tracking can be achieved.


[Effect of changes in blood glucose concentration on the standing potential of the rabbit eye].
The paper deals with the study of the influence of changes of glucose blood concentration in rabbits on the standing potential of the eye, which reflects the functioning of the retinal pigment epithelium and photoreceptors. Recordings of the electrooculogram (EOG), determination of glucose blood level and ophthalmoscopic and histological examinations of the eye were performed in 48 rabbits. Short-term increase of glucose concentration was accompanied by the increase in the standing potential and Arden EOG ratio, with hypoglycemia evoking the opposite effect. A long-term (4 month) hyperglycemia was accompanied by the reversible decrease of both parameters studied. No ophthalmoscopic and histological changes were observed.


Sleep and circadian rhythm disturbance in kidney stone disease: a narrative review.
The circadian rhythm generated by circadian clock genes functions as an internal timing system. Since the circadian rhythm controls abundant physiological processes, the circadian rhythm evolved in organisms is salient for adaptation to environmental change. A disturbed circadian rhythm is a trigger for numerous pathological events. Recently, accumulated data have indicated that kidney stone disease (KSD) is related to circadian rhythm disturbance. However, the mechanism between them has not been fully elucidated. In this narrative review, we summarized existing evidence to illustrate the possible association between circadian rhythm disturbance and KSD based on the epidemiological studies and risk factors that are linked to circadian rhythm disturbance and discuss some chronotherapies for KSD. In summary, KSD is associated with systemic disorders. Metabolic syndrome, inflammatory bowel disease, and microbiome dysbiosis are the major risk factors supported by sufficient data to cause KSD in patients with circadian rhythm disturbance, while others including hypertension, vitamin D deficiency, parathyroid gland dysfunction, and renal tubular damage/dysfunction need further investigation. Then, some chronotherapies for KSD were confirmed to be effective, but the molecular mechanism is still unclear.


Constitutively active K-cyclin/cdk6 kinase in Kaposi sarcoma-associated herpesvirus-infected cells.
Kaposi sarcoma-associated human herpesvirus (KSHV) encodes K-cyclin, a homologue of D-type cellular cyclins, which binds cyclin-dependent kinases to phosphorylate various substrates. K-cyclin/cdk phosphorylates a subset of substrates normally targeted by cyclins D, E, and A. We used cells naturally infected with KSHV to further characterize the biochemical features of K-cyclin. We used immunoprecipitation with K-cyclin antibodies to examine the association of K-cyclin with cdk2, cdk6, p21Cip1, and p27Kip1 proteins in BC3 cells. We separated populations of BC3 cells enriched in cells in G1, S, or G2/M phases by elutriation and measured K-cyclin protein and the kinase activity of K-cyclin/cdk6 complexes. The half-life of K-cyclin and cyclin D2 proteins was determined by blocking protein synthesis with cycloheximide and measuring proteins in cell lysates by western blot analysis. We fused the entire K-cyclin sequence to the carboxyl-terminal sequence of cellular cyclin D that contains the PEST degradation sequence to produce K-cyclin/D2 and transfected K-cyclin/D2 into K-cyclin-negative cells to investigate the effect of the PEST sequence on K-cyclin's stability. Viral K-cyclin interacted with cyclin-dependent kinases cdk2, cdk4, and cdk6 and with the cyclin/cdk inhibitory proteins p21Cip1 and p27Kip1 in BC3 cell lysates. Unlike D-type cyclins, whose expression is cell cycle dependent, the level of K-cyclin was stable throughout the cell cycle, and the kinase associated with the K-cyclin/cdk6 complex was constitutively active. The half-life of K-cyclin (6.9 hours) was much longer than that of cellular cyclin D2 (0.6 hour) and that of K-cyclin/D2 (0.5 hour), probably because K-cyclin lacks the PEST degradation sequence present in D-type cyclins. The constitutive activation of K-cyclin/cdk complexes in KSHV-infected cells appears to result from the extended half-life of K-cyclin and may explain its role in Kaposi sarcoma.


Comparison of effects of propofol and ketofol (Ketamine-Propofol mixture) on emergence agitation in children undergoing tonsillectomy.
The aim of this study was to compare the effect of propofol and ketofol (ketamine-propofol mixture) on EA in children undergoing tonsillectomy. In this randomized clinical trial, 87 ASA class I and II patients, aged 3-12 years, who underwent tonsillectomy, were divided into two groups to receive either propofol 100 µg/kg/min (group p, n=44) or ketofol : ketamine 25 µg/kg/min + propofol 75 µg/kg/min (group k, n= 43). Incidence and severity of EA was evaluated using the Pediatric Anesthesia Emergence Delirium (PAED) scales on arrival at the recovery room, and 10 and 30 min after that time. There was no statistically significant difference in demographic data between the two groups. In the ketofol group, the need for agitation treatment and also mean recovery duration were lower than in the propofol group (30 and 41%, and 29.9 and 32.7 min), without statistically significant difference (P value=0.143 and P value=0.187). Laryngospasm or bronchospasm occurred in 2 patients in each group and bleeding was observed in only one individual in the ketofol group. Infusion of ketofol in children undergoing tonsillectomy provides shorter recovery time and lower incidence of EA despite the non significant difference with propofol.


Spiritual skepticism? Heterogeneous science skepticism in the Netherlands.
Recent work points to the heterogeneous nature of science skepticism. However, most research on science skepticism has been conducted in the United States. The current work addresses the generalizability of the knowledge acquired so far by investigating individuals from a Western European country (The Netherlands). Results indicate that various previously reported findings hold up: Mirroring North American patterns, climate change skepticism is associated with political conservatism (but only modestly), and scientific literacy does not contribute to skepticism, except about genetic modification (Study 1 only) and vaccine skepticism (Study 2 only). Results also reveal a crucial difference: Religiosity does not consistently contribute to science skepticism, except about evolution. Instead, spirituality is found to most consistently predict vaccine skepticism and low general faith in science-which in turn predicts willingness to support science. Concerns about societal impact play an additional role. These findings speak to the generalizability of previous findings, improving our understanding of science skepticism.


Reductive nanocomplex encapsulation of cRGD-siRNA conjugates for enhanced targeting to cancer cells.
In this study, through covalent conjugation and lipid material entrapment, a combined modification strategy was established for effective delivery of small interfering RNA (siRNA). Single strands of siRNA targeting to BRAFV600E gene (siMB3) conjugated with cRGD peptide at 3'-terminus or 5'-terminus via cleavable disulfide bond was synthesized and then annealed with corresponding strands to obtain single and bis-cRGD-siRNA conjugates. A cationic lipid material (CLD) developed by our laboratory was mixed with the conjugates to generate nanocomplexes; their uniformity and electrical property were revealed by particle size and zeta potential measurement. Compared with CLD/siBraf, CLD/cRGD-siBraf achieved higher cell uptake and more excellent tumor-targeting ability, especially 21 (sense-5'/antisense-3″-cRGD-congjugate) nanocomplex. Moreover, they can regulate multiple pathways to varying degree and reduce acidification of endosome. Compared with the gene silencing of different conjugates, single or bis-cRGD-conjugated siRNA showed little differences except 22 (5/5) which cRGD was conjugated at 5'-terminus of antisense strand and sense strand. However bis-cRGD conjugate 21 nanocomplex exhibited better specific target gene silencing at multiple time points. Furthermore, the serum stabilities of the bis-cRGD conjugates were higher than those of the single-cRGD conjugates. In conclusion, all these data suggested that CLD/bis-conjugates, especially CLD/21, can be an effective system for delivery of siRNA to target BRAFV600E gene for therapy of melanoma.


[Randomized placebo-controlled trial of orlistat for weight loss and prevention of weight regain in obese patients].
We undertook a randomised controlled trial to assess the efficacy and tolerance of orlistat, a gastrointestinal lipase inhibitor, in promoting weight loss and preventing weight regain in obese patients over a 2-year period. 743 patients (body-mass index 28-47 kg/m2), recruited at 15 European centres, entered a 4-week, single-blind, placebo lead-in period on a slightly hypocaloric diet (600 kcal/day deficit). 688 patients who completed the lead-in were assigned double-blind treatment with orlistat 120 mg (three times a day) or placebo for 1 year in conjunction with the hypocaloric diet. In a second 52-week double-blind period patients were reassigned orlistat or placebo with a weight maintenance (eucaloric) diet. From the start of lead-in to the end of year 1, the orlistat group lost, on average, more bodyweight than the placebo group (10.2% [10.3 kg] vs 6.1% [6.1 kg]; LSM difference 3.9 kg [p < 0.001] from randomisation to the end of year 1). During year 2, patients who continued with orlistat regained, on average, half as much weight as those patients switched to placebo (p < 0.001). Patients switched from placebo to orlistat lost an additional 0.9 kg during year 2, compared with a mean regain of 2.5 kg in patients who continued on placebo (p < 0.001). Total cholesterol, low-density lipoprotein (LDL) cholesterol, LDL/high-density lipoprotein ratio, and concentrations of glucose and insulin decreased more in the orlistat group than in the placebo group. Gastrointestinal adverse events were more common in the orlistat group. Other adverse symptoms occurred at a similar frequency during both treatments. Orlistat taken with an appropriate diet promotes clinically significant weight loss and reduces weight regain in obese patients over a 2-year period. The use of orlistat beyond 2 years needs careful monitoring with respect to efficacy and adverse events.


