Methylation status of CpG islands in the promoter region of genes differentially expressed in colonic mucosa from adenoma patients and controls in response to altered vegetable intake.
Vegetables may protect against colorectal cancer (CRC) via changes in gene expression involved in anticarcinogenic mechanisms. There is considerable evidence that aberrant DNA methylation plays an important role in carcinogenesis. Furthermore, DNA methylation can be affected by dietary components. Therefore, in the present study, we investigated the DNA methylation status of CpG dinucleotides within the promoter region of the four genes protein kinase C b 1, ornithine decarboxylase 1, fos proto-oncogene and 5,10-methylenetetrahydrofolate reductase in the colon of female sporadic adenoma patients and healthy controls. These genes were chosen because their expression was modulated in response to altered vegetable intake, they are functionally relevant for CRC; they have CpG islands in their promoter region, and a methylation-specific restriction enzyme is available to permit quantitative assay. No significant differences in extent of methylation in colon DNA were detected for any of the four genes in both adenoma polyp patients and healthy controls after altering vegetable intake. Interestingly, before the intervention, ornithine decarboxylase 1 promoter methylation was lower in the colonic mucosa of the adenoma polyp patients when compared with healthy control subjects, which may explain the increased ornithine decarboxylase 1 activity in CRC reported in the literature. In conclusion, we found no evidence that changes in promoter methylation were responsible for differences in expression of four genes in the human colonic mucosa in response to altered vegetable intake. The mechanism(s) responsible for this altered gene expression and, indeed, potential effects on methylation of other genes remain to be determined.


Coupling to modes of a near-confocal optical resonator using a digital light modulator.
Digital micromirror devices (DMD) provide a robust platform with which to implement digital holography, in principle providing the means to rapidly generate propagating transverse electromagnetic fields with arbitrary mode profiles at visible and IR wavelengths. We use a DMD to probe a Fabry-Pérot cavity in single-mode and near-degenerate confocal configurations. Pumping arbitrary modes of the cavity is possible with excellent specificity by virtue of the spatial overlap between the incident light field and the cavity mode.


Development of the Italian Version of the Neck Disability Index: cross-cultural adaptation, factor analysis, reliability, validity, and sensitivity to change.
Evaluation of the psychometric properties of a translated and culturally adapted questionnaire. Translating, culturally adapting, and validating the Italian version of the Neck Disability Index (NDI-I) to allow its use with Italian-speaking patients with neck pain (NP). More attention is being given to standardized outcome measures to improve interventions for NP. A translated form of the NDI has never been validated in Italian patients with NP. The NDI-I was developed by forward-backward translation, a final review by an expert committee, and a test of the prefinal version to establish its correspondence with the original English version. The psychometric testing included factor analysis, reliability by internal consistency (Cronbach α) and test-retest reliability (intraclass coefficient correlation), construct validity by comparing NDI-I with the Neck Pain and Disability Scale, a numerical rating scale, the Hospital Anxiety and Depression Scale, and the 36-Item Short Form Health Survey (Spearman correlation), and sensitivity to change by calculating the smallest detectable change. The questionnaire was administered to 101 subjects with chronic NP and proved to be acceptable. Factor analysis revealed a 2-factor 10-item solution (explained variance: 56%). The questionnaire showed good internal consistency (α = 0.842) and test-retest reliability (intraclass coefficient correlation = 0.846). Construct validity showed a good correlation with Neck Pain and Disability Scale (ρ = 0.687), moderate correlations with the numerical rating scale (ρ = 0.545), and Hospital Anxiety and Depression Scale (ρ = 0.422 for the Anxiety score and ρ = 0.546 for the Depression score), and poor correlations with the 36-Item Short Form Health Survey subscales (ρ = 0.066 to -0.286). The psychometric analyses of the subscales and total scale were similar. The smallest detectable change of the NDI-I was 3. The NDI was successfully translated into Italian and proved to have a good factorial structure and psychometric properties that replicated the results of other versions. Its use is recommended for research purposes.


Hypoglycemia and the origin of hypoxia-induced reduction in human fetal growth.
The most well known reproductive consequence of residence at high altitude (HA >2700 m) is reduction in fetal growth. Reduced fetoplacental oxygenation is an underlying cause of pregnancy pathologies, including intrauterine growth restriction and preeclampsia, which are more common at HA. Therefore, altitude is a natural experimental model to study the etiology of pregnancy pathophysiologies. We have shown that the proximate cause of decreased fetal growth is not reduced oxygen availability, delivery, or consumption. We therefore asked whether glucose, the primary substrate for fetal growth, might be decreased and/or whether altered fetoplacental glucose metabolism might account for reduced fetal growth at HA. Doppler and ultrasound were used to measure maternal uterine and fetal umbilical blood flows in 69 and 58 residents of 400 vs 3600 m. Arterial and venous blood samples from mother and fetus were collected at elective cesarean delivery and analyzed for glucose, lactate and insulin. Maternal delivery and fetal uptakes for oxygen and glucose were calculated. The maternal arterial - venous glucose concentration difference was greater at HA. However, umbilical venous and arterial glucose concentrations were markedly decreased, resulting in lower glucose delivery at 3600 m. Fetal glucose consumption was reduced by >28%, but strongly correlated with glucose delivery, highlighting the relevance of glucose concentration to fetal uptake. At altitude, fetal lactate levels were increased, insulin concentrations decreased, and the expression of GLUT1 glucose transporter protein in the placental basal membrane was reduced. Our results support that preferential anaerobic consumption of glucose by the placenta at high altitude spares oxygen for fetal use, but limits glucose availability for fetal growth. Thus reduced fetal growth at high altitude is associated with fetal hypoglycemia, hypoinsulinemia and a trend towards lactacidemia. Our data support that placentally-mediated reduction in glucose transport is an initiating factor for reduced fetal growth under conditions of chronic hypoxemia.


Cooperative elastic fluctuations provide tuning of the metal-insulator transition.
Metal-to-insulator transitions1 driven by strong electronic correlations occur frequently in condensed matter systems, and are associated with remarkable collective phenomena in solids, including superconductivity and magnetism. Tuning and control of the transition holds the promise of low-power, ultrafast electronics2, but the relative roles of doping, chemistry, elastic strain and other applied fields have made systematic understanding of such transitions difficult. Here we show that existing data3-5 on the tuning of metal-to-insulator transitions in perovskite transition-metal oxides through ionic size effects provides evidence of large systematic effects on the phase transition owing to dynamical fluctuations of the elastic strain, which have usually been neglected6. We illustrate this using a simple yet quantitative statistical mechanical calculation in a model that incorporates cooperative lattice distortions coupled to the electronic degrees of freedom. We reproduce the observed dependence of the transition temperature on the cation radius in the well studied manganite7 and nickelate8 materials. Because elastic couplings are generally strong, we anticipate that these conclusions will generalize to all metal-to-insulator transitions that couple to a change in lattice symmetry.


Development and validation of an IA-LC/MS method to quantitate active and total B-type natriuretic peptide in human plasma.
Patients with elevated levels of B-type natriuretic peptide (BNP) and/or NT-proBNP as measured by clinical tests have an elevated risk of heart failure (HF). Despite utility in large clinical studies, both assays are plagued by large biological variability and specificity issues. To address these concerns and further investigate BNP in the HF setting, we developed an LC/MS assay to characterize the ratio of active to total BNP. We have developed and validated a novel immunoaffinity LC/MS assay to measure BNP-derived fragments, as well as 'total BNP' in human plasma. The ratio of active BNP1-32 to total BNP in 11 HF subjects was found to be <8%, and the sum of detectable BNP fragments contributed approximately 20% of total BNP. We developed an assay with the specificity to measure the active form of BNP, which may aid in the accurate diagnosis and better management of HF.


Virus-free induction of pluripotency and subsequent excision of reprogramming factors.
Reprogramming of somatic cells to pluripotency, thereby creating induced pluripotent stem (iPS) cells, promises to transform regenerative medicine. Most instances of direct reprogramming have been achieved by forced expression of defined factors using multiple viral vectors. However, such iPS cells contain a large number of viral vector integrations, any one of which could cause unpredictable genetic dysfunction. Whereas c-Myc is dispensable for reprogramming, complete elimination of the other exogenous factors is also desired because ectopic expression of either Oct4 (also known as Pou5f1) or Klf4 can induce dysplasia. Two transient transfection-reprogramming methods have been published to address this issue. However, the efficiency of both approaches is extremely low, and neither has been applied successfully to human cells so far. Here we show that non-viral transfection of a single multiprotein expression vector, which comprises the coding sequences of c-Myc, Klf4, Oct4 and Sox2 linked with 2A peptides, can reprogram both mouse and human fibroblasts. Moreover, the transgene can be removed once reprogramming has been achieved. iPS cells produced with this non-viral vector show robust expression of pluripotency markers, indicating a reprogrammed state confirmed functionally by in vitro differentiation assays and formation of adult chimaeric mice. When the single-vector reprogramming system was combined with a piggyBac transposon, we succeeded in establishing reprogrammed human cell lines from embryonic fibroblasts with robust expression of pluripotency markers. This system minimizes genome modification in iPS cells and enables complete elimination of exogenous reprogramming factors, efficiently providing iPS cells that are applicable to regenerative medicine, drug screening and the establishment of disease models.


G protein activation by serotonin type 4 receptor dimers: evidence that turning on two protomers is more efficient.
The discovery that class C G protein-coupled receptors (GPCRs) function as obligatory dimeric entities has generated major interest in GPCR oligomerization. Oligomerization now appears to be a common feature among all GPCR classes. However, the functional significance of this process remains unclear because, in vitro, some monomeric GPCRs, such as rhodopsin and β(2)-adrenergic receptors, activate G proteins. By using wild type and mutant serotonin type 4 receptors (5-HT(4)Rs) (including a 5-HT(4)-RASSL) expressed in COS-7 cells as models of class A GPCRs, we show that activation of one protomer in a dimer was sufficient to stimulate G proteins. However, coupling efficiency was 2 times higher when both protomers were activated. Expression of combinations of 5-HT(4), in which both protomers were able to bind to agonists but only one could couple to G proteins, suggested that upon agonist occupancy, protomers did not independently couple to G proteins but rather that only one G protein was activated. Coupling of a single heterotrimeric G(s) protein to a receptor dimer was further confirmed in vitro, using the purified recombinant WT RASSL 5-HT(4)R obligatory heterodimer. These results, together with previous findings, demonstrate that, differently from class C GPCR dimers, class A GPCR dimers have pleiotropic activation mechanisms.


