A model for the conformational activation of the structurally quiescent metalloprotease ADAMTS13 by Von Willebrand factor

Blood loss is prevented by the multi-domain glycoprotein von Willebrand factor (VWF), which binds exposed collagen at damaged vessels and captures platelets. VWF is regulated by the metalloprotease ADAMTS13, which, in turn, is conformationally activated by VWF. To delineate the structural requirements for VWF-mediated conformational activation of ADAMTS13, we performed binding and functional studies with a panel of truncated ADAMTS13 variants. We demonstrate that both the isolated CUB1 and CUB2 domains in ADAMTS13 bind to the spacer domain exosite of a truncated ADAMTS13 variant, MDTCS (KD of 135 ± 10.1 nM and 86.9 ± 9.0 nM, respectively). However, only the CUB1 domain inhibited proteolytic activity of MDTCS. Moreover, ADAMTS13∆CUB2, unlike ADAMTS13∆CUB1-2, exhibited activity similar to wild-type ADAMTS13 and could be activated by VWF D4-CK. The CUB2 domain is therefore not essential for maintaining the inactive conformation of ADAMTS13. Both CUB domains could bind to the VWF D4-CK domain fragment (KD of 53.7± 2.1 nM and 84.3 ± 2.0 nM, respectively). However, deletion of both CUB domains did not prevent VWF D4-CK binding, suggesting that competition for CUB-domain binding to the spacer domain is not the dominant mechanism behind the conformational activation. ADAMTS13∆TSP8-CUB2 could no longer bind to VWF D4-CK, and deletion of TSP8 abrogated ADAMTS13 conformational activation. These findings support an ADAMTS13-activation model in which VWF D4-CK engages the TSP8-CUB2 domains, inducing the conformational change that disrupts the CUB1-spacer domain interaction and thereby activates ADAMTS13

Evaluation of community perspectives on national health insurance policy to health service delivery in Ghana

The paper aims to examine National Health Insurance Scheme (NHIS) policy from the perspective of local communities. Qualitative data from nine key informant interviews and seventy-two community respondents in nine focus group discussions (FGD) were conducted. The FGDs took place in seven different communities in seven regions of Ghana. The data were analysed using thematic network approach. The findings noted a disconnection between NHIS policy and community life. More so, the findings showed an explicit link between equity and access with its impacts on health outcomes. The qualitative results indicated lack of voice and understanding as features limiting people from accessing the NHIS facilities. Understanding of equity as a key theme revealed: first, lack of stakeholder engagement and consultation or participation in the NHIS decision making process. Second it was established that while the policy indicated a will to include all the core poor for an equitable NHIS, there is lack of willingness to implement this aim fully. Finally, despite being a pro- poor intervention by name, practical management of the NHIS is transmitted down vertical silos from the national level, with the lack of joined-up government at the centre undermining local partnerships. Thus, not only are national expectations being dashed locally, local expectations are dashed nationally. The paper proposes that community viewpoints should be given higher priority given that NHIS has since its inception been associated with medical treatments and biomedical paradigm. Promoting community participation, understanding and voices should be recognised to shape the future NHIS policy and practice

Exploring community perspectives on national health insurance policy in Ghana

Even though a number of studies have examined the implementation of National Health Insurance Scheme (NHIS) in Ghana, its equity and access significance in relation to community perspective has not been assessed nationally. We looked at the NHIS from the perspective of local communities in focus groups discussions (FGDs) and key informant interviews. Qualitative data from nine key informant interviews and seventy-two community respondents in nine focus group discussions were collected. The focus groups took place in seven different communities purposely selected. The data were analysed using thematic network approach. The key informants and focus groups were analysed together. The findings noted a disconnection between NHIS policy and community life. More so, the findings showed an explicit link between equity and access with its impacts on health outcomes. The qualitative results noted lack of voice and understanding as features limiting people from accessing the NHIS facilities. Understanding of equity as a key theme revealed: first, lack of stakeholder engagement and consultation or participation in the NHIS decision making process. Second, the NHIS displayed an overreliance on healthcare and an emphasis on a biomedical approach. Finally, despite being a pro- poor intervention by name, practical management of the NHIS is transmitted down vertical silos from the national level, with the lack of joined-up government at the centre undermining local partnerships. Thus, not only are national expectations being dashed locally, local expectations are dashed nationally. We proposed that community viewpoints should be given higher priority given that NHIS has since its inception been associated with medical treatments and biomedical paradigm. Promoting community participation, understanding and voices should be recognised to shape the future NHIS policy and practice