[Tackling hemodynamic analysis of the carotid artery using open-source software and computational fluid dynamics].
The aim of this study was to evaluate the impact of wall share stress (WSS) in the carotid artery using a computed fluid dynamics analysis system and adopting open-source software. The dependence of element number (computation time and analytical accuracy) were considered with simple vessel models. We evaluated WSS and flow velocity using a carotid artery model that was based on the outcome of simple vessel models. When the number of elements was 10(5) or more, the flow velocity error of the outlet decreased to 0.5% or below when using simple vessel models. The carotid bifurcation model showed a whirlpool and a decrease in flow velocity in the carotid bulb part. An analysis system was built using open source software. The results from the carotid bifurcation model suggested that hemodynamics contributes to the development of carotid stenosis.


Trichosporon infection in a cat.
A protruding nasal mass in a domestic shorthair cat with nasal discharge and recurrent fever was determined to be caused by infection with the fungus Trichosporon pullulans, as determined by cytologic, histologic, and fungal culture methods. Initially, the cat was treated orally with ketoconazole and the mass decreased in size. When signs of hepatotoxicosis developed, treatment was reduced to an alternate-day basis. However, the nasal mass increased in size with this regimen, and therapy with amphotericin B and 5-flucytosine was begun. It also was unsuccessful.


[Linburg-Comstock syndrome. Epidemiologic and anatomic study, clinical applications].
The Linburg-Comstock (LC) syndrome is distinguished by the inability to actively flex the interphalangeal (IP) joint of the thumb without simultaneously flexing the distal IP joint of the index finger. Any resistance to this 'parasitic' reaction causes pain on the palmar side of the wrist or in the distal part of the forearm; this is due to an anomalous tendinous connection between the flexor pollicus longus (FPL) and the flexor digitorum profundus (FDP). An epidemiological study was carried out on 264 individuals (a total of 528 hands were examined), and the LC syndrome was found in 98 subjects (37%); women were more frequently affected than men, and bilaterally rather than unilaterally. In addition, we dissected 26 fresh cadaver upper limbs, and in seven cases found an anomalous connection between FPL and FDP. We also examined the case of a young violinist with bilateral LC syndrome, who complained of pain in the distal part of the left forearm after prolonged musical exercises. Surgical investigation determined a complete fusion between FPL and FDP of the index with a common tendon. Treatment consisted of splitting this common tendon to form two separate tendons, thereby permitting a certain degree of independence between the thumb and index finger, and which considerably improved the violinist's musical performance. A review of the literature showed that there was a large quantity of anatomical descriptions available on these types of connection. Certain publications also provide an extremely precise report on the anthropological significance of these anomalies.


Glioma growth inhibition by neurostatin and O-But GD1b.
In spite of their low incidence, central nervous system tumors have elevated morbidity and mortality, being responsible for 2.3% of total cancer deaths. The ganglioside O-acetylated GD1b (O-Ac GD1b; neurostatin), present in the mammalian brain, and the semi-synthetic O-butyrylated GD1b (O-But GD1b) are potent glioma proliferation inhibitors, appearing as possible candidates for the treatment of nervous system tumors. Tumoral cell division inhibitory activity in culture correlated with growth inhibition of glioma xenotransplants in Foxn1(nu) nude mice and intracranial glioma allotransplants. Both O-Ac GD1b and O-But GD1b inhibited in vivo cell proliferation, induced cell cycle arrest, and potentiated immune cell response to the tumor. Furthermore, the increased stability of the butyrylated compound (O-But GD1b) enhanced its activity with respect to the acetylated ganglioside (neurostatin). These results are the first report of the antitumoral activity of neurostatin and a neurostatin-like compound in vivo and indicate that semi-synthetic O-acetylated and O-butyrylated gangliosides are potent antitumoral compounds that should be considered in strategies for brain tumor treatment.


RelB-p50 NF-kappa B complexes are selectively induced by cytomegalovirus immediate-early protein 1: differential regulation of Bcl-x(L) promoter activity by NF-kappa B family members.
The NF-kappa B/Rel family has been implicated in control of transcription of the Bcl-x(L) gene, a target which mediates cell survival signals. The cytomegalovirus (CMV) immediate-early protein 1 (IE1) was previously shown to induce NF-kappa B activity. Here, we report that in both vascular smooth muscle cells (SMCs) and NIH 3T3 cells, surprisingly, IE1 failed to induce Bcl-x(L) promoter activity, although it induced activity of E8-CAT, a reporter construct driven by two copies of the NF-kappa B element upstream of the c-myc promoter (upstream regulatory element [URE]). Thus, the subunit nature of the NF-kappa B/Rel factors induced by IE1 was examined using immunofluorescence and immunoblotting. IE1 was found to selectively induce nuclear RelB and p50 in SMCs and NIH 3T3 cells. An increase in RelB protein mediated by IE1 could, in part, be related to an increase in steady-state relB mRNA levels. Consistent with this subunit identification, IE1 was unable to induce E8-CAT activity in relB(-/-) murine embryonic fibroblast cells. In cotransfection analysis of SMCs and NIH 3T3 cells, RelB and p50 proteins failed to induce Bcl-x(L) promoter activity while inducing E8-CAT. Furthermore, the NF-kappa B element of the Bcl-x(L) promoter only weakly bound RelB-p50 complexes compared to the URE NF-kappa B element. Overall, these findings demonstrate in SMCs and NIH 3T3 cells that the CMV IE1 protein selectively induces RelB and p50, which fail to activate the Bcl-x(L) promoter, indicating a strong specificity of binding and activity for the RelB member of the NF-kappa B family. Furthermore, our results implicate RelB in CMV infection of cells such as vascular SMCs.


Aligning climate scenarios to emissions inventories shifts global benchmarks.
Taking stock of global progress towards achieving the Paris Agreement requires consistently measuring aggregate national actions and pledges against modelled mitigation pathways1. However, national greenhouse gas inventories (NGHGIs) and scientific assessments of anthropogenic emissions follow different accounting conventions for land-based carbon fluxes resulting in a large difference in the present emission estimates2,3, a gap that will evolve over time. Using state-of-the-art methodologies4 and a land carbon-cycle emulator5, we align the Intergovernmental Panel on Climate Change (IPCC)-assessed mitigation pathways with the NGHGIs to make a comparison. We find that the key global mitigation benchmarks become harder to achieve when calculated using the NGHGI conventions, requiring both earlier net-zero CO2 timing and lower cumulative emissions. Furthermore, weakening natural carbon removal processes such as carbon fertilization can mask anthropogenic land-based removal efforts, with the result that land-based carbon fluxes in NGHGIs may ultimately become sources of emissions by 2100. Our results are important for the Global Stocktake6, suggesting that nations will need to increase the collective ambition of their climate targets to remain consistent with the global temperature goals.


Emergency management of acute coma in children.
A deteriorating neurologic state in a child with an acute brain insult is a life-threatening situation, and the decisions made during the first few minutes of management will frequently determine the outcome. Often, the family physician is the first person contacted by the parents to evaluate the sudden change in a child. Attention must be first directed toward ensuring an airway and maintaining adequate circulation. Computed tomographic study, toxicology screen and determination of acid-base status should follow. Possible causes of acute coma in children include trauma, child abuse, intracerebral hemorrhage, tumor, meningitis and poisoning.


The Application of Exercise Training for Diabetic Peripheral Neuropathy.
Diabetic peripheral neuropathy (DPN) is the presence of symptoms and/or signs of peripheral nerve dysfunction in people with diabetes after the exclusion of other causes. It is associated with pain, paresthesia, sensory loss, muscle atrophy with fat infiltration, and muscular dysfunction typically starting distally in the feet and progressing proximally. Muscle deterioration within the leg and foot can lead to muscle dysfunction, reduced mobility, and increases the risk of disability, ulceration, and amputation. Exercise training is an established method for increasing the different components of physical fitness, including enhancing body composition and improving neuromuscular strength. A number of experimental studies have utilized exercise training to treat various impairments associated with DPN, such as nerve conduction velocity, pain tolerance, and balance. However, the broad spectrum of exercise training modalities implemented and differences in target outcome measurements have made it difficult to understand the efficacy of exercise training interventions or provide appropriate exercise prescription recommendations. Therefore, the aims of this review were to (1) briefly describe the pathophysiology of DPN and (2) discuss the effects of exercise training interventions on sensorimotor, metabolic, and physical functions in people with DPN.