Linkage of inflammation and oxidative stress via release of glutathionylated peroxiredoxin-2, which acts as a danger signal.
The mechanism by which oxidative stress induces inflammation and vice versa is unclear but is of great importance, being apparently linked to many chronic inflammatory diseases. We show here that inflammatory stimuli induce release of oxidized peroxiredoxin-2 (PRDX2), a ubiquitous redox-active intracellular enzyme. Once released, the extracellular PRDX2 acts as a redox-dependent inflammatory mediator, triggering macrophages to produce and release TNF-α. The oxidative coupling of glutathione (GSH) to PRDX2 cysteine residues (i.e., protein glutathionylation) occurs before or during PRDX2 release, a process central to the regulation of immunity. We identified PRDX2 among the glutathionylated proteins released in vitro by LPS-stimulated macrophages using mass spectrometry proteomic methods. Consistent with being part of an inflammatory cascade, we find that PRDX2 then induces TNF-α release. Unlike classical inflammatory cytokines, PRDX2 release does not reflect LPS-mediated induction of mRNA or protein synthesis; instead, PRDX2 is constitutively present in macrophages, mainly in the reduced form, and is released in the oxidized form on LPS stimulation. Release of PRDX2 is also observed in human embryonic kidney cells treated with TNF-α. Importantly, the PRDX2 substrate thioredoxin (TRX) is also released along with PRDX2, enabling an oxidative cascade that can alter the -SH status of surface proteins and thereby facilitate activation via cytokine and Toll-like receptors. Thus, our findings suggest a model in which the release of PRDX2 and TRX from macrophages can modify the redox status of cell surface receptors and enable induction of inflammatory responses. This pathway warrants further exploration as a potential novel therapeutic target for chronic inflammatory diseases.


Prospective study of gut hormone and metabolic changes after adjustable gastric banding and Roux-en-Y gastric bypass.
The objective of this study was to quantify hormones that regulate energy and glucose homeostasis to establish possible mechanisms for the greater efficacy of Roux-en-Y gastric bypass (RYGB) compared with laparoscopic adjustable gastric banding (LAGB) in achieving weight loss and improved insulin sensitivity. Longitudinal study of patients undergoing LAGB (n=15) and RYGB (n=28) who were studied before surgery and at 2, 12, 26 and 52 weeks afterwards. Fasting blood samples were drawn at each visit. Postprandial blood samples were also obtained before surgery and at 26 and 52 weeks. Samples were assayed for peptide YY (PYY), ghrelin, glucagon-like peptide-1 (GLP-1), glucose, insulin, leptin, thyrotropic hormone, free T(4) and free T(3). At 1 year there was greater weight loss in RYGB compared with LAGB patients (30 vs 15%), but final body mass index was similar (34 vs 33 kg m(-2)). At week 52, area under the curve (AUC) for PYY in RYGB subjects was greater than LAGB (P<0.01). GLP-1 levels at 30 min after meal were threefold greater after RYGB compared with LAGB (P<0.001). Conversely, ghrelin AUC increased after LAGB at week 52 (P<0.05) but tended to decrease after RYGB. Fasting glucose, insulin, and leptin and homeostasis model of assessment (HOMA-IR) decreased in both groups over time but were significantly lower at week 52 after RYGB compared with LAGB. The change in leptin correlated significantly with weight loss in LAGB (r=0.86) and RYGB (r=0.77), however, HOMA-IR correlated significantly with weight loss only in LAGB (r=0.78), and not RYGB (r=0.15). There was a significant decrease in free T(3) (P<0.01) after RYGB. Differences in levels of gut hormones may play a role in promoting greater weight loss and insulin sensitivity after RYGB compared with LAGB, however, weight loss may be limited by decreases in free T(3) and leptin.


Single-cell phenomics reveals intra-species variation of phenotypic noise in yeast.
Most quantitative measures of phenotypic traits represent macroscopic contributions of large numbers of cells. Yet, cells of a tissue do not behave similarly, and molecular studies on several organisms have shown that regulations can be highly stochastic, sometimes generating diversified cellular phenotypes within tissues. Phenotypic noise, defined here as trait variability among isogenic cells of the same type and sharing a common environment, has therefore received a lot of attention. Given the potential fitness advantage provided by phenotypic noise in fluctuating environments, the possibility that it is directly subjected to evolutionary selection is being considered. For selection to act, phenotypic noise must differ between contemporary genotypes. Whether this is the case or not remains, however, unclear because phenotypic noise has very rarely been quantified in natural populations. Using automated image analysis, we describe here the phenotypic diversity of S. cerevisiae morphology at single-cell resolution. We profiled hundreds of quantitative traits in more than 1,000 cells of 37 natural strains, which represent various geographical and ecological origins of the species. We observed abundant trait variation between strains, with no correlation with their ecological origin or population history. Phenotypic noise strongly depended on the strain background. Noise variation was largely trait-specific (specific strains showing elevated noise for subset of traits) but also global (a few strains displaying elevated noise for many unrelated traits). Our results demonstrate that phenotypic noise does differ quantitatively between natural populations. This supports the possibility that, if noise is adaptive, microevolution may tune it in the wild. This tuning may happen on specific traits or by varying the degree of global phenotypic buffering.


Excretion of administered and endogenous photobilirubins in the bile of the jaundice gunn rat.
Radiolabeled photobilirubins, prepared in vitro by anaerobic illumination of [34C]bilirubin, were injected intravenously into homozygous jaundiced Gunn rats with an external bile fistula. With the animals kept in darkness, the labeled photobilirubins were excreted rapidly in bile. Photobilirubins IA and IB were excreted primarily as unconjugated bilirubin, whereas photobilirubin II was excreted primarily as photobilirubin II and not converted into bilirubin. Bile of Gunn rats given no exogenous pigments, but undergoing phototherapy, contained a large proportion of photobilirubin II and, if collected in liquid nitrogen, traces of photobilirubins I; neither was found in bile when these rats were kept in the dark. Because there is prior evidence that these rats were kept in the dark. Because there is prior evidence that these photobilirubins are isomers of bilirubin, these experiments indicate that the major mechanism of phototherapy is photoisomerization of bilirubin. Photobilirubin II is the unidentified major photoderivative described previously, whereas formation of photobilirubins IA and IB, and their reversion to bilirubin-IXalpha, account for the remarkably increased output of the parent pigment during phototherapy.


Diagnostic Classification and Biomarker Identification of Alzheimer's Disease with Random Forest Algorithm.
Random Forest (RF) is a bagging ensemble model and has many important advantages, such as robustness to noise, an effective structure for complex multimodal data and parallel computing, and also provides important features that help investigate biomarkers. Despite these benefits, RF is not used actively to predict Alzheimer's disease (AD) with brain MRIs. Recent studies have reported RF's effectiveness in predicting AD, but the test sample sizes were too small to draw any solid conclusions. Thus, it is timely to compare RF with other learning model methods, including deep learning, particularly with large amounts of data. In this study, we tested RF and various machine learning models with regional volumes from 2250 brain MRIs: 687 normal controls (NC), 1094 mild cognitive impairment (MCI), and 469 AD that ADNI (Alzheimer's Disease Neuroimaging Initiative database) provided. Three types of features sets (63, 29, and 22 features) were selected, and classification accuracies were computed with RF, Support vector machine (SVM), Multi-layer perceptron (MLP), and Convolutional neural network (CNN). As a result, RF, MLP, and CNN showed high performances of 90.2%, 89.6%, and 90.5% with 63 features. Interestingly, when 22 features were used, RF showed the smallest decrease in accuracy, -3.8%, and the standard deviation did not change significantly, while MLP and CNN yielded decreases in accuracy of -6.8% and -4.5% with changes in the standard deviation from 3.3% to 4.0% for MLP and 2.1% to 7.0% for CNN, indicating that RF predicts AD more reliably with fewer features. In addition, we investigated the importance of the features that RF provides, and identified the hippocampus, amygdala, and inferior lateral ventricle as the major contributors in classifying NC, MCI, and AD. On average, AD showed smaller hippocampus and amygdala volumes and a larger volume of inferior lateral ventricle than those of MCI and NC.


The influence of emotions on inhibitory functioning in borderline personality disorder.
Borderline personality disorder (BPD) is characterized by an emotionally unstable and impulsive cognitive and behavioral style. Inhibitory dysfunction has been hypothesized as playing a crucial role in BPD psychopathology. This study aimed to systematically investigate differential inhibitory functions in patients with BPD as compared to healthy controls, and to investigate their expected impairment in the context of aversive emotions by comparing performances in neuropsychological tasks that present both neutral and emotional material. Unmedicated female patients with BPD (n=28) were compared with age-matched healthy female controls (n=30) in the following tasks: the emotional Stroop test (inhibition of interference), directed forgetting (intentional, resource-dependent inhibition), and an emotional variant of the negative priming task (automatic, resource-independent inhibition). In comparison with the controls, the BPD patients showed reduced inhibition of negative material in the directed forgetting task and in the negative priming task. No effect was found in the emotional Stroop test. Significant correlations with current affect as well as trait anxiety and anger (but not impulsiveness) were found in the BPD group specifically for negative stimuli, while no such correlations were found in the control group. In addition to inhibitory deficiencies, BPD patients had difficulties remembering positive words in the directed forgetting task. Our data suggest that individuals with BPD have difficulties in actively suppressing irrelevant information when it is of an aversive nature. Inhibitory dysfunction appears to be closely related to state and trait variables of unstable affect, but not to self-reported impulsiveness.


Isobactins: O-acyl isopeptide prodrugs of teixobactin and teixobactin derivatives.
The antibiotic teixobactin is a promising drug candidate against drug-resistant pathogens, such as MRSA and VRE, but forms insoluble gels that may limit intravenous administration. O-Acyl isopeptide prodrug analogues of teixobactin circumvent the problem of gel formation while retaining antibiotic activity. The teixobactin prodrug analogues contain ester linkages between Ile6 and Ser7, Ile2 and Ser3, or between both Ile6 and Ser7 and Ile2 and Ser3. Upon exposure to physiological pH, the prodrug analogues undergo clean conversion to the corresponding amides, with half-lives between 13 and 115 min. Prodrug analogues containing lysine, arginine, or leucine at position 10 exhibit good antibiotic activity against a variety of Gram-positive bacteria while exhibiting little or no cytotoxicity or hemolytic activity. Because O-acyl isopeptide prodrug analogues of teixobactin exhibit clean conversion to the corresponding teixobactin analogues with reduced propensity to form gels, it is anticipated that teixobactin prodrugs will be superior to teixobactin as drug candidates.


