Real Time Enzyme Inhibition Assays Provide Insights into Differences in Binding of Neuraminidase Inhibitors to Wild Type and Mutant Influenza Viruses

The influenza neuraminidase (NA) inhibitors zanamivir, oseltamivir and peramivir were all designed based on the knowledge that the transition state analogue of the cleaved sialic acid, 2-deoxy,2,3-dehydro N-acetyl neuraminic acid (DANA) was a weak inhibitor of NA. While DANA bound rapidly to the NA, modifications leading to the improved potency of these new inhibitors also conferred a time dependent or slow binding phenotype. Many mutations in the NA leading to decreased susceptibility result in loss of slow binding, hence this is a phenotypic marker of many but not all resistant NAs. We present here a simplified approach to determine whether an inhibitor is fast or slow binding by extending the endpoint fluorescent enzyme inhibition assay to a real time assay and monitoring the changes in IC50s with time. We carried out two reactions, one with a 30 min preincubation with inhibitor and the second without. The enzymatic reaction was started via addition of substrate and IC50s were calculated after each 10 min interval up to 60 min. Results showed that without preincubation IC50s for the wild type viruses started high and although they decreased continuously over the 60 min reaction time the final IC50s remained higher than for pre-incubated samples. These results indicate a slow equilibrium of association and dissociation and are consistent with slow binding of the inhibitors. In contrast, for viruses with decreased susceptibility, preincubation had minimal effect on the IC50s, consistent with fast binding. Therefore this modified assay provides additional phenotypic information about the rate of inhibitor binding in addition to the IC50, and critically demonstrates the differential effect of incubation times on the IC50 and Ki values of wild type and mutant viruses for each of the inhibitors

Understanding pregnancy planning in a low-income country setting: validation of the London measure of unplanned pregnancy in Malawi

This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.Background: The London Measure of Unplanned Pregnancy (LMUP) is a new and psychometrically valid measure of pregnancy intention that was developed in the United Kingdom. An improved understanding of pregnancy intention in low-income countries, where unintended pregnancies are common and maternal and neonatal deaths are high, is necessary to inform policies to address the unmet need for family planning. To this end this research aimed to validate the LMUP for use in the Chichewa language in Malawi.Methods: Three Chichewa speakers translated the LMUP and one translation was agreed which was back-translated and pre-tested on five pregnant women using cognitive interviews. The measure was field tested with pregnant women who were recruited at antenatal clinics and data were analysed using classical test theory and hypothesis testing.Results: 125 women aged 15-43 (median 23), with parities of 1-8 (median 2) completed the Chichewa LMUP. There were no missing data. The full range of LMUP scores was captured. In terms of reliability, the scale was internally consistent (Cronbach's alpha = 0.78) and test-retest data from 70 women showed good stability (weighted Kappa 0.80). In terms of validity, hypothesis testing confirmed that unmarried women (p = 0.003), women who had four or more children alive (p = 0.0051) and women who were below 20 or over 29 (p = 0.0115) were all more likely to have unintended pregnancies. Principal component analysis showed that five of the six items loaded onto one factor, with a further item borderline. A sensitivity analysis to assess the effect of the removal of the weakest item of the scale showed slightly improved performance but as the LMUP was not significantly adversely affected by its inclusion we recommend retaining the six-item score.Conclusion: The Chichewa LMUP is a valid and reliable measure of pregnancy intention in Malawi and can now be used in research and/or surveillance. This is the first validation of this tool in a low-income country, helping to demonstrate that the concept of pregnancy planning is applicable in such a setting. Use of the Chichewa LMUP can enhance our understanding of pregnancy intention in Malawi, giving insight into the family planning services that are required to better meet women's needs and save lives. © 2013 Hall et al.; licensee BioMed Central Ltd.Dr Hall’s Wellcome Trust Research Training Fellowship, grant number 097268/Z/11/Z

Report from the PredictER Expert Panel Meeting, November 2, 2007

On November 2, 2007, the Indiana University Center for Bioethics convened an expert panel on predictive health research (PHR) as part of the Center’s Program in Predictive Health Ethics Research (http://www.bioethics.iu.edu/predicter.asp) which is supported by a grant from the Richard M. Fairbanks Foundation. The goal of this meeting was to identify the major obstacles and opportunities for engaging the community in PHR. PredictER intends to use the results of this meeting as a first step toward more fully engaging the Indianapolis community in discussions about PHR.Richard M. Fairbanks Foundatio

Traumatic Brain Injury-Practice Based Evidence Study: Design and Patients, Centers, Treatments, and Outcomes

OBJECTIVES: To describe study design, patients, centers, treatments, and outcomes of a traumatic brain injury (TBI) practice-based evidence (PBE) study and to evaluate the generalizability of the findings to the U.S. TBI inpatient rehabilitation population. DESIGN: Prospective, longitudinal, observational study. SETTING: Ten inpatient rehabilitation centers. PARTICIPANTS: Patients (N=2130) enrolled between October 2008 and September 2011 and admitted for inpatient rehabilitation after an index TBI injury. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Return to acute care during rehabilitation, rehabilitation length of stay, FIM at discharge, residence at discharge, and 9 months postdischarge rehospitalization, FIM, participation, and subjective well-being. RESULTS: The level of admission FIM cognitive score was found to create relatively homogeneous subgroups for the subsequent analysis of best treatment combinations. There were significant differences in patient and injury characteristics, treatments, rehabilitation course, and outcomes by admission FIM cognitive subgroups. TBI-PBE study patients were overall similar to U.S. national TBI inpatient rehabilitation populations. CONCLUSIONS: This TBI-PBE study succeeded in capturing naturally occurring variation in patients and treatments, offering opportunities to study best treatments for specific patient impairments. Subsequent articles in this issue report differences between patients and treatments and associations with outcomes in greater detail