Septicemia, access and cardiovascular disease in dialysis patients: the USRDS Wave 2 study.
Microinflammation is linked to cardiovascular disease, and is highly prevalent in dialysis patients. It is logical to postulate that septicemia, a common macroinflammatory occurrence in dialysis patients, contributes to their large burden of cardiovascular disease. The Dialysis Morbidity and Mortality Wave 2 was a randomly selected prospective cohort of incident dialysis patients. Admission claims data were used to define and calculate rates of septicemia or bacteremia and cardiovascular events in those with Medicare as the primary payer. Utilizing Cox proportional hazard models we determined the association between baseline access and the development of bacteremia or sepsis, and also the association between bacteremia or sepsis episodes and subsequent cardiovascular events. The 2358 (59%) patients with Medicare as primary payer were older and more likely to have heart failure than those with other payers, but had similar comorbidity-adjusted mortality hazards. Rates of first septicemia, bacteremia, or either condition, were 7.0, 5.9 and 10.4 events per 100-patient years, respectively. Cox regression identified initial dialysis access as the main antecedent of septicemia or bacteremia. Hazards ratios for hemodialysis with permanent catheters, temporary catheters, and grafts were 1.95 (95% CI 1.47-2.57), 1.76 (95% CI 1.29-2.41), and 1.05 (95% CI 0.82-1.35), respectively, while that for peritoneal dialysis was 0.96 (95% CI 0.75-1.23) (reference arteriovenous fistula). After adjustment for baseline factors, septicemia or bacteremia, as a time-dependent covariate, was associated with subsequent death [hazards ratio (HR) 2.33, 95% CI 1.38-2.28], myocardial infarction (HR 1.78, 95% CI 1.38-2.28), heart failure (HR 1.64, 95% CI 1.39-1.95), peripheral vascular disease (HR 1.64, 95% CI 1.34-2.0), and stroke (HR 2.04, 95% CI 1.27-3.28). Septicemia appears to be an important, potentially preventable, cardiovascular risk factor in dialysis patients.


[The interrelationship between immune status characteristics and the trace element level in men with ischemic heart disease].
A comparative analysis was carried out between the hair content of basic trace metals in patients with atherosclerosis, ischemic heart disease (IHD) and findings from their immunogrammes. Close correlation was found out between changes in concentrations of Mn, Cu, Fe, Si, Zn, Ca and processes of activation of basic leucocyte populations characteristic of IHD course and alterations in humoral immune response peculiar to the above patients. Conclusions were reached about presence of the aforementioned changes in the pathogenesis of atherosclerosis, IHD.


In situ cross-linkable hyaluronan hydrogel enhances chondrogenesis.
The present work describes the feasibility of a cross-linkable injectable hyaluronan hydrogel for cartilage repair. The hydrogel used is a two-component system based on aldehyde-modified hyaluronan and hydrazide-modified polyvinyl alcohol, which are rapidly cross-linked in situ upon mixing. The in vitro study showed that chondrocytes and mesenchymal cells cultured in the gel form cartilage-like tissue, rich in glycosaminoglycans, collagen type II and aggrecan. In a rabbit animal model the injection of the hydrogel improved the healing of a full-thickness cartilage defect created in the knee as compared to non-treated controls. This rabbit study showed that the regenerated cartilage defects stained more intensely for type II collagen upon treatment with the hydrogel. The hyaluronan-based hydrogel may be used as a delivery vehicle for both growth factors and/or cells for cartilage repair. The in vivo study also indicated that the hydrogel alone has a beneficial effect on cartilage regeneration.


Investigation of the hypothalamo-pituitary-adrenal axis (HPA) by 1 microg ACTH test and metyrapone test in patients with primary fibromyalgia syndrome.
Primary fibromyalgia syndrome (PFS) is characterized by widespread chronic pain that affects the musculoskeletal system, fatigue, anxiety, sleep disturbance, headache and postural hypotension. The pathophysiology of PFS is unknown. The hypothalamic-pituitary-adrenal (HPA) axis seems to play an important role in PFS. Both hyperactivity and hypoactivity of the HPA axis have been reported in patients with PFS. In this study we assessed the HPA axis by 1 microg ACTH stimulation test and metyrapone test in 22 patients with PFS and in 15 age-, sex-, and body mass index (BMI)- matched controls. Metyrapone (30 mg/kg) was administered orally at 23:00 h and blood was sampled at 08:30 h the following morning for 11-deoxycortisol. ACTH stimulation test was carried out by using 1 microg (iv) ACTH as a bolus injection after an overnight fast, and blood samples were drawn at 0, 30 and 60 min. Peak cortisol level (659.4 +/- 207.2 nmol/l) was lower in the patients with PFS than peak cortisol level (838.7 +/- 129.6 nmol/l) in the control subjects (p < 0.05). Ten patients (45%) with PFS had peak cortisol responses to 1 microg ACTH test lower than the lowest peak cortisol detected in healthy controls. After metyrapone test 11-deoxycortisol level was 123.7 +/- 26 nmol/l in patients with PFS and 184.2 +/- 17.3 nmol/l in the controls (p < 0.05). Ninety five percent of the patients with PFS had lower 11-deoxycortisol level after metyrapone than the lowest 11-deoxycortisol level after metyrapone detected in healthy controls. We also compared the adrenal size of the patients with that of the healthy subjects and we found that the adrenal size between the groups was similar. This study clearly shows that HPA axis is underactivated in PFS, rather than overactivated.


Reduction in mortality after epidural anaesthesia and analgesia in patients undergoing rectal but not colonic cancer surgery: a retrospective analysis of data from 655 patients in central Sweden.
There is some evidence that epidural analgesia (EDA) reduces tumour recurrence after breast and prostatic cancer surgery. We assessed whether EDA reduces long-term mortality after colorectal cancer surgery. All patients having colorectal cancer surgery between January 2004 and January 2008 at Linköping and Örebro were included. Exclusion criteria were: emergency operations, laparoscopic-assisted colorectal resection, and stage 4 cancer. Statistical information was obtained from the Swedish National Register for Deaths. Patients were analysed in two groups: EDA group or patient-controlled analgesia (PCA group) as the primary method of analgesia. A total of 655 patients could be included. All-cause mortality for colorectal cancer (stages 1-3) was 22.7% (colon: 20%, rectal: 26%) after 1-5 yr of surgery. Multivariate regression analysis identified the following statistically significant factors for death after colon cancer (P<0.05): age (>72 yr) and cancer stage 3 (compared with stage 1). A similar model for rectal cancer found that age (>72 yr) and the use of PCA rather than EDA and cancer stages 2 and 3 (compared with stage 1) were associated with a higher risk for death. No significant risk of death was found for colon cancer when comparing EDA with PCA (P=0.23), but a significantly increased risk of death was seen after rectal cancer when PCA was used compared with EDA (P=0.049) [hazards ratio: 0.52 (0.27-1.00)]. We found a reduction in all-cause mortality after rectal but not colon cancer in patients having EDA compared with PCA technique.


Intimate partner violence, partner notification, and expedited partner therapy: a qualitative study.
Over one-third of women experience intimate partner violence (IPV) in their lifetime. IPV increases the risk of infection and re-infection with sexually transmitted infections (STIs). The extent to which health care providers consider IPV when recommending partner notification and expedited partner therapy is unknown. The objective of this qualitative study was to understand health care providers' views on IPV and STIs when recommending partner treatment to patients with chlamydia. Using a purposive sampling strategy to include health care providers who treat young women at risk for chlamydia, 23 semi-structured, in-depth interviews were conducted. While some health care providers expressed concern for their patients' safety and believed assessing for IPV was needed before provision of expedited partner therapy, nearly a third had not considered the links between IPV and STIs. Strategies used by health care providers to assess for IPV did not include inquiry about specific behaviours related to IPV, STI risk, and sexual coercion. Many health care providers understand the risk for IPV in the setting of STI treatment, yet a significant portion of those interviewed failed to recognise the link between IPV and STIs. Provider education is necessary to increase knowledge and implement more effective inquiry and counselling about IPV to more safely recommend expedited partner therapy.