Reduced neural synchronization of gamma-band MEG oscillations in first-degree relatives of children with autism.
Gamma-band oscillations recorded from human electrophysiological recordings, which may be associated with perceptual binding and neuronal connectivity, have been shown to be altered in people with autism. Transient auditory gamma-band responses, however, have not yet been investigated in autism or in the first-degree relatives of persons with the autism. We measured transient evoked and induced magnetic gamma-band power and inter-trial phase-locking consistency in the magnetoencephalographic recordings of 16 parents of children with autism, 11 adults with autism and 16 control participants. Source space projection was used to separate left and right hemisphere transient gamma-band measures of power and phase-locking. Induced gamma-power at 40 Hz was significantly higher in the parent and autism groups than in controls, while evoked gamma-band power was reduced compared to controls. The phase-locking factor, a measure of phase consistency of neuronal responses with external stimuli, was significantly lower in the subjects with autism and the autism parent group, potentially explaining the difference between the evoked and induced power results. These findings, especially in first degree relatives, suggest that gamma-band phase consistency and changes in induced versus induced power may be potentially useful endophenotypes for autism, particularly given emerging molecular mechanisms concerning the generation of gamma-band signals.


Shape Effect of Surface Defects on Nanohardness by Quasicontinuum Method.
Nanoindentation on a platinum thin film with surface defects in a rectangular shape and triangular shape was simulated using the quasicontinuum method to study the shape effect of surface defects on nanohardness. The results show that the nanohardness of thin film with triangular defects is basically larger than those with rectangular defects, which is closely related to the height of the surface defects at the boundary near to the indenter. Moreover, the triangular defect might have an enhancement effect on nanohardness by a certain size of the defects and the boundary orientation of the defect, where such an enhancement effect increases as the defect grows. Furthermore, the nanohardness decreases when the defect is folded from wide to narrow in the same atom cavity, and particularly expresses a more obvious drop when the height of the defects increases. In addition, larger sizes of the rectangular defect induce more reduction in nanohardness, while the nanohardness of the triangular surface defect is sensitive to the periodic arrangement of atoms changed by the boundary orientation of the defect, which is well explained and demonstrated by the calculation formula theory of necessary load for dislocation emission.


Genomic surveillance of Rift Valley fever virus: from sequencing to lineage assignment.
Genetic evolution of Rift Valley fever virus (RVFV) in Africa has been shaped mainly by environmental changes such as abnormal rainfall patterns and climate change that has occurred over the last few decades. These gradual environmental changes are believed to have effected gene migration from macro (geographical) to micro (reassortment) levels. Presently, 15 lineages of RVFV have been identified to be circulating within the Sub-Saharan Africa. International trade in livestock and movement of mosquitoes are thought to be responsible for the outbreaks occurring outside endemic or enzootic regions. Virus spillover events contribute to outbreaks as was demonstrated by the largest epidemic of 1977 in Egypt. Genomic surveillance of the virus evolution is crucial in developing intervention strategies. Therefore, we have developed a computational tool for rapidly classifying and assigning lineages of the RVFV isolates. The computational method is presented both as a command line tool and a web application hosted at https://www.genomedetective.com/app/typingtool/rvfv/ . Validation of the tool has been performed on a large dataset using glycoprotein gene (Gn) and whole genome sequences of the Large (L), Medium (M) and Small (S) segments of the RVFV retrieved from the National Center for Biotechnology Information (NCBI) GenBank database. Using the Gn nucleotide sequences, the RVFV typing tool was able to correctly classify all 234 RVFV sequences at species level with 100% specificity, sensitivity and accuracy. All the sequences in lineages A (n = 10), B (n = 1), C (n = 88), D (n = 1), E (n = 3), F (n = 2), G (n = 2), H (n = 105), I (n = 2), J (n = 1), K (n = 4), L (n = 8), M (n = 1), N (n = 5) and O (n = 1) were also correctly classified at phylogenetic level. Lineage assignment using whole RVFV genome sequences (L, M and S-segments) did not achieve 100% specificity, sensitivity and accuracy for all the sequences analyzed. We further tested our tool using genomic data that we generated by sequencing 5 samples collected following a recent RVF outbreak in Kenya. All the 5 samples were assigned lineage C by both the partial (Gn) and whole genome sequence classifiers. The tool is useful in tracing the origin of outbreaks and supporting surveillance efforts.Availability: https://github.com/ajodeh-juma/rvfvtyping.


Cutibacterium acnes (formerly Propionibacterium acnes) isolated from prosthetic joint infections is less susceptible to oxacillin than to benzylpenicillin.
Introduction: The frequency of prosthetic joint infections (PJIs) due to Cutibacterium acnes (formerly Propionibacterium acnes) is increasing, especially shoulder PJIs. The recommended antibiotic prophylaxis for hip and knee arthroplasties is beta-lactam antibiotics, predominantly cephalosporins. However, for example in Sweden, isoxazolyl-penicillin cloxacillin is used. No specific recommendations for shoulder arthroplasties are available. The aim of the present study was to determine the minimum inhibitory concentration (MIC) values for different antibiotics for C. acnes; and, more specifically, to compare the MIC values for benzylpenicillin and oxacillin. Materials and methods: Minimum inhibitory concentration (MIC) values for nine different antibiotic agents were obtained by gradient test (Etest) using strains of C. acnes (n= 57) isolated from PJIs from shoulders (n=31), hips (n=21), and knees (n=5). Results: All isolates had low MIC values for most of the tested antibiotic agents, and showed a wild type MIC distribution. The exception was clindamycin with 9% of the isolates displaying decreased susceptibility. The MIC values obtained for benzylpenicillin were significantly lower than the MIC values for isoxazolyl-penicillin (oxacillin). Conclusion: These in vitro results indicate that benzylpenicillin might be a more effective prophylactic treatment to prevent shoulder PJIs caused by C. acnes. However, further studies on the subject are needed, and the effectiveness of the prophylactic treatment should be evaluated using randomized controlled studies and/or register-based studies.


A mixed-methods sequential explanatory design comparison between COVID-19 infection control guidelines' applicability and their protective value as perceived by Israeli healthcare workers, and healthcare executives' response.
Healthcare workers (HCWs) are on the front line of the COVID-19 outbreak, and their constant exposure to infected patients and contaminated surfaces puts them at risk of acquiring and transmitting the infection. Therefore, they must employ protective measures. In practice, HCWs in Israel were not fully prepared for this sudden COVID-19 outbreak. This research aimed to identify and compare: (1) Israeli HCWs' perceptions regarding the official COVID-19 guidelines' applicability and their protective value, and (2) HCWs executives' response to HWCs' concern regarding personal protective equipment (PPE) shortage. A mixed-methods sequential explanatory design consists of: (1) An online survey of 242 HCWs about the application of the guidelines and PPE, and (2) Personal interviews of 15 HCWs executives regarding PPE shortage and the measures they are taking to address it. A significant difference between the perceived applicability and protective value was found for most of the guidelines. Some of the guidelines were perceived as more applicable than protective (hand hygiene, signage at entrance, alcohol rub sanitizers at entrance, and mask for contact with symptomatic patients). Other were perceived as less applicable than protective (prohibited gathering of over 10 people, maintaining a distance of 2 m', and remote services). HCWs need the support of the healthcare authorities not only to provide missing equipment, but also to communicate the risk to them. Conveying the information with full transparency, while addressing the uncertainty element and engaging the HCWs in evaluating the guidelines, are critical for establishing trust.


3D printed TCP-based scaffold incorporating VEGF-loaded PLGA microspheres for craniofacial tissue engineering.
Vascularization is a critical process during bone regeneration/repair and the lack of tissue vascularization is recognized as a major challenge in applying bone tissue engineering methods for cranial and maxillofacial surgeries. The aim of our study is to fabricate a vascular endothelial growth factor (VEGF)-loaded gelatin/alginate/β-TCP composite scaffold by 3D printing method using a computer-assisted design (CAD) model. The paste, composed of (VEGF-loaded PLGA)-containing gelatin/alginate/β-TCP in water, was loaded into standard Nordson cartridges and promptly employed for printing the scaffolds. Rheological characterization of various gelatin/alginate/β-TCP formulations led to an optimized paste as a printable bioink at room temperature. The in vitro release kinetics of the loaded VEGF revealed that the designed scaffolds fulfill the bioavailability of VEGF required for vascularization in the early stages of tissue regeneration. The results were confirmed by two times increment of proliferation of human umbilical vein endothelial cells (HUVECs) seeded on the scaffolds after 10 days. The compressive modulus of the scaffolds, 98±11MPa, was found to be in the range of cancellous bone suggesting their potential application for craniofacial tissue engineering. Osteoblast culture on the scaffolds showed that the construct supports cell viability, adhesion and proliferation. It was found that the ALP activity increased over 50% using VEGF-loaded scaffolds after 2 weeks of culture. The 3D printed gelatin/alginate/β-TCP scaffold with slow releasing of VEGF can be considered as a potential candidate for regeneration of craniofacial defects.


Deep Learning-Based Framework for In Vivo Identification of Glioblastoma Tumor using Hyperspectral Images of Human Brain.
The main goal of brain cancer surgery is to perform an accurate resection of the tumor, preserving as much normal brain tissue as possible for the patient. The development of a non-contact and label-free method to provide reliable support for tumor resection in real-time during neurosurgical procedures is a current clinical need. Hyperspectral imaging is a non-contact, non-ionizing, and label-free imaging modality that can assist surgeons during this challenging task without using any contrast agent. In this work, we present a deep learning-based framework for processing hyperspectral images of in vivo human brain tissue. The proposed framework was evaluated by our human image database, which includes 26 in vivo hyperspectral cubes from 16 different patients, among which 258,810 pixels were labeled. The proposed framework is able to generate a thematic map where the parenchymal area of the brain is delineated and the location of the tumor is identified, providing guidance to the operating surgeon for a successful and precise tumor resection. The deep learning pipeline achieves an overall accuracy of 80% for multiclass classification, improving the results obtained with traditional support vector machine (SVM)-based approaches. In addition, an aid visualization system is presented, where the final thematic map can be adjusted by the operating surgeon to find the optimal classification threshold for the current situation during the surgical procedure.