Conformational activation of ADAMTS13

A disintegrin and metalloprotease with thrombospondin motifs 13 (ADAMTS13) is a metalloprotease that regulates von Willebrand factor (VWF) function. ADAMTS13-mediated proteolysis is determined by conformational changes in VWF, but also may depend on its own conformational activation. Kinetic analysis of WT ADAMTS13 revealed ∼2.5-fold reduced activity compared with ADAMTS13 lacking its C-terminal tail (MDTCS) or its CUB1-2 domains (WTΔCUB1-2), suggesting that the CUB domains naturally limit ADAMTS13 function. Consistent with this suggestion, WT ADAMTS13 activity was enhanced ∼2.5-fold by preincubation with either an anti-CUB mAb (20E9) or VWF D4CK (the natural binding partner for the CUB domains). Furthermore, the isolated CUB1-2 domains not only bound MDTCS, but also inhibited activity by up to 2.5-fold. Interestingly, a gain-of-function (GoF) ADAMTS13 spacer domain variant (R568K/F592Y/R660K/Y661F/Y665F) was ∼2.5-fold more active than WT ADAMTS13, but could not be further activated by 20E9 mAb or VWF D4CK and was unable to bind or to be inhibited by the CUB1-2 domains, suggesting that the inhibitory effects of the CUB domains involve an interaction with the spacer domain that is disrupted in GoF ADAMTS13. Electron microscopy demonstrated a “closed” conformation of WT ADAMTS13 and suggested a more “open” conformation for GoF ADAMTS13. The cryptic spacer domain epitope revealed by conformational unfolding also represents the core antigenic target for autoantibodies in thrombotic thrombocytopenic purpura. We propose that ADAMTS13 circulates in a closed conformation, which is maintained by a CUB–spacer domain binding interaction. ADAMTS13 becomes conformationally activated on demand through interaction of its C-terminal CUB domains with VWF, making it susceptible to immune recognition

Understanding qualitative and community indicators of poverty for national health insurance scheme exemptions in Ghana

The exemptions of the National Health Insurance Scheme (NHIS) for poor people in Ghana have not been sufficiently explored. Using a qualitative approach that involved gathering and analysing viewpoints from the community, this paper investigates the factors that are used to determine NHIS exemptions in Ghana. Focus group discussions (FGDs) and interviews of key informants (KIs) were conducted during the period (August 2015 to August 2016) within the Ashanti and Greater Accra regions in Ghana. Nine FGDs were conducted in nine different communities with 72 respondents. Nine KIs, including local and national policy-makers, civil servants and local community members were also interviewed. A sampling method was adopted to capture a range of understandings of community indicators of poverty for NHIS exemptions. Community perceptions of the indicators of poverty included interconnected themes of income, employment, education, assets, and social marginalisation. The findings highlight contextual and community descriptions associated with exemptions, of which poverty is a predictor, and discovered that to identify the indigent for exemptions based only on income and quantitative measures is not enough. The results are significant for a number of reasons including poor fit with official measures and the neglect of lay perspectives. For practice implications, our findings show that communities should be principal stakeholders for describing the disadvantaged groups that require exemptions

Enhanced activity of an ADAMTS-13 variant (R568K/F592Y/R660K/Y661F/Y665F) against platelet agglutination in vitro and in a murine model of acute ischemic stroke.