Disease management programs for the underserved.
Disease management has become an important tool for improving population patient outcomes. The Louisiana State University Health Care Services Division (HCSD) has used this tool to provide care to a largely uninsured population for approximately 10 years. Eight programs currently exist within the HCSD focusing on diabetes, asthma, congestive heart failure, HIV, cancer screening, smoking cessation, chronic kidney disease, and diet, exercise, and weight control. These programs operate at hospital and clinic sites located in 8 population centers throughout southern Louisiana. The programs are structured to be managed at the system level with a clinical expert for each area guiding the scope of the program and defining new goals. Care largely adheres to evidence-based guidelines set forth by professional organizations. To monitor quality of care, indicators are defined within each area and benchmarked to achieve the most effective measures in our population. For example, hemoglobin A1c levels have shown improvements with nearly 54% of the population <7.0%. To support these management efforts, HCSD utilizes an electronic data repository that allows physicians to track patient labs and other tests as well as reminders. To ensure appropriate treatment, patients are able to enroll in the Medication Assistance program. This largely improves adherence to medications for those patients unable to afford them otherwise.


Regulation of hypoxia-inducible factor-1alpha, vascular endothelial growth factor, and angiogenesis by an insulin-like growth factor-I receptor autocrine loop in human pancreatic cancer.
Activation of the insulin-like growth factor-I receptor (IGF-IR) was recently shown to modulate angiogenesis by up-regulating the expression of vascular endothelial growth factor (VEGF). We hypothesized that inhibiting IGF-IR function would inhibit angiogenesis and growth of pancreatic cancer in vivo and sought to identify major signaling pathways regulated by IGF-IR in pancreatic cancer cells. Human pancreatic cancer cells (L3.6pl) were stably transfected with a dominant-negative form of IGF-IR (IGF-IR DN) or an empty vector (pcDNA). In vitro, IGF-IR DN cells exhibited a decrease in both constitutive and inducible phosphorylation of IGF-IR and Erk1/2. Constitutive expression of nuclear hypoxia-inducible factor-1alpha and secreted VEGF (P < 0.01) protein levels also were significantly lower in IGF-IR DN cells than in pcDNA cells. In vivo, IGF-IR inhibition led to decreases in pancreatic tumor volume and weight, vessel density, and tumor cell proliferation (P < 0.01 for all) and increases in tumor cell apoptosis (P < 0.02). Our results suggest that autocrine activation of the IGF-IR system significantly affects VEGF expression and angiogenesis in human pancreatic cancer. Thus, IGF-IR may be a valid target in the treatment of pancreatic cancer.


The selective use of magnetic resonance cholangiopancreatography in the imaging of the axial biliary tree in patients with acute gallstone pancreatitis.
Magnetic resonance cholangiopancreatography (MRCP) is an emerging modality in the management of acute gallstone pancreatitis (AGP). The aim of this study was to assess the impact following the introduction of MRCP in the management of AGP in a tertiary referral unit. Patients presenting with AGP from January 2002 to December 2004 were reviewed to assess the impact of the introduction of MRCP in June 2003. The indication for MRCP was suspected common bile duct (CBD) stones in the absence of biliary sepsis. Definitive treatment for AGP was laparoscopic cholecystectomy, with endoscopic retrograde cholangiopancreatography (ERCP) and sphincterotomy reserved for patients unfit for cholecystectomy and those with biliary sepsis. 249 patients were identified of whom 36 (14.5%) underwent ERCP and sphincterotomy as definitive treatment. 96 patients with a non-dilated CBD and normal or resolving liver function tests proceeded to laparosocopic cholecystectomy and intraoperative cholangiogram (IOC), 8 (8.5%) of whom had CBD stones intraoperatively. Eleven patients underwent cholecystectomy during pancreatic necrosectomy. Of those undergoing preoperative diagnostic biliary tract imaging, ERCP was undertaken in 57 patients and MRCP in 49 patients. There was no significant difference in serum bilirubin levels [ERCP 43 mmol/l (18-204) vs. MRCP 39 mmol/l (24-180), p = NS] or the proportion of patients with CBD stones [ERCP 10 (17.5%) vs. MRCP 7 (14.2%), p = NS] between the two groups. Patients who underwent MRCP had a shorter median hospital stay [MRCP 5 days (range: 3-14) vs. ERCP 9 days (range: 4-20), p < 0.01] and higher rate of cholecystectomy during the index admission (MRCP 83.3% vs. ERCP 67.2%, p < 0.05). There was a high degree of correlation between preoperative MRCP results and findings of subsequent IOC or therapeutic ERCP (area under ROC curve: 0.94). MRCP is an accurate modality for imaging the axial biliary tree in patients with AGP. Selective use of MRCP reduces the need for ERCP and results in shorter hospital stay. and IAP.


Recommendations for Face Coverings While Exercising During the COVID-19 Pandemic.
In an effort to reduce transmission and number of infections of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or COVID-19) virus, governments and official bodies around the world have produced guidelines on the use of face masks and face coverings. While there is a growing body of recommendations for healthcare professionals and the wider population to use facial protection in "enclosed spaces" where minimal distancing from other individuals is not possible, there is a dearth of clear guidelines for individuals undertaking exercise and sporting activity. The present viewpoint aims to propose recommendations for face coverings while exercising during the COVID-19 pandemic that consider physical distancing, the environment, the density of active cases associated with the specific sports activity, and the practical use of face coverings in order to reduce potential viral transmission. Recommendations are provided on the basis of very limited available evidence in conjunction with the extensive collective clinical experience of the authors and acknowledging the need to consider the likelihood of the presence of the SARS-CoV-2 in the general population. We recommend that face coverings should be used in any environment considered to be of a high or moderate transmission risk, where tolerated and after individual risk assessment. In addition, as national caseloads fluctuate, individual sporting bodies should consider up to date guidance on the use of face coverings during sport and exercise, alongside other preventative measures.


Profilometer evaluation of the effect of various polishing methods on the surface roughness in dental ceramics of different structures subjected to repeated firings.
To evaluate the changes in surface roughness of dental ceramics with various substances/instruments and polishing methods during the course of repeated firings. Three types of dental ceramic (IPS Classic, Empress Esthetic, and Empress 2 [all by Ivoclar Vivadent]) were used to form three groups, respectively. The samples from the first group were subjected to glaze, the -second group to natural glaze, and the third group to mechanical polishing. As many as seven firings were performed on each sample, and surface roughness measurements of the samples were carried out at certain stages. Statistical evaluation was performed through repeated-measures ANOVA, and the Duncan test proved that the differences among the mean values were by no means a coincidence. The interaction among the substrates, the method used, and the time was statistically significant (P < .05). Mechanical polishing significantly increased surface roughness (P < .05), and the greater number of firings significantly decreased the amount of surface roughness (P < .05). In terms of surface roughness, the best method available is the glaze method and the best material is IPS Classic. The mechanical polishing process generates rough surfaces. Repeated firings should be avoided as much as possible because they have a destructive effect on the glaze layer and deform the surface.


Stereoselectivity of ectonucleotidases on vascular endothelial cells.
We have investigated the stereoselectivity of ectonucleotidases (nucleoside triphosphatase, EC 184.108.40.206; nucleoside diphosphatase, EC 22.214.171.124; 5'-nucleotidase, EC 220.127.116.11) on pig aortic endothelial cells using two classes of nucleotide analogue. In experiments with nucleotide enantiomers in which the natural D-ribofuranosyl moiety is replaced by an L-ribofuranosyl moiety, the rate of catabolism of 100 microM-L-ATP was one-fifth that of D-ATP, the rate of catabolism of 100 microM-L-ADP was one-fifteenth that of D-ADP and there was no detectable catabolism of 100 microM-L-AMP. Each of the L-enantiomers inhibited, apparently competitively, the catabolism of the corresponding D-enantiomer; Ki values were approx. 0.6 mM, 1.0 mM and 3.9 mM for L-ATP, L-ADP and L-AMP respectively. Experiments with adenosine 5'-[beta, gamma-imido]triphosphate and with D- and L-enantiomers of adenosine 5'-[beta, gamma-methylene]triphosphate revealed modest ectopyrophosphatase activity, undetectable in experiments with natural nucleotides, which was also stereoselective. Use of phosphorothioate nucleotide analogues demonstrated that ATP catabolism was virtually stereospecific with respect to the geometry of the thiol group substituted on the beta-phosphate: the Rp isomer was degraded, whereas there was little or no breakdown of the Sp isomer. ADP catabolism was also stereospecific with respect to the geometry of the thiol group substituted on the alpha-phosphate: the Sp isomer but not the Rp isomer was degraded. The geometry of thiol-group substitution on the alpha-phosphate had no effect on ATP catabolism to ADP. There was no detectable catabolism of analogues with thiol-group substitution on the terminal phosphate. Each of the phosphorothioate analogues that was catabolized broke down at a rate similar to that of the natural nucleotide from which it was derived. These results demonstrate that the ectonucleotidases on pig aortic endothelial cells exhibit a high degree of stereoselectivity, characteristic for each enzyme, both with respect to the ribofuranosyl moiety and to the phosphate side chain.