Incidence of pulmonary infection due to Nocardia species in Nigeria.
Pulmonary infection due to Nocardia sp. was investigated in 600 patients of various bronchopulmonary disorders by direct microscopy of sputum samples and cultures on appropriate media. Pathogenic Nocardia species were isolated from 25 patients, giving 4.1% incidence of infection. The species of Nocardia represented were N. asteroides (21 isolates), N. otitidis-caviarum and N. brasiliensis (2 isolates each). Paraffin baiting technique proved to be more efficacious than culturing on glucose nutrient agar.


Biocompatible Coatings from Smart Biopolymer Nanoparticles for Enzymatically Induced Drug Release.
Nanoparticles can be used as a smart drug delivery system, when they release the drug only upon degradation by specific enzymes. A method to create such responsive materials is the formation of hydrogel nanoparticles, which have enzymatically degradable crosslinkers. Such hydrogel nanoparticles were prepared by ionotropic gelation sodium alginate with lysine-rich peptide sequences-either α-poly-L-lysine (PLL) or the aggrecanase-labile sequence KKKK-GRD-ARGSV↓NITEGE-DRG-KKKK. The nanoparticle suspensions obtained were analyzed by means of dynamic light scattering and nanoparticle tracking analysis. Degradation experiments carried out with the nanoparticles in suspension revealed enzyme-induced lability. Drugs present in the polymer solution during the ionotropic gelation can be encapsulated in the nanoparticles. Drug loading was investigated for interferon-β (IFN-β) as a model, using a bioluminescence assay with MX2Luc2 cells. The encapsulation efficiency for IFN-β was found to be approximately 25%. The nanoparticles suspension can be used to spray-coat titanium alloys (Ti-6Al-4V) as a common implant material. The coatings were proven by ellipsometry, reflection-absorption infrared spectroscopy, and X-ray photoelectron spectroscopy. An enzyme-responsive decrease in layer thickness is observed due to the degradation of the coatings. The Alg/peptide coatings were cytocompatible for human gingival fibroblasts (HGFIB), which was investigated by CellTiterBlue and lactate dehydrogenase (LDH) assay. However, HGFIBs showed poor adhesion and proliferation on the Alg/peptide coatings, but these could be improved by modification of the alginate with a RGD-peptide sequence. The smart drug release system presented can be further tailored to have the right release kinetics and cell adhesion properties.


Multiple common susceptibility variants near BMP pathway loci GREM1, BMP4, and BMP2 explain part of the missing heritability of colorectal cancer.
Genome-wide association studies (GWAS) have identified 14 tagging single nucleotide polymorphisms (tagSNPs) that are associated with the risk of colorectal cancer (CRC), and several of these tagSNPs are near bone morphogenetic protein (BMP) pathway loci. The penalty of multiple testing implicit in GWAS increases the attraction of complementary approaches for disease gene discovery, including candidate gene- or pathway-based analyses. The strongest candidate loci for additional predisposition SNPs are arguably those already known both to have functional relevance and to be involved in disease risk. To investigate this proposition, we searched for novel CRC susceptibility variants close to the BMP pathway genes GREM1 (15q13.3), BMP4 (14q22.2), and BMP2 (20p12.3) using sample sets totalling 24,910 CRC cases and 26,275 controls. We identified new, independent CRC predisposition SNPs close to BMP4 (rs1957636, P = 3.93×10(-10)) and BMP2 (rs4813802, P = 4.65×10(-11)). Near GREM1, we found using fine-mapping that the previously-identified association between tagSNP rs4779584 and CRC actually resulted from two independent signals represented by rs16969681 (P = 5.33×10(-8)) and rs11632715 (P = 2.30×10(-10)). As low-penetrance predisposition variants become harder to identify-owing to small effect sizes and/or low risk allele frequencies-approaches based on informed candidate gene selection may become increasingly attractive. Our data emphasise that genetic fine-mapping studies can deconvolute associations that have arisen owing to independent correlation of a tagSNP with more than one functional SNP, thus explaining some of the apparently missing heritability of common diseases.


Management of simple and retained hemothorax: A practice management guideline from the Eastern Association for the Surgery of Trauma.
Traumatic hemothorax poses diagnostic and therapeutic challenges both acutely and chronically. A working group of the Eastern Association for the Surgery of Trauma convened to formulate a practice management guideline for traumatic hemothorax. We formulated four questions: whether tube thoracostomy vs observation be performed, should pigtail catheter versus thoracostomy tube be placed to drain hemothorax, should thrombolytic therapy be attempted versus immediate thoracoscopic assisted drainage (VATS) in retained hemothorax (rHTX), and should early VATS (≤4 days) versus late VATS (>4 days) be performed? A systematic review was undertaken from articles identified in multiple databases. A total of 6391 articles were identified, 14 were selected for guideline construction. Most articles were retrospective with very low-quality evidence. We performed meta-analysis for some of the outcomes for three of the questions. For traumatic hemothorax we conditionally recommend pigtail catheters, in hemodynamically stable patients. In patients with rHTX, we conditionally recommend VATS rather than attempting thrombolytic therapy and recommend that it should be performed early (≤4 days).


THE PROGRESSION TO CARCINOMA OF VIRUS-INDUCED RABBIT PAPILLOMAS (SHOPE).
The papillomas induced in domestic rabbits with virus procured from cottontails undergo progressive changes in the direction of malignancy when they grow vigorously. From the beginning they exhibit the traits whereby tumors are characterized, and they have malignant potentialities. In seven animals of a group of ten carrying papillomas for more than 200 days, cancer has developed, and in an eighth a tumor of problematic malignancy has arisen. One of the remaining two rabbits died early in the cancer period, and the papillomas of the other eventually retrogressed. Ten cottontails with induced growths of much longer duration have not developed cancer. The malignant tumors have all been acanthomatous in type, and have arisen directly from the papillomas by graded, continuous alterations. These have often gone further after malignancy has been attained, and have eventuated in great anaplasia. Metastasis has been frequent, and transplantation to another host has proved successful. Individual growths have occurred expressive of each stage of the transformation to cancer, as if through a stabilization at this stage; yet despite the variety thus afforded, the tumors must all be looked upon as the consequence of alterations in cells of a single sort, namely epidermal cells affected by the virus, and the alterations themselves have taken a single direction. In the morphology of many of the cancers the influence of the virus is still manifest. The better the papilloma grew, the more likely was cancer to occur, and the greater was the tendency to multiple tumors. In the most favorable rabbits malignant changes took place at numerous locations in the papillomatous tissue, and were imminent at many others. Intercurrent factors had much to do with determining frank carcinosis; and when the tendency to it was not marked their influence sometimes seemed crucial. Analogous instances of a graded alteration from papilloma to cancer are frequent in human pathology. The virus that gives rise to the rabbit papillomas must be looked upon as the primary cause of the cancers developing therefrom. Whether it is their proximate cause has yet to be determined.


[Use of health services and progression of AIDS in persons with HIV infection in Belo Horizonte (Minas Gerais), Brazil].
A nonconcurrent prospective study was done on the relationship between a number of variables and the progression to AIDS among persons diagnosed with the human immunodeficiency virus (HIV). The variables included sociodemographic ones, behavioral ones, and, most importantly, those persons' difficulties in obtaining public health services for HIV/AIDS. The course of the infection was monitored from the first to the last visit to the health services by means of an individual, diagnostic-based classification, using categories established in 1993 by the Centers for Disease Control and Prevention of the United States of America. Participating in the study were 758 patients seen between 1989 and 1992 in the public AIDS referral services of the city of Belo Horizonte (Minas Gerais). All the persons had been diagnosed with HIV and classified in a pre-AIDS stage. Both the patients who developed AIDS during the study as well as those who did not were assessed according to the selected study characteristics. During the study period, 39.5% of the patients developed AIDS. For the group as a whole, the median time without AIDS was 32.4 months. Multivariate analysis showed that the patients who had less risk of developing AIDS were those who had had fewer than 8.8 medical consultations per year (relative risk = 0.36; 95% confidence interval, 0.26 to 0.50) and an interval of at least 6 months between consultations (RR = 0.37; 95% CI, 0.25 to 0.55). The risk was greater in patients age 30 and older (RR = 1.37; 95% CI, 1.03 to 1.84), in those who were not treated with zidovudine (AZT) (RR = 1.91; 95% CI, 1.37 to 2.64), and those who were initially classified in stage "B" of the disease (RR = 4.83; 95% CI, 3.59 to 6.48). The results of this investigation show the dynamics of the supply and demand of services by these patients, and the information will be useful in planning and organizing care for persons with HIV. Recommendations include giving priority to early intervention with a focus on ongoing outpatient care, and more study of the process that persons with HIV follow in seeking and obtaining health care.


Modelling biomechanics of bark patterning in grasstrees.
Bark patterns are a visually important characteristic of trees, typically attributed to fractures occurring during secondary growth of the trunk and branches. An understanding of bark pattern formation has been hampered by insufficient information regarding the biomechanical properties of bark and the corresponding difficulties in faithfully modelling bark fractures using continuum mechanics. This study focuses on the genus Xanthorrhoea (grasstrees), which have an unusual bark-like structure composed of distinct leaf bases connected by sticky resin. Due to its discrete character, this structure is well suited for computational studies. A dynamic computational model of grasstree development was created. The model captures both the phyllotactic pattern of leaf bases during primary growth and the changes in the trunk's width during secondary growth. A biomechanical representation based on a system of masses connected by springs is used for the surface of the trunk, permitting the emergence of fractures during secondary growth to be simulated. The resulting fracture patterns were analysed statistically and compared with images of real trees. The model reproduces key features of grasstree bark patterns, including their variability, spanning elongated and reticulate forms. The patterns produced by the model have the same statistical character as those seen in real trees. The model was able to support the general hypothesis that the patterns observed in the grasstree bark-like layer may be explained in terms of mechanical fractures driven by secondary growth. Although the generality of the results is limited by the unusual structure of grasstree bark, it supports the hypothesis that bark pattern formation is primarily a biomechanical phenomenon.