Essentials ADAMTS13 requires a substrate-induced conformational change to attain full activity in vitro. The efficacy of wild type ADAMTS13 in models of thrombosis/stroke may be enhanced by pre-activation. A pre-activated ADAMTS13 variant exhibits enhanced proteolysis of platelet agglutinates. This ADAMTS13 variant is protective in a murine model of stroke at a lower dose than WT ADAMTS13. SUMMARY: Background ADAMTS-13 circulates in a closed conformation, only achieving full proteolytic activity against von Willebrand factor (VWF) following a substrate-induced conformational change. A gain-of-function (GoF) ADAMTS-13 variant (R568K/F592Y/R660K/Y661F/Y665F) is conformationally preactivated. Objectives To establish how the hyperactivity of GoF ADAMTS-13 is manifested in experimental models mimicking the occlusive arterial thrombi present in acute ischemic stroke. Methods The ability of GoF ADAMTS-13 to dissolve VWF-platelet agglutinates was examined with an assay of ristocetin-induced platelet agglutination and in parallel-flow models of arterial thrombosis. A murine model of focal ischemia was used to assess the thrombolytic potential of GoF ADAMTS-13. Results Wild-type (WT) ADAMTS-13 required conformational activation to attain full activity against VWF-mediated platelet capture under flow. In this assay, GoF ADAMTS-13 had an EC50 value more than five-fold lower than that of WT ADAMTS-13 (0.73 ± 0.21 nm and 3.81 ± 0.97 nm, respectively). The proteolytic activity of GoF ADAMTS-13 against preformed platelet agglutinates under flow was enhanced more than four-fold as compared with WT ADAMTS-13 (EC50 values of 2.5 ± 1.1 nm and 10.2 ± 5.6 nm, respectively). In a murine stroke model, GoF ADAMTS-13 restored cerebral blood flow at a lower dose than WT ADAMTS-13, and partially retained the ability to recanalize vessels when administration was delayed by 1 h. Conclusions The limited proteolytic activity of WT ADAMTS-13 in in vitro models of arterial thrombosis suggests an in vivo requirement for conformational activation. The enhanced activity of the GoF ADAMTS-13 variant translates to a more pronounced protective effect in experimental stroke

Knowledge and use of emergency contraception among women in the Western Cape province of South Africa: a cross-sectional study

BACKGROUND: Emergency contraception (EC) is widely available free of charge at public sector clinics in South Africa. At the same time, rates of teenage and unintended pregnancy in South Africa remain high, and there are few data on knowledge of EC in the general population in South Africa, as in other resource-limited settings. METHODS: We conducted a cross-sectional, interviewer-administered survey among 831 sexually active women at 26 randomly selected public sector clinics in the Western Cape province. RESULTS: Overall, 30% of the women had ever heard of EC when asked directly, after the method was described to them. Only 15% mentioned EC by name or description spontaneously. Knowledge of EC was independently associated with higher education, being married, and living in an urban setting. Four percent of women had ever used EC. DISCUSSION: These data suggest that knowledge of EC in this setting is more common among women of higher socioeconomic status living in urban areas. For EC to play a role in decreasing unintended pregnancy in South Africa, specific interventions are necessary to increase knowledge of the method, where to get it, and the appropriate time interval for its use before the need for EC arises. Future health promotion campaigns should target rural and low socioeconomic status communities

Surgical and medical second trimester abortion in South Africa: A cross-sectional study

<p>Abstract</p> <p>Background</p> <p>A high percentage of abortions performed in South Africa are in the second trimester. However, little research focuses on women's experiences seeking second trimester abortion or the efficacy and safety of these services.</p> <p>The objectives are to document clinical and acceptability outcomes of second trimester medical and surgical abortion as performed at public hospitals in the Western Cape Province.</p> <p>Methods</p> <p>We performed a cross-sectional study of women undergoing abortion at 12.1-20.9 weeks at five hospitals in Western Cape Province, South Africa in 2008. Two hundred and twenty women underwent D&E with misoprostol cervical priming, and 84 underwent induction with misoprostol alone. Information was obtained about the procedure and immediate complications, and women were interviewed after recovery.</p> <p>Results</p> <p>Median gestational age at abortion was earlier for D&E clients compared to induction (16.0 weeks vs. 18.1 weeks, p < 0.001). D&E clients reported shorter intervals between first clinic visit and abortion (median 17 vs. 30 days, p < 0.001). D&E was more effective than induction (99.5% vs. 50.0% of cases completed on-site without unplanned surgical procedure, p < 0.001). Although immediate complications were similar (43.8% D&E vs. 52.4% induction), all three major complications occurred with induction. Early fetal expulsion occurred in 43.3% of D&E cases. While D&E clients reported higher pain levels and emotional discomfort, most women were satisfied with their experience.</p> <p>Conclusions</p> <p>As currently performed in South Africa, second trimester abortions by D&E were more effective than induction procedures, required shorter hospital stay, had fewer major immediate complications and were associated with shorter delays accessing care. Both services can be improved by implementing evidence-based protocols.</p