Pseudomonas aeruginosa exotoxin A reduces chemoresistance of oral squamous carcinoma cell via inhibition of heat shock proteins 70 (HSP70).
Oral squamous carcinoma (OSCC) cells exhibit resistance to chemotherapeutic agent-mediated apoptosis in the late stage of malignancy. Increased levels of heat shock proteins 70 (HSP70) in cancer cells are known to confer resistance to apoptosis. Since recent advances in the understanding of bacterial toxins have produced new strategies for the treatment of cancers, we investigated the effect of Pseudomonas aeruginosa exotoxin A (PEA) on HSP70 expression and induction of apoptosis in chemoresistant OSCC cell line (YD-9). The apoptotic effect of PEA on chemoresistant YD-9 cells was confirmed by MTT, Hoechst and TUNEL stains, DNA electrophoresis, and Western blot analysis. While YD-9 cells showed high resistance to chemotherapeutic agents such as etoposide and 5-fluorouraci (5-FU), HSP70 antisense oligonucelotides sensitized chemoresistant YD-9 cells to etoposide and 5-FU. On the other hand, PEA significantly decreased the viability of YD-9 cells by deteriorating the HSP70-relating protecting system through inhibition of HSP70 expression and inducing apoptosis in YD-9 cells. Apoptotic manifestations were evidenced by changes in nuclear morphology, generation of DNA fragmentation, and activation of caspases. While p53, p21, and E2F-1 were upregulated, cdk2 and cyclin B were downregulated by PEA treatment, suggesting that PEA caused cell cycle arrest at the G2/M checkpoint. Therefore, these results indicate that PEA reduced the chemoresistance through inhibition of HSP70 expression and also induced apoptosis in chemoresistant YD-9 cells.


Microfabricated porous layer open tubular (PLOT) column.
Development of micro gas chromatography (μGC) is aimed at rapid and in situ analysis of volatile organic compounds (VOCs) for environmental protection, industrial monitoring, and toxicology. However, due to the lack of appropriate microcolumns and associated stationary phases, current μGC is unable to separate highly volatile chemicals such as methane, methanol, and formaldehyde, which are of great interest for their high toxicity and carcinogenicity. This inability has significantly limited μGC field applicability. To address this deficiency, this paper reports the development and characterization of a microfabricated porous layer open tubular (μPLOT) column with a divinylbenzene-based stationary phase. The separation capabilities of the μPLOT column are demonstrated by three distinct analyses of light alkanes, formaldehyde solution, and organic solvents, exhibiting its general utility for a wide range of highly volatile compounds. Further characterization shows the robust performance of the μPLOT column in the presence of high moisture and at high temperatures (up to 300 °C). The small footprint and the ability to separate highly volatile chemicals make the μPLOT column highly suitable for integration into μGC systems, thus significantly broadening μGC's applicability to rapid, field analysis of VOCs.


Optical Analysis and Reappraisal of the Peripheral Light Focusing Theory of Nasal Pterygia Formation.
Pterygia are much more common nasally than temporally. Ultraviolet (UV) radiation is a major risk factor. Coroneo proposed that the nasal preference is caused by the "peripheral light focusing effect," (PLF), in which UV at an oblique angle passes through temporal cornea and is concentrated on and damages nasal limbal stem cells. This study evaluates whether the PLF is sufficient to explain the nasal preference. Whereas Coroneo and colleagues derived the maximum PLF intensity gain (UV concentration factor) as a function of incident angle (i.e., different nasal limbal positions were used for different incident angles) the current analysis derived intensity gain at a fixed position such at the nasal corneo-limbal junction (CLJ). This provided a measure of the total PLF irradiation at this position, which was compared to total direct irradiation of nasal and temporal limbus at the corresponding positions (e.g., CLJs). In Part 1, analysis was performed like that of Coroneo, using horizontally incident UV; in Part 2, the analysis was extended to include incident rays above and below the horizontal. In both part 1 and part 2 of the study, the limbal UV irradiation of the nasal limbus from the PLF was not sufficient to explain the strong nasal location preference of pterygia. The analysis calls into question the PLF explanation of nasal location preference. Other explanations of the nasal preference, and of pterygium pathogenesis, should be considered, such as temporal to nasal tear flow carrying substances such as cytokines to the nasal limbus.


Packaging the male germline in plants.
The development of plant lateral organs is interesting because, although many of the same genes seem to be involved in the early growth of primordia, completely different gene combinations are required for the complete development of organs such as leaves and stamens. Thus, the genes common to the development of most organs, which generally form and polarize the primordial 'envelope', must at some stage interact with those that 'install' the functional content of the organ--in the case of the stamen, the four microsporangia. Although distinct genetic pathways of organ initiation, polarity establishment and setting up the reproductive cell line can readily be recognized, they do not occur sequentially. Rather, they are activated early and run in parallel. There is evidence for continuing crosstalk between these pathways.


Overall survival and causes of death in elderly patients with Hodgkin lymphoma: a Norwegian population-based case-control study.
Elderly Hodgkin Lymphoma (HL) patients are poorly characterized and underrepresented in studies. In this national population-based study, we investigated cause-specific survival using competing-risk analysis in elderly HL patients compared to the normal population. Patients ≥ 60 years diagnosed between 2000-2015 were identified by Cancer Registry of Norway, records reviewed in detail and compared to data from Norwegian Cause of Death Registry for patients and cancer-free controls. Of 492 patients, 81 (17%) were ineligible for treatment directed specifically towards HL, mostly because of an underlying other lymphoma entity, whereas 74 (15%) and 337 (69%) were treated with palliative or curative intent, respectively. Median overall survival in patients ineligible for assessment of HLdirected therapies was 0.5 years (95% confidence interval [CI] 0.4-0.6), and for palliatively and curatively treated patients 0.8 (0.4-1.2) and 9.1 (7.5-10.7) years, respectively. After correction of discrepancies in registry data, with 359 deaths, 108 (30%) died of HL, the most common cause of death. In curatively treated patients, treatment-related mortality was 6.5% and the risk-difference of dying from HL compared to controls was 28% (95% CI 23-33%) after 10 years. These numbers indicate disease control in a majority of elderly patients eligible for curative treatment, compared to risk-differences for death from HL of 59% (48-71%) and 42% (31-53%) after 10 years in the palliative and ineligible groups, respectively. There was an increased risk of dying from hematological malignancies other than HL in all groups, but not from other competing causes of death, showing no excess mortality from long-term treatment complications.


Therapeutic effectiveness of Galphimia glauca vs. lorazepam in generalized anxiety disorder. A controlled 15-week clinical trial.
Galphimia glauca Cav. has demonstrated anxiolytic activity attributable to nor-seco-triterpenes denominated galphimines, the most active of which is galphimine-B. Galphimine-B inhibits ventral tegmental area dopaminergic neurons and interacts with the serotoninergic system of the dorsal hippocampus. A previous clinical study that administered a G. glauca herbal medicinal product for 4 weeks evidenced high percentages of therapeutic effectiveness and safety in patients with generalized anxiety disorder. Based on the previous findings, the goal of the present study was to evaluate the effectiveness, safety, and tolerability of G. glauca herbal medicinal product administered during 15 weeks in patients with generalized anxiety disorder. double-blind, randomized, lorazepam-controlled clinical trial. adult males and females, ambulatory, diagnosed with generalized anxiety disorder, with 20 or more points on the Hamilton anxiety scale, without data of depression, and without anxiolytic treatment in the previous month. Interventions were as follows. Experimental treatment: G. glauca herbal medicinal product in capsules containing the dry extract of G. glauca standardized in 0.175 mg of galphimine-B, one or two capsules twice a day, during 12 weeks plus 3 withdrawal weeks, and control treatment: lorazepam 0.5 mg with the same presentation and posology. anxiolytic effectiveness (≥ 50 % reduction of initial Hamilton anxiety scale score). tolerability and safety. One hundred ninety-one patients initiated the study with 94 in the experimental group. One hundred four patients concluded the study, 51 of these in the experimental group. Anxiolytic effectiveness, measured as 0 in a negative case and as 1 in a positive case, was assessed 593 times in the experimental group and 631 in the control; the mean effectiveness observed was 0.686 ± 0.019 vs. 0.588 ± 0.019 (repeated-measures ANOVA; p = 0.0003). In the same way, G. glauca-herbal medicinal product diminished the score in the Hamilton anxiety scale to 11.51 ± 8.27 points and lorazepam to 12.40 ± 8.07 points (repeated-measures ANOVA; p = 0.05). The tolerability analysis, which comprised patients who concluded the treatment plus 11 patients who withdrew due to adverse reactions did not show differences between treatments (p = 0.35), nor did therapeutic safety demonstrate differences between groups (p = 0.21). There were no cases of tolerance, intoxication, dependence, or suppression syndrome. We concluded that G. glauca herbal medicinal product, standardized in 0.175 mg of galphimine-B and administered for 15 weeks to patients with generalized anxiety disorder, showed greater anxiolytic effectiveness than that obtained with lorazepam, with high percentages of therapeutic tolerability and safety.