Regulation of cone cell formation by Canoe and Ras in the developing Drosophila eye.
Cone cells are lens-secreting cells in ommatidia, the unit eyes that compose the compound eye of Drosophila. Each ommatidium contains four cone cells derived from precursor cells of the R7 equivalence group which express the gene sevenless (sev). When a constitutively active form of Ras1 (Ras1V12) is expressed in the R7 equivalence group cells using the sev promoter (sev-Ras1V12), additional cone cells are formed in the ommatidium. Expression of Ras1N17, a dominant negative form of Ras1, results in the formation of 1-3 fewer cone cells than normal in the ommatidium. The effects of Ras1 variants on cone cell formation are modulated by changing the gene dosage at the canoe (cno) locus, which encodes a cytoplasmic protein with Ras-binding activity. An increase or decrease in gene dosage potentiates the sev-Ras1v12 action, leading to marked induction of cone cells. A decrease in cno+ activity also enhances the sev-Ras1N17 action, resulting in a further decrease in the number of cone cells contained in the ommatidium. In the absence of expression of sev-Ras1V12 or sev-Ras1N17, an overdose of wild-type cno (cno+) promotes cone cell formation while a significant reduction in cno+ activity results in the formation of 1-3 fewer cone cells than normal in the ommatidium. We propose that there are two signaling pathways in cone cell development, one for its promotion and the other for its repression, and Cno functions as a negative regulator for both pathways. We also postulate that Cno predominantly acts on a prevailing pathway in a given developmental context, thereby resulting in either an increase or a decrease in the number of cone cells per ommatidium. The extra cone cells resulting from the interplay of Ras1v12 and Cno are generated from a pool of undifferentiated cells that are normally fated to develop into pigment cells or undergo apoptosis.


The Age-Varying Association of Student Status with Excessive Alcohol Use: Ages 18 to 30 Years.
There is a well-known link between attending college and engaging in excessive alcohol use. This study examines in a national sample how the association between student status and excessive alcohol use changes from late adolescence through young adulthood and whether the association of student status with excessive alcohol use is different for students residing with versus away from parents during the school year. This study used cross-sectional data from the National Epidemiologic Survey on Alcohol and Related Conditions, a nationally representative sample of noninstitutionalized civilian adults residing in the United States. Our analyses included nonhigh school young adults who were ages 18 to 30 years (n = 8,645). Excessive alcohol use included past-year (i) high-intensity drinking (men: ≥10 standard drinks; women: ≥8) and (ii) exceeding weekly drinking guidelines (men: >14 drinks per week; women: >7). Students who resided away from their parents and students who lived with their parents during the school year were compared to nonstudents. Analyses using time-varying effect modeling showed that the relationship of student status with excessive alcohol use varied as a function of age. Overall student status lost its association with excessive alcohol use in the early 20s, after controlling for demographics and other adult social roles. The association between student status and excessive alcohol use also varied considerably across age and depending on whether the student was residing with or away from parents. The association of student status with excessive alcohol use is heterogeneous in terms of both age and living arrangements, suggesting opportunities for interventions targeting problematic alcohol use. Future research should examine additional sources of heterogeneity of students in their risk for excessive alcohol use.


Research advances on the usage of traditional Chinese medicine for neuroprotection in glaucoma.
Progressive loss of retinal ganglion cells (RGCs) and their axons is the main pathogenesis of glaucoma. The cause of glaucoma is not fully understood, but the neurodegeneration of glaucoma involves many mechanisms such as oxidative stress, glutamate toxicity and ischemia/reperfusion insult. In order to target these mechanisms, multiple neuroprotective interventions have been investigated to prevent the death of RGCs. Of note are some tonic herbs from the traditional Chinese medicine (TCM) pharmacopeia that have shown neuroprotective effects in glaucoma. TCM differs from Western medicine in that TCM exhibits complicated bioactive components, triggering many signaling pathways and extensive actions on vital organs. Modern scientific approaches have demonstrated some of their underlying mechanisms. In this review, we used Lycium barbarum and Ginkgo biloba as examples to elaborate the characteristics of TCM and their potential applications in neuroprotection in glaucoma.


Consistency of interrater scoring of student performances of osteopathic manipulative treatment on COMLEX-USA Level 2-PE.
Assessment of osteopathic manipulative treatment (OMT) is included in the National Board of Osteopathic Medical Examiners' Comprehensive Osteopathic Medical Licensing Examination-USA Level 2-Performance Evaluation (COMLEX-USA Level 2-PE). The scores earned for OMT should be equivalent among all raters regardless of which technique is scored or which rater is scoring the performance. As a quality assurance measure, selected examination dates and the encounters within the administration of COMLEX-USA Level 2-PE are scored by 2 raters: first by a "live" rater and next by a quality assurance rater. Neither rater knows if he or she is the first or second rater. To compare candidate's scores recorded for OMT on COMLEX-USA Level 2-PE to determine whether differences exist among raters and techniques scored. The authors evaluated candidate performances that took place from July through November 2012. For each performance, 2 raters scored the same technique or different techniques using the OMT scoring rubric. Discrepancies between scores were compared using t tests. Statistical significance was set at P<.05 for most analyses. Of the 708 performances, there was no statistically significant difference in scoring whether the OMT raters scored the same technique or different techniques when the students performed more than 1. There were no statistically significant differences between these results and instances when only a single technique was performed and scored. The present study provides reliability evidence for the use of the global OMT scoring tool in the evaluation of OMT in COMLEX-USA Level 2-PE.


Modeling proximal tubule cell homeostasis: tracking changes in luminal flow.
During normal kidney function, there are routinely wide swings in proximal tubule fluid flow and proportional changes in Na(+) reabsorption across tubule epithelial cells. This "glomerulotubular balance" occurs in the absence of any substantial change in cell volume, and is thus a challenge to coordinate luminal membrane solute entry with peritubular membrane solute exit. In this work, linear optimal control theory is applied to generate a configuration of regulated transporters that could achieve this result. A previously developed model of rat proximal tubule epithelium is linearized about a physiologic reference condition; the approximate linear system is recast as a dynamical system; and a Riccati equation is solved to yield the optimal linear feedback that stabilizes Na(+) flux, cell volume, and cell pH. The first observation is that optimal feedback control is largely consigned to three physiologic variables, cell volume, cell electrical potential, and lateral intercellular hydrostatic pressure. Parameter modulation by cell volume stabilizes cell volume; parameter modulation by electrical potential or interspace pressure act to stabilize Na(+) flux and cell pH. This feedback control is utilized in a tracking problem, in which reabsorptive Na(+) flux varies over a factor of two, in order to represent a substantial excursion of glomerulotubular balance. The resulting control parameters consist of two terms, an autonomous term and a feedback term, and both terms include transporters on both luminal and peritubular cell membranes. Overall, the increase in Na(+) flux is achieved with upregulation of luminal Na(+)/H(+) exchange and Na(+)-glucose cotransport, with increased peritubular Na(+)-3HCO(3)(-) and K(+)-Cl(-) cotransport, and with increased Na(+), K(+)-ATPase activity. The configuration of activated transporters emerges as a testable hypothesis of the molecular basis for glomerulotubular balance. It is suggested that the autonomous control component at each cell membrane could represent the cytoskeletal effects of luminal flow.


Incarnation and animation: physical versus representational deficits of body integrity.
Two apparently disparate phenomena of defective body integrity are reviewed. The first concerns dysmelia, characterized by the congenital absence or deformation of limbs, and the focus of the review is on phantom sensations of people with this kind of physical integrity disorder. The second phenomenon consists of non-psychotic individuals' desire to have a healthy limb amputated, which is interpreted as a mismatch between the physical integrity of a particular limb and its representation in multimodal cortical areas of the brain. We outlined commonalities and differences between the two conditions and note the absence, in both areas of research, of a unified theory that would account for the reported phenomenology. Phantom limbs in dysmelia and the desire for limb amputation most likely are the consequence of very early developmental dissociations between physical and phenomenal body shapes. They are mirror images of one another in that the former constitutes an "animation without incarnation" and, the latter, an "incarnation without animation".


The Saccharomyces cerevisiae Mcm6/2 and Mcm5/3 ATPase active sites contribute to the function of the putative Mcm2-7 'gate'.
The Mcm2-7 complex is the eukaryotic replicative helicase, a toroidal AAA(+) molecular motor that uses adenosine triphosphate (ATP) binding and hydrolysis to separate duplex DNA strands during replication. This heterohexameric helicase contains six different and essential subunits (Mcm2 through Mcm7), with the corresponding dimer interfaces forming ATPase active sites from conserved motifs of adjacent subunits. As all other known hexameric helicases are formed from six identical subunits, the function of the unique heterohexameric organization of Mcm2-7 is of particular interest. Indeed, prior work using mutations in the conserved Walker A box ATPase structural motif strongly suggests that individual ATPase active sites contribute differentially to Mcm2-7 activity. Although only a specific subset of active sites is required for helicase activity, another ATPase active site (Mcm2/5) may serve as a reversible ATP-dependent discontinuity ('gate') within the hexameric ring structure. This study analyzes the contribution that two other structural motifs, the Walker B box and arginine finger, make to each Mcm2-7 ATPase active site. Mutational analysis of these motifs not only confirms that Mcm ATPase active sites contribute unequally to activity but implicates the involvement of at least two additional active sites (Mcm5/3 and 6/2) in modulating the activity of the putative Mcm2/5 gate.


Method of estimating time scales of the atmospheric piston and its application at Dome C (Antarctica).
Analysis of the first interferometric fringes recorded at Dome C, Antarctica are presented. Measurements were taken 31 January and 1 February 2005 during daytime. Our purpose in performing the analysis was to measure temporal fluctuations of the atmospheric piston, which are critical for interferometers, and determine their sensitivity. These scales are derived through the motion of the image that is formed in the focal plane of a Fizeau interferometer. We could establish a lower limit to the coherence time by studying the decay rate of correlation between successive fringes. Coherence times are measured to be larger than 10 ms, i.e., at least three times higher than the median coherence time measured at the site of Paranal (3.3 ms).


On the causes of persistent apical periodontitis: a review.
Apical periodontitis is a chronic inflammatory disorder of periradicular tissues caused by aetiological agents of endodontic origin. Persistent apical periodontitis occurs when root canal treatment of apical periodontitis has not adequately eliminated intraradicular infection. Problems that lead to persistent apical periodontitis include: inadequate aseptic control, poor access cavity design, missed canals, inadequate instrumentation, debridement and leaking temporary or permanent restorations. Even when the most stringent procedures are followed, apical periodontitis may still persist as asymptomatic radiolucencies, because of the complexity of the root canal system formed by the main and accessory canals, their ramifications and anastomoses where residual infection can persist. Further, there are extraradicular factors -- located within the inflamed periapical tissue -- that can interfere with post-treatment healing of apical periodontitis. The causes of apical periodontitis persisting after root canal treatment have not been well characterized. During the 1990s, a series of investigations have shown that there are six biological factors that lead to asymptomatic radiolucencies persisting after root canal treatment. These are: (i) intraradicular infection persisting in the complex apical root canal system; (ii) extraradicular infection, generally in the form of periapical actinomycosis; (iii) extruded root canal filling or other exogenous materials that cause a foreign body reaction; (iv) accumulation of endogenous cholesterol crystals that irritate periapical tissues; (v) true cystic lesions, and (vi) scar tissue healing of the lesion. This article provides a comprehensive overview of the causative factors of non-resolving periapical lesions that are seen as asymptomatic radiolucencies post-treatment.