The human papillomavirus E7 proteins associate with p190RhoGAP and alter its function

Using mass spectrometry, we identified p190RhoGAP (p190) as a binding partner of human papillomavirus 16 (HPV16) E7. p190 belongs to the GTPase activating protein (GAP) family and is one of the primary GAPs for RhoA. GAPs stimulate the intrinsic GTPase activity of the Rho proteins, leading to Rho inactivation and influencing numerous biological processes. RhoA is one of the best-characterized Rho proteins and is specifically involved in formation of focal adhesions and stress fibers, thereby regulating cell migration and cell spreading. Since this is the first report that E7 associates with p190, we carried out detailed interaction studies. We show that E7 proteins from other HPV types also bind p190. Furthermore, we found that conserved region 3 (CR3) of E7 and the middle domain of p190 are important for this interaction. More specifically, we identified two residues in CR3 of E7 that are necessary for p190 binding and used mutants of E7 with mutations of these residues to determine the biological consequences of the E7-p190 interaction. Our data suggest that the interaction of E7 with p190 dysregulates this GAP and alters the actin cytoskeleton. We also found that this interaction negatively regulates cell spreading on a fibronectin substrate and therefore likely contributes to important aspects of the HPV life cycle or HPV-induced tumorigenesis. © 2014, American Society for Microbiology

Associations of cannabis use disorder with cognition, brain structure, and brain function in African Americans

Although previous studies have highlighted associations of cannabis use with cognition and brain morphometry, critical questions remain with regard to the association between cannabis use and brain structural and functional connectivity. In a cross-sectional community sample of 205 African Americans (age 18–70) we tested for associations of cannabis use disorder (CUD, n = 57) with multi-domain cognitive measures and structural, diffusion, and resting state brain-imaging phenotypes. Post hoc model evidence was computed with Bayes factors (BF) and posterior probabilities of association (PPA) to account for multiple testing. General cognitive functioning, verbal intelligence, verbal memory, working memory, and motor speed were lower in the CUD group compared with nonusers (p \u3c .011; 1.9 \u3c BF \u3c 3,217). CUD was associated with altered functional connectivity in a network comprising the motor-hand region in the superior parietal gyri and the anterior insula (p \u3c .04). These differences were not explained by alcohol, other drug use, or education. No associations with CUD were observed in cortical thickness, cortical surface area, subcortical or cerebellar volumes (0.12 \u3c BF \u3c 1.5), or graph-theoretical metrics of resting state connectivity (PPA \u3c 0.01). In a large sample collected irrespective of cannabis used to minimize recruitment bias, we confirm the literature on poorer cognitive functioning in CUD, and an absence of volumetric brain differences between CUD and non-CUD. We did not find evidence for or against a disruption of structural connectivity, whereas we did find localized resting state functional dysconnectivity in CUD. There was sufficient proof, however, that organization of functional connectivity as determined via graph metrics does not differ between CUD and non-user group

Quantitative MRI brain in congenital adrenal hyperplasia: in vivo assessment of the cognitive and structural impact of steroid hormones

Abstract Context Brain white matter hyper-intensities are seen on routine clinical imaging in 46% of adults with congenital adrenal hyperplasia (CAH). The extent and functional relevance of these abnormalities have not been studied using quantitative MRI analysis. Objective To examine white matter microstructure, neural volumes and CNS metabolites in CAH due to 21-hydroxylase deficiency (21OHD) and to determine whether identified abnormalities are associated with cognition, glucocorticoid and androgen exposure. Design, setting and participants A cross-sectional study at a tertiary hospital including 19 females (18-50 years) with 21OHD and 19 age-matched healthy females. Main outcome measure Recruits underwent cognitive assessment and brain imaging including; diffusion weighted imaging of white matter, T1-weighted volumetry and magnetic resonance spectroscopy for neural metabolites. We evaluated white matter microstructure using tract-based spatial statistics. We compared cognitive scores, neural volumes and metabolites between groups and relationships between glucocorticoid exposure, MRI and neurologic outcomes. Results Patients with 21OHD had widespread reductions in white matter structural integrity, reduced volumes of right hippocampus, bilateral thalami, cerebellum and brainstem, and reduced mesial temporal lobe total choline content. Working memory, processing speed, and digit span and matrix reasoning scores were reduced in patients with 21OHD, despite similar education and intelligence to controls. 21OHD individuals exposed to higher glucocorticoid doses had greater abnormalities in white matter microstructure and cognitive performance. Conclusion For the first time we demonstrate that 21OHD and current glucocorticoid replacement regimens have a profound impact on brain morphology and function. If reversible, these CNS markers represent a potential target for treatment

Developing inclusive practice in Scotland: the National Framework for Inclusion

Introduction and context Developing the Framework Principles of the revised National Framework for Inclusion Reflections on the collaborative processes in developing the Framework Using the Framework Conclusion References Full Article Figures & data References Citations Metrics Reprints & Permissions PDF Abstract This paper reports on the collaborative development of a ‘National Framework for Inclusion’ under the auspices of the Scottish Teacher Education Committee by a working party representing each of the Scottish Universities providing initial teacher education. Recent research, international legislation and Scottish education policy have refocused the notion of ‘special educational needs’ based on ideas of individual deficit to support and provision for all learners. As teachers are therefore charged with responsibility for an increasingly diverse population of learners, the National Framework of Inclusion was developed to support both pre-service and qualified teachers to work inclusively to provide fair and meaningful experiences for all learners. The paper examines the underpinning principles of the Framework, describes the collaborative process of its development and provides one innovative example of its use