Cutting edge: roles of Toll-like receptor 4 and IL-23 in IL-17 expression in response to Klebsiella pneumoniae infection.
Local production of IL-17 is a significant factor in effective host defense against Gram-negative bacteria. However, the proximal events mediating IL-17 elaboration by T cells remain unclear. In this study, we show in vivo that intact Toll-like receptor 4 signaling in the lung is required for induction of both the p19 transcript of IL-23 and IL-17 protein elaboration in response to Klebsiella pneumoniae. Although IL-17 is widely considered a CD4(+) T cell product, we also demonstrate significant in vitro IL-17 production by CD8(+) T cells after culture in medium from dendritic cells exposed to these bacteria. The dominant portion of this IL-17-inducing activity for both CD4(+) and CD8(+) T cells is IL-23. These data demonstrate the critical signaling pathway for IL-17 induction in the host response to Gram-negative pulmonary infection and suggest a direct role for IL-23 in CD8(+) T cell IL-17 production.


The conceptualization of perceived insufficient milk among Mexican mothers.
This study sought to provide a comprehensive picture of maternal conceptualization of Perceived Insufficient Milk (PIM) using qualitative methods (semistructured interviews, free lists, and rankings) in Mexico. Two hundred and seven first-time mothers of infants younger than 6 months and mothers-to-be who were (or intended to be) breastfeeding were interviewed in waiting areas at a hospital and a health clinic in Mexico City, Mexico, from September 2000 to January 2001. Pattern searching and triangulation of the 3 qualitative methods indicated that women viewed crying as the chief symptom of PIM. Maternal diet and liquid intake were cited as both the most salient causes and treatments for PIM. The main coping strategy in the event of PIM was formula feeding. These findings highlight the need for addressing maternal concerns in relation to PIM, especially regarding the role of crying as the initiator of the PIM cycle.


[Dysontogenetic median maxillary cysts].
Nasopalatine duct cysts, median palatine cysts and nasoalveolar cysts belong to the developmental cysts of the upper jaw. They are situated at the midline in the fusion areas of embryonic facial processes and for this reason are known as "fissural cysts". They are, therefore, not of odontogenic origin and, on account of their relationship to the mouth, the nose and the nasal cavities, are of practical relevance to the otolaryngologist. The various clinical symptoms and the histological findings are described, and a review of the literature is presented.


[Current Topics on Vitamin D. Vitamin D and chronic kidney disease].
CKD-MBD is a new clinical concept which defines a systemic disorder of mineral and bone metabolism as a result of CKD. Although various mineral metabolic factors are involved in its pathogenesis, alteration in vitamin D metabolism plays a crucial role among them. Vitamin D receptor activators (VDRAs), which are traditionally considered a cardinal treatment of secondary hyperparathyroidism, have attracted lots of attention for their potential pleiotropic effects through the activation of the vitamin D receptor expressed in multiple organs. In recent years, the deficiency of vitamin D in CKD patients also has received growing interest. The serum 25 (OH) D levels are considered as a useful prognostic marker, and the significances of nutritional vitamin D supplementation are reviewed from a new perspective.


Maternal and family factors differentiate profiles of psychiatric impairments in very preterm children at age 5-years.
Very preterm (VPT; <30 weeks gestation) children are a heterogeneous group, yet the co-occurrence of psychiatric and neurodevelopmental impairments remains unclear. Moreover, the clinical and socio-environmental factors that promote resilient developmental outcomes among VPT children are poorly understood. One hundred and twenty five children (85 VPT and 40 full-term) underwent neurodevelopmental evaluation at age 5-years. Parents and teachers completed measures of internalizing, externalizing, attention-deficit/hyperactivity (ADHD), and autism symptoms. Psychiatric and neurodevelopmental measures were analyzed using Latent Profile Analysis. Multinomial regression examined the extent that infant, sociodemographic, and family factors, collected prospectively from birth to follow-up, independently differentiated resilient and impaired children. Four latent profiles were identified, including a Typically Developing Group which represented 27.1% of the VPT group and 65.0% of the full-term group, an At-Risk Group with mild psychiatric and neurodevelopmental problems (VPT 44.7%, full-term 22.5%), a Psychiatric Group with moderate-to-severe psychiatric ratings (VPT 12.9%, full-term 10.0%), and a school-based Inattentive/Hyperactive Group (VPT 15.3%, full-term 2.5%). Clinical diagnoses were highest among the Psychiatric Group (80%). Factors that differentiated resilient and impaired subgroups of VPT children included prolonged exposure to maternal psychosocial distress (p ≤ .04), current family dysfunction (p ≤ .05), and maternal ADHD symptoms (p ≤ .02), whereas social risk index scores differentiated resilient and impaired full-term children (p < .03). Lower levels of maternal distress, family dysfunction, and maternal ADHD symptoms were associated with resilience among VPT children. Maternal distress and family dysfunction are modifiable factors to be targeted as part of psychiatric interventions embedded in the long-term care of VPT children.


Does physical activity moderate the association between alcohol drinking and all-cause, cancer and cardiovascular diseases mortality? A pooled analysis of eight British population cohorts.
To examine whether physical activity (PA) moderates the association between alcohol intake and all-cause mortality, cancer mortality and cardiovascular diseases (CVDs) mortality. Prospective study using 8 British population-based surveys, each linked to cause-specific mortality: Health Survey for England (1994, 1998, 1999, 2003, 2004 and 2006) and Scottish Health Survey (1998 and 2003). 36 370 men and women aged 40 years and over were included with a corresponding 5735 deaths and a mean of 353 049 person-years of follow-up. 6 sex-specific categories of alcohol intake (UK units/week) were defined: (1) never drunk; (2) ex-drinkers; (3) occasional drinkers; (4) within guidelines (<14 (women); <21 (men)); (5) hazardous (14-35 (women); 21-49 (men)) and (6) harmful (>35 (women) >49 (men)). PA was categorised as inactive (≤7 MET-hour/week), active at the lower (>7.5 MET-hour/week) and upper (>15 MET-hour/week) of recommended levels. Cox proportional-hazard models were used to examine associations between alcohol consumption and all-cause, cancer and CVD mortality risk after adjusting for several confounders. Stratified analyses were performed to evaluate mortality risks within each PA stratum. We found a direct association between alcohol consumption and cancer mortality risk starting from drinking within guidelines (HR (95% CI) hazardous drinking: 1.40 (1.11 to 1.78)). Stratified analyses showed that the association between alcohol intake and mortality risk was attenuated (all-cause) or nearly nullified (cancer) among individuals who met the PA recommendations (HR (95% CI)). Meeting the current PA public health recommendations offsets some of the cancer and all-cause mortality risk associated with alcohol drinking.


Two-dimensional vector accelerometer based on Bragg gratings inscribed in a multi-core fiber.
We propose and demonstrate a novel orientation-sensitive two-dimensional accelerometer based on fiber Bragg gratings inscribed in a multi-core fiber. Through monitoring of the wavelength shifts of three of the seven cores, including the central core and two outer cores which are not aligned in a straight line, information on vibration orientation as well as acceleration can be obtained simultaneously. Performance of the proposed accelerometer in terms of frequency, acceleration and vibration orientation are experimentally investigated. The designed two-dimensional accelerometer is capable of obtaining all these three parameters simultaneously. A sensitivity which is strongly dependent on the orientation is achieved, with a best orientation accuracy of 0.127° over a range of 0-180°. Moreover, the resonance frequency and the sensitivity can be optimized through adjusting the length and weight of the free-fiber. The ease of fabrication as well as the versatility of the proposed sensor makes it potentially useful in dynamic monitoring for industrial applications.