Local tissue reaction after the application of topical hemostatic agents in a rat partial nephrectomy model.
Various hemostatics are used for renal surgical procedures. We investigated the hemostatic efficacy of cellulose derivatives on the model of partial nephrectomy in rats focusing on the local reaction of renal parenchyma. A total of 50 Wistar rats were divided into five groups of 10 animals each. Partial nephrectomy of the caudal pole without hilar vascular control was performed. Oxidized cellulose (OC), sodium salt of oxycellulose (OCN), carboxymethyl cellulose (CMC), dialdehyde cellulose (DAC), and gelatin-based hemostatic (C) were applied to the bleeding wounds. The time to hemostasis was monitored. Half of the animals were euthanized after 3 days, the second half 30 days from the experiment start date. The left kidney was excised and subjected to histopathological examination. The biochemical data was subjected to statistical analysis. The time to hemostasis in all groups was significantly less than in the C group (in OC p = 0.0057, OCN p = 0.0039, CMC and DAC p = 0.0001). In the C group, massive hemorrhages and necrosis did occur. In the OC and OCN groups, there were regenerative changes, a receding inflammatory reaction and hemorrhage. DAC caused an immune reaction and massive interstitial hemorrhages with biochemical signs of liver damage. Parenchyma in CMC revealed a reduction of necrosis and interstitial hemorrhages with regenerative processes. The most effective hemostatics were CMC and OC, achieving the best results both in the time to hemostasis, and for histopathological evaluation.


Signal representation on the angular Poincaré sphere, based on second-order moments.
Based on the analysis of second-order moments, a generalized canonical representation of a two-dimensional optical signal is proposed, which is associated with the angular Poincaré sphere. Vortex-free (or zero-twist) optical beams arise on the equator of this sphere, while beams with a maximum vorticity (or maximum twist) are located at the poles. An easy way is shown how the latitude on the sphere, which is a measure for the degree of vorticity, can be derived from the second-order moments. The latitude is invariant when the beam propagates through a first-order optical system between conjugate planes. To change the vorticity of a beam, a system that does not operate between conjugate planes is needed, with the gyrator as the prime representative of such a system. A direct way is derived to find an optical system (consisting of a lens, a magnifier, a rotator, and a gyrator) that transforms a beam with an arbitrary moment matrix into its canonical form.


Echocardiographic assessment of raised pulmonary vascular resistance: application to diagnosis and follow-up of pulmonary hypertension.
To optimise an echocardiographic estimation of pulmonary vascular resistance (PVR(e)) for diagnosis and follow-up of pulmonary hypertension (PHT). Cross-sectional study. Tertiary referral centre. Patients undergoing right heart catheterisation and echocardiography for assessment of suspected PHT. PVR(e) ([tricuspid regurgitation velocity ×10/(right ventricular outflow tract velocity-time integral+0.16) and invasive PVR(i) ((mean pulmonary artery systolic pressure-wedge pressure)/cardiac output) were compared in 72 patients. Other echo data included right ventricular systolic pressure (RVSP), estimated right atrial pressure, and E/e' ratio. Difference between PVR(e) and PVR(i) at various levels of PVR was sought using Bland-Altman analysis. Corrected PVR(c) ((RVSP-E/e')/RVOT(VTI)) (RVOT, RV outflow time; VTI, velocity time integral) was developed in the training group and tested in a separate validation group of 42 patients with established PHT. PVR(e)>2.0 had high sensitivity (93%) and specificity (91%) for recognition of PVR(i)>2.0, and PVR(c) provided similar sensitivities and specificities. PVR(e) and PVR(i) correlated well (r=0.77, p<0.01), but PVR(e) underestimated marked elevation of PVR(i)-a trend avoided by PVR(c). PVR(c) and PVR(e) were tested against PVR(i) in a separate validation group (n=42). The mean difference between PVR(e) and PVR(i) exceeded that between PVR(c) and PVR(i) (2.8±2.7 vs 0.8±3.0 Wood units; p<0.001). A drop in PVR(i) by at least one SD occurred in 10 patients over 6 months; this was detected in one patient by PVR(e) and eight patients by PVR(c) (p=0.002). PVR(e) distinguishes normal from abnormal PVR(i) but underestimates high PVR(i). PVR(c) identifies the severity of PHT and may be used to assess treatment response.


The Gene expression Grade Index: a potential predictor of relapse for endocrine-treated breast cancer patients in the BIG 1-98 trial.
We have previously shown that the Gene expression Grade Index (GGI) was able to identify two subtypes of estrogen receptor (ER)-positive tumors that were associated with statistically distinct clinical outcomes in both untreated and tamoxifen-treated patients. Here, we aim to investigate the ability of the GGI to predict relapses in postmenopausal women who were treated with tamoxifen (T) or letrozole (L) within the BIG 1-98 trial. We generated gene expression profiles (Affymetrix) and computed the GGI for a matched, case-control sample of patients enrolled in the BIG 1-98 trial from the two hospitals where frozen samples were available. All relapses (cases) were identified from patients randomized to receive monotherapy or from the switching treatment arms for whom relapse occurred before the switch. Each case was randomly matched with four controls based upon nodal status and treatment (T or L). The prognostic value of GGI was assessed as a continuous predictor and divided at the median. Predictive accuracy of GGI was estimated using time-dependent area under the curve (AUC) of the ROC curves. Frozen samples were analyzable for 48 patients (10 cases and 38 controls). Seven of the 10 cases had been assigned to receive L. Cases and controls were comparable with respect to menopausal and nodal status, local and chemotherapy, and HER2 positivity. Cases were slightly older than controls and had a larger proportion of large, poorly differentiated ER+/PgR- tumors. The GGI was significantly and linearly related to risk of relapse: each 10-unit increase in GGI resulted in an increase of approximately 11% in the hazard rate (p = 0.02). Within the subgroups of patients with node-positive disease or who were treated with L, the hazard of relapse was significantly greater for patients with GGI at or above the median. AUC reached a maximum of 78% at 27 months. This analysis supports the GGI as a good predictor of relapse for ER-positive patients, even among patients who receive L. Validation of these results, in a larger series from BIG 1-98, is planned using the simplified GGI represented by a smaller set of genes and tested by qRT-PCR on paraffin-embedded tissues.


Stabilization of lignin peroxidases in white rot fungi by tryptophan.
Supplementation of various cultures of white rot fungi with tryptophan was found to have a large stimulatory effect on lignin peroxidase activity levels. This enhancement was greater than that observed in the presence of the lignin peroxidase recycling agent veratryl alcohol. Using reverse transcription-PCR, we found that tryptophan does not act to induce lignin peroxidase expression at the level of gene transcription. Instead, the activity enhancement observed is likely to result from the protective effect of tryptophan against H2O2 inactivation. In experiments using a partially purified lignin peroxidase preparation, tryptophan and its derivative indole were determined to function in the same way as veratryl alcohol in converting compound II, an oxidized form of lignin peroxidase, to ferric enzyme, thereby completing the catalytic cycle. Furthermore, tryptophan was found to be a better substrate for lignin peroxidase than veratryl alcohol. Inclusion of either tryptophan or indole enhanced the oxidation of the azo dyes methyl orange and Eriochrome blue black. Stimulation of azo dye oxidations by veratryl alcohol has previously been shown to be due to its enzyme recycling function. Our data allow us to propose that tryptophan stabilizes lignin peroxidase by acting as a reductant for the enzyme.


Cross-sectional associations of active transport, employment status and objectively measured physical activity: analyses from the National Health and Nutrition Examination Survey.
To investigate associations between active transport, employment status and objectively measured moderate-to-vigorous physical activity (MVPA) in a representative sample of US adults. Cross-sectional analyses of data from the National Health and Nutrition Examination Survey. A total of 5180 adults (50.2 years old, 49.0% men) were classified by levels of active transportation and employment status. Outcome measure was weekly time spent in MVPA as recorded by the Actigraph accelerometer. Associations between active transport, employment status and objectively measured MVPA were examined using multivariable linear regression models adjusted for age, body mass index, race and ethnicity, education level, marital status, smoking status, working hour duration (among the employed only) and self-reported leisure time physical activity. Patterns of active transport were similar between the employed (n=2897) and unemployed (n=2283), such that 76.0% employed and 77.5% unemployed engaged in no active transport. For employed adults, those engaging in high levels of active transport (≥90 min/week) had higher amount of MVPA than those who did not engage in active transport. This translated to 40.8 (95% CI 15.7 to 65.9) additional minutes MVPA per week in men and 57.9 (95% CI 32.1 to 83.7) additional minutes MVPA per week in women. Among the unemployed adults, higher levels of active transport were associated with more MVPA among men (44.8 min/week MVPA, 95% CI 9.2 to 80.5) only. Findings from the present study support interventions to promote active transport to increase population level physical activity. Additional strategies are likely required to promote physical activity among unemployed women.


Local and regional mechanical characterisation of a collagen-glycosaminoglycan scaffold using high-resolution finite element analysis.
Artificial tissue growth requires cells to proliferate and differentiate within the host scaffold. As cell function is governed by mechano-sensitive selection, tissue type is influenced by the microscopic forces exposed to the cells, which is a product of macroscopically straining the scaffold. Accordingly, the microscopic strain environment within a CG scaffold is offered here. Using muCT to characterise CG scaffold architecture, two high-resolution 3D FE models were used to predict the deformation mechanics. While also providing an analysis of region-specific features, such as relative density, pore diameters and microstructural elastic stability, the deformation patterns afforded strains to be inferred for seeded cells. The results indicate a regional dependence, in terms of architectural and mechanical properties. Specifically, the peripheral regions demonstrated the lowest volume fraction, the highest stress concentrations and the greatest potential for elastic instability. Conversely, the mid-region exhibited the most homogeneous environment. Based on the proviso of mechano-sensitive proliferation and differentiation, the findings suggest cell function will vary between CG scaffold regions. Further work should investigate the possibility of improving the fabrication process in order to deliver a construct in line with the mid-region, or alternatively, isolation of the mid-region may prove beneficial for cell culturing.


The genome of the African trypanosome Trypanosoma brucei.
African trypanosomes cause human sleeping sickness and livestock trypanosomiasis in sub-Saharan Africa. We present the sequence and analysis of the 11 megabase-sized chromosomes of Trypanosoma brucei. The 26-megabase genome contains 9068 predicted genes, including approximately 900 pseudogenes and approximately 1700 T. brucei-specific genes. Large subtelomeric arrays contain an archive of 806 variant surface glycoprotein (VSG) genes used by the parasite to evade the mammalian immune system. Most VSG genes are pseudogenes, which may be used to generate expressed mosaic genes by ectopic recombination. Comparisons of the cytoskeleton and endocytic trafficking systems with those of humans and other eukaryotic organisms reveal major differences. A comparison of metabolic pathways encoded by the genomes of T. brucei, T. cruzi, and Leishmania major reveals the least overall metabolic capability in T. brucei and the greatest in L. major. Horizontal transfer of genes of bacterial origin has contributed to some of the metabolic differences in these parasites, and a number of novel potential drug targets have been identified.


Paroxysmal nocturnal hemoglobinuria.
Paroxysmal nocturnal hemoglobinuria (PNH) is a rare bone marrow failure disorder that manifests with hemolytic anemia, thrombosis, and peripheral blood cytopenias. The absence of two glycosylphosphatidylinositol (GPI)-anchored proteins, CD55 and CD59, leads to uncontrolled complement activation that accounts for hemolysis and other PNH manifestations. GPI anchor protein deficiency is almost always due to somatic mutations in phosphatidylinositol glycan class A (PIGA), a gene involved in the first step of GPI anchor biosynthesis; however, alternative mutations that cause PNH have recently been discovered. In addition, hypomorphic germ-line PIGA mutations that do not cause PNH have been shown to be responsible for a condition known as multiple congenital anomalies-hypotonia-seizures syndrome 2. Eculizumab, a first-in-class monoclonal antibody that inhibits terminal complement, is the treatment of choice for patients with severe manifestations of PNH. Bone marrow transplantation remains the only cure for PNH but should be reserved for patients with suboptimal response to eculizumab.


Strategies for controlling non-transmissible infection outbreaks using a large human movement data set.
Prediction and control of the spread of infectious disease in human populations benefits greatly from our growing capacity to quantify human movement behavior. Here we develop a mathematical model for non-transmissible infections contracted from a localized environmental source, informed by a detailed description of movement patterns of the population of Great Britain. The model is applied to outbreaks of Legionnaires' disease, a potentially life-threatening form of pneumonia caused by the bacteria Legionella pneumophilia. We use case-report data from three recent outbreaks that have occurred in Great Britain where the source has already been identified by public health agencies. We first demonstrate that the amount of individual-level heterogeneity incorporated in the movement data greatly influences our ability to predict the source location. The most accurate predictions were obtained using reported travel histories to describe movements of infected individuals, but using detailed simulation models to estimate movement patterns offers an effective fast alternative. Secondly, once the source is identified, we show that our model can be used to accurately determine the population likely to have been exposed to the pathogen, and hence predict the residential locations of infected individuals. The results give rise to an effective control strategy that can be implemented rapidly in response to an outbreak.


Small Supernumerary Marker Chromosomes in Human Infertility.
Small supernumerary marker chromosomes (sSMC) are structurally abnormal chromosomes that cannot be unambiguously identified by banding cytogenetics. The objective of this study was to provide an overview of sSMC frequency and characterization in a context of infertility and to review the literature describing sSMC in relation with male and female infertility. Therefore, a systematic literature review on sSMC associated with infertility was conducted by means of a PubMed literature and a sSMC database (http://ssmc-tl.com/sSMC.html) search. A total of 234 patients with infertility were identified as carriers of sSMC. All chromosomes, except chromosomes 10, 19 and the X, were involved in sSMC, and in 72% the sSMC originated from acrocentric chromosomes. Euchromatic imbalances were caused by the presence of sSMC in 30% of the cases. Putative genes have been identified in only 1.2% of sSMC associated with infertility. The implication of sSMC in infertility could be due to a partial trisomy of some genes but also to mechanical effects perturbing meiosis. Further precise molecular and interphase-architecture studies on sSMC are needed in the future to characterize the relationship between this chromosomal anomaly and human infertility.


Axons regulate Schwann cell expression of the major myelin and NGF receptor genes.
The elaboration of myelin by Schwann cells is triggered by contact with appropriate peripheral axons. Among the most prominent features of this interaction is the activation and high-level expression of the genes encoding the major myelin proteins P0 and Myelin Basic Protein (MBP). Although the initial induction of these genes is thought to be dependent upon contact with axons, neither the inductive signal of the axon nor the receptor and associated second messenger system of the Schwann cell that transduces this signal has been identified. In this report, we demonstrate that expression of the P0 and MBP genes in rapidly myelinating Schwann cells is sharply reduced upon withdrawal of axons, but that this expression can be substantially restored by agents that raise the intracellular concentration of cyclic AMP. We further show that Schwann cell expression of a third gene, i.e. that encoding the Nerve Growth Factor receptor, is strongly activated by the withdrawal of axons, and that this activation is largely independent of cAMP.


snRNAs contain specific SMN-binding domains that are essential for snRNP assembly.
To serve in its function as an assembly machine for spliceosomal small nuclear ribonucleoprotein particles (snRNPs), the survival of motor neurons (SMN) protein complex binds directly to the Sm proteins and the U snRNAs. A specific domain unique to U1 snRNA, stem-loop 1 (SL1), is required for SMN complex binding and U1 snRNP Sm core assembly. Here, we show that each of the major spliceosomal U snRNAs (U2, U4, and U5), as well as the minor splicing pathway U11 snRNA, contains a domain to which the SMN complex binds directly and with remarkable affinity (low nanomolar concentration). The SMN-binding domains of the U snRNAs do not have any significant nucleotide sequence similarity yet they compete for binding to the SMN complex in a manner that suggests the presence of at least two binding sites. Furthermore, the SMN complex-binding domain and the Sm site are both necessary and sufficient for Sm core assembly and their relative positions are critical for snRNP assembly. These findings indicate that the SMN complex stringently scrutinizes RNAs for specific structural features that are not obvious from the sequence of the RNAs but are required for their identification as bona fide snRNAs. It is likely that this surveillance capacity of the SMN complex ensures assembly of Sm cores on the correct RNAs only and prevents illicit, potentially deleterious, assembly of Sm cores on random RNAs.


Establishment and identification of a hypoxia-ischemia brain damage model in neonatal rats.
The present study was designed to set up a reliable model of severe hypoxia-ischemia brain damage (HIBD) in neonatal rats and several methods were used to identify whether the model was successful. A total of 40 healthy 7-day-old Sprague-Dawley rats were randomly divided into 2 groups: The sham-surgery group (n=18) and the HIBD model group (n=22). The HIBD model was produced according to the traditional Rice method. The rats were anesthetized with ethyl ether. The left common carotid artery (CCA) was exposed, ligated and cut. Following this, the rats were exposed to hypoxia in a normobaric chamber filled with 8% oxygen and 92% nitrogen for 2 h. In the sham-surgery group, the left CCA was exposed but was not ligated, cut or exposed to hypoxia. The neurobehavioral changes of the rats were observed in the 24 h after HIBD. The brains were collected after 72 h to observe the pathological morphological changes of the brain tissue. The behavioral ability and neurobehavioral changes were studied in each group. The water maze test was used for evaluating the learning-memory ability when the rats were 28 days old. Compared with the sham-surgery group, all the HIBD model rats had a lag of motor development. The rats had evident changes in anatomy and Nissl staining, and cognitive impairment was shown through the result of the water maze. Therefore, the model of HIBD in neonatal rats is feasible and provides a reliable model for subsequent studies.


[LDL oxidation in homozygous familial hypercholesterolemia: effects of selective LDL-apheresis treatment].
Homozygous familial hypercholesterolemia (HFH) results from a mutation affecting both the structure and function of a cell surface receptor that removes low density lipoproteins (LDL) from plasma. The disorder is characterized by autosomal dominant inheritance, a lifelong elevation in the concentration of LDL-bound cholesterol in blood and by cholesterol deposits that form xanthomas and early coronary artery disease. HFH patients, as a result of the increased levels and prolonged residence time of LDL in plasma, have a strong tendency toward accumulation of LDL-cholesterol in the arterial wall causing premature atherosclerosis. Selective LDL-apheresis (LA) on dextran/sulphate cellulose columns is the best therapy reducing mortality of these patients. We previously showed that prolonged lifelong enhanced LDL oxidation in HFH. LDL undergo oxidation before being taken up by macrophages then transformed into foam cells. At the present time, the relevance of the in vitro macrophages studies to the accumulation of cholesterol esters in scavenger cells of HFH patients is not yet established. The aim of this study was to investigate LDL oxidation, induced by xanthine (2 mM)+xanthine oxidase (100 mU), and cholesterol esterification in macrophages, in 8 HFH patients before and after LA. LDL peroxidation by conjugated-diene absorbance showed an increased resistance against oxidation after LA: lag time 129 +/- 25 vs 112 +/- 27 min, p < 0.05; diene production 9.1 +/- 2.1 vs 13.9 +/- 2.5 nM/min/mg LDL, p < 0.01. Peroxidation was also evaluated from lipid peroxides (158 +/- 34 vs 57 +/- 18 nM/mg protein after LA, p < 0.05) and malonyldialdehyde (38 +/- 12 vs 27 +/- 8 nM/mg protein after LA, p < 0.05) content. When oxidized LDL was run on polyacrylamide gel extensive apo-B100 fragmentation was observed in LDL before LA, vs a less fragmentation after LA. A similar reduction was obtained in LDL agarose mobility after LA (1.7 +/- 0.2 vs 2.5 +/- 0.2, p < 0.05). Cholesterol esterification in mouse peritoneal macrophages was also decreased after LA (8.5 +/- 1.8 vs 14.6 +/- 2.7 nM/mg cell protein/12 hours, p < 0.05). Vitamin E content of LDL (mg/g protein) was increased after LA (4.44 +/- 1.0 vs 3.9 +/- 1.2, p < 0.05). Thus, selective LA, not only decreases the pool of LDL, but it also induces changes that render LDL less susceptible to oxidation and decreased high cholesterol esterification in macrophages. The prevention of these mechanisms by LA contributes actively to retard atherogenesis in HFH patients.