Preliminary experience with the multisensor HeartLogic algorithm for heart failure monitoring: a retrospective case series report.
In the Multisensor Chronic Evaluation in Ambulatory Heart Failure Patients study, a novel algorithm for heart failure (HF) monitoring was implemented. The HeartLogic (Boston Scientific) index combines data from multiple implantable cardioverter defibrillator (ICD)-based sensors and has proved to be a sensitive and timely predictor of impending HF decompensation. The remote monitoring of HF patients by means of HeartLogic has never been described in clinical practice. We report post-implantation data collected from sensors, the combined index, and their association with clinical events during follow-up in a group of patients who received a HeartLogic-enabled device in clinical practice. Patients with ICD and cardiac resynchronization therapy ICD were remotely monitored. In December 2017, the HeartLogic feature was activated on the remote monitoring platform, and multiple ICD-based sensor data collected since device implantation were made available: HeartLogic index, heart rate, heart sounds, thoracic impedance, respiration, and activity. Their association with clinical events was retrospectively analysed. Data from 58 patients were analysed. During a mean follow-up of 5 ± 3 months, the HeartLogic index crossed the threshold value (set by default to 16) 24 times (over 24 person-years, 0.99 alerts/patient-year) in 16 patients. HeartLogic alerts preceded five HF hospitalizations and five unplanned in-office visits for HF. Symptoms or signs of HF were also reported at the time of five scheduled visits. The median early warning time and the time spent in alert were longer in the case of hospitalizations than in the case of minor events of clinical deterioration of HF. HeartLogic contributing sensors detected changes in heart sound amplitude (increased third sound and decreased first sound) in all cases of alerts. Patients with HeartLogic alerts during the observation period had higher New York Heart Association class (P = 0.025) and lower ejection fraction (P = 0.016) at the time of activation. Our retrospective analysis indicates that the HeartLogic algorithm might be useful to detect gradual worsening of HF and to stratify risk of HF decompensation.


Vesicular-arbuscular mycorrhizae in relation to plant disease.
Vesicular-arbuscular mycorrhizae (VAM) enhance plant growth through increased nutrient uptake, stress tolerance and disease resistance. As an integral part of the root system, they interact with other microorganisms in soil and result in increased root exudation approaching about 25% of the plant dry matter production. Roots support a multitude of microorganisms that, in concert, can have profound influence on growth and survival of the plant. VAM fungi can alter the root exudation pattern, enhance chitinolytic activity and alter photosynthetic/respiratory deficiencies. VAM-positive plants are known to exhibit varied resistance towards soil-borne and foliar pathogens. The known interactions include a number of mechanisms, such as exclusion of the pathogen, lignification of plant cell walls, changed phosphate nutrition resulting in altered exudation by roots, and formation of inhibitory low molecular weight compounds. The purpose of this review is to discuss VAM-plant-pathogen interactions and the possible mechanisms involved in altered resistance. Based on these observations, a working model is proposed to explain the VAM-disease interaction under varied environmental conditions.


Laboratory performance in albumin and total protein measurement using a commutable specimen: results of a College of American Pathologists study.
Discrepant results for serum constituents were observed among peer groups in the College of American Pathologists Comprehensive Chemistry Survey. To assess the performance of serum albumin and total protein measurement procedures and to evaluate the commutability of the conventional survey specimens. A fresh frozen, off-the-clot serum sample was included along with 4 conventional survey specimens. The fresh frozen, off-the-clot serum sample was prepared in a manner expected to confer commutability with native clinical samples. For the fresh frozen, off-the-clot serum sample, the mean values for 17 peer-groups were -0.07 to 0.32 g/dL from the bromocresol green albumin designated comparison method, whereas 4 VITROS (Ortho Clinical Diagnostics, Rochester, New York) peer groups differed by -0.29 to -0.37 g/dL (15 of 21 differences [71%] had P < .001). For bromocresol purple albumin methods, the mean differences from the designated comparison method from 8 peer groups were 0.25 to 0.47 g/dL (all had P < .001). For total protein methods, 23 peer group mean values were -0.07 to 0.15 g/dL from the reference measurement procedure (12 of 24 [50%] had P < .001). The Beckman (Fullerton, California) Synchron LX20 had a bias of -0.30 g/dL (P <.001). The commutability of the conventional specimens was acceptable for 23 of 24 bromocresol green method-material combinations (96%) and 13 of 16 bromocresol purple albumin method-material combinations (81%). All (100%) of the 36 method-material combinations had acceptable commutability for total protein. One (2.2%) of the instrument systems (Synchron) using bromocresol green and none (0%) of the instrument systems using bromocresol purple had satisfactory total-error performance for albumin measurement. Differences in results between bromocresol green and bromocresol purple methods precluded using common reference intervals for interpreting results for serum albumin. Eight of 9 instrument systems (86.5%) had satisfactory total-error performance for total protein measurement.


Genetic and evolutionary patterns of treatment resistance in relapsed B-cell lymphoma.
Diffuse large B-cell lymphoma (DLBCL) patients are typically treated with immunochemotherapy containing rituximab (rituximab, cyclophosphamide, hydroxydaunorubicin-vincristine (Oncovin), and prednisone [R-CHOP]); however, prognosis is extremely poor if R-CHOP fails. To identify genetic mechanisms contributing to primary or acquired R-CHOP resistance, we performed target-panel sequencing of 135 relapsed/refractory DLBCLs (rrDLBCLs), primarily comprising circulating tumor DNA from patients on clinical trials. Comparison with a metacohort of 1670 diagnostic DLBCLs identified 6 genes significantly enriched for mutations upon relapse. TP53 and KMT2D were mutated in the majority of rrDLBCLs, and these mutations remained clonally persistent throughout treatment in paired diagnostic-relapse samples, suggesting a role in primary treatment resistance. Nonsense and missense mutations affecting MS4A1, which encodes CD20, are exceedingly rare in diagnostic samples but show recurrent patterns of clonal expansion following rituximab-based therapy. MS4A1 missense mutations within the transmembrane domains lead to loss of CD20 in vitro, and patient tumors harboring these mutations lacked CD20 protein expression. In a time series from a patient treated with multiple rounds of therapy, tumor heterogeneity and minor MS4A1-harboring subclones contributed to rapid disease recurrence, with MS4A1 mutations as founding events for these subclones. TP53 and KMT2D mutation status, in combination with other prognostic factors, may be used to identify high-risk patients prior to R-CHOP for posttreatment monitoring. Using liquid biopsies, we show the potential to identify tumors with loss of CD20 surface expression stemming from MS4A1 mutations. Implementation of noninvasive assays to detect such features of acquired treatment resistance may allow timely transition to more effective treatment regimens.


The relationship of patient medical and laboratory characteristics to changes in functional health status in children and adolescents after the Fontan procedure.
Despite hypothesized concerns about deterioration beginning in adolescence, longitudinal data and associated factors regarding standardized assessment of physical functioning are not available for Fontan patients. Parents who participated in the Fontan Cross-Sectional Study completed the Child Health Questionnaire at 2 time points for 245 subjects ages 6-18 years. Associations between change in Physical Functioning Summary Score and baseline patient, medical, and laboratory characteristics (mean age 9.5 ± 1.7 years) and follow-up patient and medical characteristics (mean age 16.2 ± 1.6 years) were determined by regression analyses. During a mean of 6.7 ± 0.4 years, a small (not clinically important) but statistically significant decrease in score from 46.2 ± 11.7 to 44.5 ± 12.1 (p < 0.03) was noted. Subjects with higher baseline scores had a greater decrease in score (r = -0.48; p < 0.001). A multivariable model of patient and medical characteristics (R(2) = 0.11) showed that a greater decrease in score was significantly associated with interim development of asthma (n = 13; parameter estimate [PE] -6.6; p < 0.05) or other chronic respiratory, lung, or breathing problems (n = 13; PE -12.5; p < 0.001) and the presence of protein-losing enteropathy at any time (n = 12; PE -9.4; p = 0.006). Change in score was not significantly associated with baseline laboratory measures of exercise capacity and ventricular characteristics and function. Therefore, although physical functioning may be stable during adolescence for many Fontan patients, deterioration occurs in some in association with respiratory conditions and protein-losing enteropathy. Further longitudinal study is necessary to better understand the relationship between clinical morbidities and functional health status as these patients transition into adulthood.