Coactivation of the CLOCK-BMAL1 complex by CBP mediates resetting of the circadian clock.
The transcription factor CLOCK-BMAL1 is a core component of the molecular clock machinery that drives circadian gene expression and physiology in mammals. Recently, we reported that this heterodimeric transcription factor functions as a signaling molecule in response to the resetting stimuli via the Ca²+-dependent protein kinase C pathway. Here, we demonstrate that the CREB-binding protein (CBP) plays a key role in rapid activation of the CLOCK-BMAL1 heterodimer that leads to phase resetting of the circadian clock. Under physiological conditions, a bimolecular fluorescence complementation (BiFC) assay revealed that CLOCK and BMAL1 dimerize in the cytoplasm and subsequently translocate into the nucleus in response to serum stimuli (mean time duration was 29.2 minutes and mean velocity 0.7 μm/minute). Concomitantly, BMAL1 rapidly recruited CBP on Per1 promoter E-box, but not p300 (a functional analog of CBP), in the discrete nuclear foci. However, recruitment of CBP by cAMP/Ca²+ response element-binding (CREB) protein on CRE was not markedly increased upon delivery of the resetting stimuli. Furthermore, overexpression of CBP greatly potentiated the CLOCK-BMAL1-mediated Per1 transcription, and this effect was completely abolished by site-directed mutation of E-box elements, but not by the mutation of CRE in the Per1 promoter. Furthermore, molecular knockdown of CBP severely dampened circadian oscillation of clock gene expression triggered by the resetting stimuli. These findings suggest that CBP recruitment by BMAL1 mediates acute transactivation of CLOCK-BMAL1, thereby inducing immediate-early Per1 transcription and phase resetting of the circadian clock.


Altered trabecular bone structure and delayed cartilage degeneration in the knees of collagen VI null mice.
Mutation or loss of collagen VI has been linked to a variety of musculoskeletal abnormalities, particularly muscular dystrophies, tissue ossification and/or fibrosis, and hip osteoarthritis. However, the role of collagen VI in bone and cartilage structure and function in the knee is unknown. In this study, we examined the role of collagen VI in the morphology and physical properties of bone and cartilage in the knee joint of Col6a1(-/-) mice by micro-computed tomography (microCT), histology, atomic force microscopy (AFM), and scanning microphotolysis (SCAMP). Col6a1(-/-) mice showed significant differences in trabecular bone structure, with lower bone volume, connectivity density, trabecular number, and trabecular thickness but higher structure model index and trabecular separation compared to Col6a1(+/+) mice. Subchondral bone thickness and mineral content increased significantly with age in Col6a1(+/+) mice, but not in Col6a1(-/-) mice. Col6a1(-/-) mice had lower cartilage degradation scores, but developed early, severe osteophytes compared to Col6a1(+/+) mice. In both groups, cartilage roughness increased with age, but neither the frictional coefficient nor compressive modulus of the cartilage changed with age or genotype, as measured by AFM. Cartilage diffusivity, measured via SCAMP, varied minimally with age or genotype. The absence of type VI collagen has profound effects on knee joint structure and morphometry, yet minimal influences on the physical properties of the cartilage. Together with previous studies showing accelerated hip osteoarthritis in Col6a1(-/-) mice, these findings suggest different roles for collagen VI at different sites in the body, consistent with clinical data.


GABA, not BOLD, reveals dissociable learning-dependent plasticity mechanisms in the human brain.
Experience and training have been shown to facilitate our ability to extract and discriminate meaningful patterns from cluttered environments. Yet, the human brain mechanisms that mediate our ability to learn by suppressing noisy and irrelevant signals remain largely unknown. To test the role of suppression in perceptual learning, we combine fMRI with MR Spectroscopy measurements of GABA, as fMRI alone does not allow us to discern inhibitory vs. excitatory mechanisms. Our results demonstrate that task-dependent GABAergic inhibition relates to functional brain plasticity and behavioral improvement. Specifically, GABAergic inhibition in the occipito-temporal cortex relates to dissociable learning mechanisms: decreased GABA for noise filtering, while increased GABA for feature template retuning. Perturbing cortical excitability during training with tDCs alters performance in a task-specific manner, providing evidence for a direct link between suppression and behavioral improvement. Our findings propose dissociable GABAergic mechanisms that optimize our ability to make perceptual decisions through training.


Comparative metagenomic analysis of plasmid encoded functions in the human gut microbiome.
Little is known regarding the pool of mobile genetic elements associated with the human gut microbiome. In this study we employed the culture independent TRACA system to isolate novel plasmids from the human gut microbiota, and a comparative metagenomic analysis to investigate the distribution and relative abundance of functions encoded by these plasmids in the human gut microbiome. Novel plasmids were acquired from the human gut microbiome, and homologous nucleotide sequences with high identity (>90%) to two plasmids (pTRACA10 and pTRACA22) were identified in the multiple human gut microbiomes analysed here. However, no homologous nucleotide sequences to these plasmids were identified in the murine gut or environmental metagenomes. Functions encoded by the plasmids pTRACA10 and pTRACA22 were found to be more prevalent in the human gut microbiome when compared to microbial communities from other environments. Among the most prevalent functions identified was a putative RelBE toxin-antitoxin (TA) addiction module, and subsequent analysis revealed that this was most closely related to putative TA modules from gut associated bacteria belonging to the Firmicutes. A broad phylogenetic distribution of RelE toxin genes was observed in gut associated bacterial species (Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria), but no RelE homologues were identified in gut associated archaeal species. We also provide indirect evidence for the horizontal transfer of these genes between bacterial species belonging to disparate phylogenetic divisions, namely Gram negative Proteobacteria and Gram positive species from the Firmicutes division. The application of a culture independent system to capture novel plasmids from the human gut mobile metagenome, coupled with subsequent comparative metagenomic analysis, highlighted the unexpected prevalence of plasmid encoded functions in the gut microbial ecosystem. In particular the increased relative abundance and broad phylogenetic distribution was identified for a putative RelBE toxin/antitoxin addiction module, a putative phosphohydrolase/phosphoesterase, and an ORF of unknown function. Our analysis also indicates that some plasmids or plasmid families are present in the gut microbiomes of geographically isolated human hosts with a broad global distribution (America, Japan and Europe), and are potentially unique to the human gut microbiome. Further investigation of the plasmid population associated with the human gut is likely to provide important insights into the development, functioning and evolution of the human gut microbiota.


Mechanisms of homomeric alpha1 glycine receptor endocytosis.
Little is known regarding the mechanism(s) by which glycine receptors are endocytosed. Here we examined the endocytosis of homomeric alpha1 glycine receptors expressed in HEK 293 cells using immunofluorescence/confocal microscopy and whole-cell patch-clamp recordings. Our studies demonstrate that constitutive endocytosis of glycine receptors is blocked by the dominant negative dynamin construct K44A and that intracellular dialysis with peptide P4, a dynamin/amphiphysin-disrupting peptide, increased whole-cell glycine-gated chloride currents. To examine whether receptor endocytosis could be regulated by PKC, experiments with the PKC activator PMA (phorbol 12-myristate 13-acetate) were performed. PMA, but not its inactive analogue PMM (phorbol 12-monomyristate), stimulated receptor endocytosis and inhibited glycine-gated chloride currents. Similar to constitutive endocytosis, PKC-stimulated endocytosis was blocked by dynamin K44A. Mutation of a putative AP2 adaptin dileucine motif (L314A, L315A) present in the receptor cytoplasmic loop blocked PMA-stimulated receptor endocytosis and also prevented PMA inhibition of glycine receptor currents. In patch-clamp experiments, intracellular dialysis of a 12-amino acid peptide corresponding to the region of the receptor containing the dileucine motif prevented PKC modulation of wild-type glycine receptors. Unlike PKC modulation of the receptor, constitutive endocytosis was not affected by mutation of this dileucine motif. These results demonstrate that PKC activation stimulates glycine receptor endocytosis, that both constitutive endocytosis and PKC-stimulated endocytosis are dynamin-dependent, and that PKC-stimulated endocytosis, but not constitutive endocytosis, occurs via the dileucine motif (L314A, L315A) within the cytoplasmic loop of the receptor.


Evidence for B0 s-->phiphi decay and measurements of branching ratio and A(CP) for B+ -->phiK+.
We present the first evidence of charmless decays of the B(0)(s) meson, the decay B(0)(s)--> phiphi, and a measurement of the branching ratio BR(B(0)(s)--> phiphi) using 180 pb(-1) of data collected by the CDF II experiment at the Fermilab Tevatron collider. In addition, the BR and direct CP asymmetry for the B+-->phiK+ decay are measured. We obtain BR(B(0)(s)--> phiphi)=[14(+6)(-5)(stat)+/-6(syst)] x 10(-6), BR(B+-->phiK+)=[7.6+/-1.3(stat)+/-0.6(syst)] x 10(-6), and A(CP)(B+-->phiK+)= -0.07+/-0.17(stat)+0.03 / -0.02(syst). Both decays are governed in the standard model by second order (penguin) b-->s(-)ss amplitudes.


Total cost of care lower among Medicare fee-for-service beneficiaries receiving care from patient-centered medical homes.
To compare health care utilization and payments between NCQA-recognized patient-centered medical home (PCMH) practices and practices without such recognition. Medicare Part A and B claims files from July 1, 2007 to June 30, 2010, 2009 Census, 2007 Health Resources and Services Administration and CMS Utilization file, Medicare's Enrollment Data Base, and the 2005 American Medical Association Physician Workforce file. This study used a longitudinal, nonexperimental design. Three annual observations (July 1, 2008-June 30, 2010) were available for each practice. We compared selected outcomes between practices with and those without NCQA PCMH recognition. Individual Medicare fee-for-service (FFS) beneficiaries and their claims and utilization data were assigned to PCMH or comparison practices based on where they received the plurality of evaluation and management services between July 1, 2007 and June 30, 2008. Relative to the comparison group, total Medicare payments, acute care payments, and the number of emergency room visits declined after practices received NCQA PCMH recognition. The decline was larger for practices with sicker than average patients, primary care practices, and solo practices. This study provides additional evidence about the potential of the PCMH model for reducing health care utilization and the cost of care.