Characteristics of early murine B-lymphocyte precursors and their direct sensitivity to negative regulators.
Recently, a collection of surface markers was exploited to isolate viable Lin(-) TdT(+) cells from murine bone marrow. These early pro-B cells were enriched for B-lineage lymphocyte precursor activity measured by short-term culture and had little responsiveness to myeloid growth factors. Early precursors can be propagated with remarkably high cloning frequencies in stromal cell-free, serum-free cultures, permitting this analysis of direct regulatory factors. Expression of the interleukin-7 receptor (IL-7Ralpha) chain marks functional precursors and IL-7 is necessary for progression beyond the CD45RA(+) CD19(-) stage. Efficient survival and differentiation were only observed when stem cell factor and Flt-3 ligand were also present. IL-7-responsive CD19(+) precursors are estrogen resistant. However, B-lineage differentiation was selectively abrogated when highly purified Lin(-) precursors were treated with hormone in the absence of stromal cells. In addition, early stages of B lymphopoiesis were arrested by limitin, a new interferon (IFN)-like cytokine as well as IFN-alpha, IFN-gamma, or transforming growth factor beta (TGF-beta), but not by epidermal growth factor (EGF). Lin(-) TdT(+) early pro-B cells are shown here to be CD27(+) AA4.1(+/-)Ki-67(+) Ly-6C(-) Ly-6A/Sca-1(Lo/-)Thy-1(-)CD43(+) CD4(+/-)CD16/32(Lo/-)CD44(Hi) and similar in some respects to the "common lymphoid progenitors" (CLP) identified by others. Although early pro-B cells have lost myeloid differentiation potential, transplantation experiments described here reveal that at least some can generate T lymphocytes. Of particular importance is the demonstration that a pivotal early stage of lymphopoiesis is directly sensitive to negative regulation by hormones and cytokines.


Role of Insulin-Like Growth Factor Binding Protein-3 in the Pathogenesis of Herpes Stromal Keratitis.
The goal of this study was to determine the role of insulin-like growth factor-binding protein-3 (IGFBP-3) in the pathogenesis of herpes stromal keratitis (HSK). In an unbiased approach, a membrane-based protein array was carried out to determine the level of expression of pro- and anti-angiogenic molecules in uninfected and HSV-1 infected corneas. Quantitative RT-PCR and ELISA assays were performed to measure the amounts of IGFBP-3 at mRNA and protein levels. Confocal microscopy documented the localization of IGFBP-3 in uninfected and infected corneal tissue. Flow cytometry assay showed the frequency of immune cell types in infected corneas from C57BL/6J (B6) and IGFBP-3 knockout (IGFBP-3-/-) mice. Slit-lamp microscopy was used to quantitate the development of opacity and neovascularization in infected corneas from both groups of mice. Quantitation of protein array dot blot showed an increased level of IGFBP-3 protein in HSV-1 infected than uninfected corneas and was confirmed with ELISA and quantitative RT-PCR assays. Cytosolic and nuclear localization of IGFBP-3 were detected in the cells of corneal epithelium, whereas scattered IGFBP-3 staining was evident in the stroma of HSK developing corneas. Increased opacity and hemangiogenesis were noted in the corneas of IGFBP-3-/- than B6 mice during the clinical period of HSK. Furthermore, an increased number of leukocytes comprising of neutrophils and CD4 T cells were found in HSK developing corneas of IGFBP-3-/- than B6 mice. Our data showed that lack of IGFBP-3 exacerbates HSK, suggesting the protective effect of IGFBP-3 protein in regulating the severity of HSK.


Expression and domain-specific function of GATA-2 during differentiation of the hematopoietic precursor cells in midgestation mouse embryos.
The aorta-gonads-mesonephros (AGM) region of the mouse embryo has been assigned as the origin of definitive hematopoiesis. The transcription factor GATA-2 has specific but unclarified roles in early hematopoiesis. To elucidate the expression profile of GATA-2, we prepared transgenic mouse lines containing the green fluorescent protein (GFP) gene driven by GATA-2 gene regulatory elements. We also prepared a mouse line in which GFP reporter sequences were inserted into the endogenous GATA-2 gene. Both mouse mutants expressed GFP in the early hematopoietic tissues. The CD45 antigen, a marker of hematopoietic cells, was expressed in a small fraction of transgene (TG)-derived GFP+ cells. The remaining TG-GFP+/CD45- cells were adherent to plastic and produced CD45+ hematopoietic cells abundantly when cultured in vitro. Exogenous expression of GATA-2 in TG-GFP+/CD45- cells from the AGM region inhibited their differentiation into CD45+ cells. Loss of GATA-2 function through the disruption of the GATA-2 locus enhanced the earlier emergence of CD45+ cells in the yolk sac of the 9.5-day conceptus. These results demonstrated that GATA-2 is expressed in the precursor of hematopoietic cells and works as a gatekeeper to preserve their immaturity. A reduction of GATA-2 expression or activity is required for the differentiation of precursors to hematopoietic cells.


Improved four-color flow cytometry method using fluo-3 and triple immunofluorescence for analysis of intracellular calcium ion ([Ca2+]i) fluxes among mouse lymph node B- and T-lymphocyte subsets.
A visible-light, dual-laser, flow cytometric method was developed for the simultaneous analysis of intracellular ionized calcium concentration ([Ca2+]i) and three cell-surface markers (CD4, CD8, and Thy-1.2 antigens) by using the calcium probe fluo-3 and using R-phycoerythrin (PE), peridinin chlorophyll-alpha protein (PerCP), and allophycocyanin (APC) conjugated monoclonal antibodies (MoAbs). This improved method was used in the analysis of [Ca2+]i mobilization upon in vitro stimulation with mitogenic lectins [phytohaemagglutinin (PHA) or concanavalin A (ConA)], anti-CD3 MoAbs, or A23187 calcium ionophore in the heterogeneous lymph node cell populations from healthy C57BL/Ka mice. The present results show that the calcium responses were heterogeneous and dependent on the cellular immunophenotype, not only on lectins or anti-CD3 MoAbs stimulation, but also on the receptor-independent A23187 ionophore stimulation. An in situ fluo-3 calibration method (using A23187 and metabolic poisons in Ca2+ /EGTA buffers with known free calcium concentrations) indicated a resting [Ca2+]i in lymphocytes of 103 +/- 23 nM (mean +/- S.D.) but with significant differences between the [Ca2+]i in B cells and in all of the T-cell subsets (CD4+Thy-1+, CD4+Thy-1-, and CD8+T cells). Both the B cells and the T-cell subsets showed an increase of fluo-3 fluorescence upon in vitro stimulation with ConA or PHA, but the calcium mobilization following lectin stimulation was time delayed in all T-cell subsets. Only the T cells, including the CD4+Thy-1- subset, responded to anti-CD3 MoAbs. The percentage of responding cells upon stimulation with ConA was higher in T cells than in B cells. By contrast, PHA gave a higher response in B cells. After stimulation with different mitogens, [Ca2+]i increased in both CD4+ and CD8+ T-cell subsets. However, the percentage of responding cells was far higher in the CD4+Thy-1+ subset than in the CD4+Thy-1- or the CD8+T-cell subsets. The stimulation with A23187 ionophore induced a higher calcium response in B cells than in T cells. Interestingly, it also induced greater Ca2+ mobilization in CD4+ than in CD8+T cells. These results demonstrate the potential use of fluo-3 simultaneously with three fluorescein (FITC)-compatible fluorochromes. This technique may be useful for investigating the role of the CD4+Thy-1-T cells, a rare subset that is abnormally expanded in a murine acquired immunodeficiency syndrome (murine AIDS).


Novel data on the innervation of the lophophore in adult phoronids (Lophophorata, Phoronida).
The structure of the lophophore nervous system may help clarify the status of the clade Lophophorata, whose monophyly is debated. In the current study, antibody labeling and confocal laser scanning microscopy revealed previously undescribed main nerve elements in the lophophore in adult phoronids: Phoronis australis and Phoronopsis harmeri. In both species, the nervous system includes a dorsal ganglion, a tentacle nerve ring, an inner nerve ring, intertentacular groups of perikarya, and tentacle nerves. The dorsal ganglion and tentacle nerve ring contain many serotonin-like immunoreactive perikarya of different sizes. The inner nerve ring is described for the first time in adult phoronids with complex lophophore. It contains a thin bundle of serotonin-like immunoreactive neurites. The tentacles possess abfrontal, frontal, and laterofrontal nerves. The abfrontal nerves originate from the tentacle nerve ring; the frontal tentacle nerves extend from the inner nerve ring in P. harmeri and from the intertentacular frontal nerves in P. australis. The intertentacular groups of perikarya are found in phoronids for the first time. These small nerve centers connect with neither the tentacle nerve ring nor the inner nerve ring, giving rise to the laterofrontal tentacle nerves. The discovery of the inner nerve ring in adult phoronids makes the architecture of the lophophore nervous system similar in all lophophorates and thereby supports the monophyly of this group. The presence of intertentacular nerves, perikarya, and groups of perikarya is a typical feature of the nervous system in lophophorate presumably coordinating movements of the tentacles and thereby increasing the efficiency of lophophore functioning.