How students perceive medical competences: a cross-cultural study between the Medical Course in Portugal and African Portuguese Speaking Countries

<p>Abstract</p> <p>Background</p> <p>A global effort has been made in the last years to establish a set of core competences that define the essential professional competence of a physician. Regardless of the environment, culture or medical education conditions, a set of core competences is required for medical practice worldwide. Evaluation of educational program is always needed to assure the best training for medical students and ultimately best care for patients. The aim of this study was to determine in what extent medical students in Portugal and Portuguese speaking African countries, felt they have acquired the core competences to start their clinical practice. For this reason, it was created a measurement tool to evaluate self-perceived competences, in different domains, across Portuguese and Portuguese-speaking African medical schools.</p> <p>Methods</p> <p>The information was collected through a questionnaire that defines the knowledge, attitudes and skills that future doctors should acquire. The Cronbach's Alpha and Principal Components Analysis (PCA) were used to evaluate the reliability of the questionnaire. In order to remove possible confounding effect, individual scores were standardized by country.</p> <p>Results</p> <p>The order of the domain's scores was similar between countries. After standardization, Personal Attitudes and Professional Behavior showed median scores above the country global median and Knowledge alone showed median score below the country global median. In Portugal, Clinical Skills showed score below the global median. In Angola, Clinical Skills and General Skills showed a similar result. There were only significant differences between countries in Personal Attitudes (p < 0.001) and Professional Behavior (p = 0.043).</p> <p>Conclusions</p> <p>The reliability of the instrument in Portuguese and Portuguese-speaking African medical schools was confirmed. Students have perceived their level of competence in personal attitudes in a high level and in opposite, knowledge and clinical skills with some weaknesses.</p

Chemical and environmental vector control as a contribution to the elimination of visceral leishmaniasis on the Indian subcontinent: cluster randomized controlled trials in Bangladesh, India and Nepal

<p>Abstract</p> <p>Background</p> <p>Bangladesh, India and Nepal are working towards the elimination of visceral leishmaniasis (VL) by 2015. In 2005 the World Health Organization/Training in Tropical Diseases launched an implementation research programme to support integrated vector management for the elimination of VL from Bangladesh, India and Nepal. The programme is conducted in different phases, from proof-of-concept to scaling up intervention. This study was designed in order to evaluate the efficacy of the three different interventions for VL vector management: indoor residual spraying (IRS); long-lasting insecticide treated nets (LLIN); and environmental modification (EVM) through plastering of walls with lime or mud.</p> <p>Methods</p> <p>Using a cluster randomized controlled trial we compared three vector control interventions with a control arm in 96 clusters (hamlets or neighbourhoods) in each of the 4 study sites: Bangladesh (one), India (one) and Nepal (two). In each site four villages with high reported VL incidences were included. In each village six clusters and in each cluster five households were randomly selected for sand fly collection on two consecutive nights. Control and intervention clusters were matched with average pre-intervention vector densities.</p> <p>In each site six clusters were randomly assigned to each of the following interventions: indoor residual spraying (IRS); long-lasting insecticide treated nets (LLIN); environmental management (EVM) or control. All the houses (50-100) in each intervention cluster underwent the intervention measures. A reduction of intra-domestic sand fly densities measured in the study households by overnight US Centres for Disease Prevention and Control light trap captures (that is the number of sand flies per trap per night) was the main outcome measure.</p> <p>Results</p> <p>IRS, and to a lesser extent EVM and LLINs, significantly reduced sand fly densities for at least 5 months in the study households irrespective of type of walls or whether or not people shared their house with cattle. IRS was effective in all sites but LLINs were only effective in Bangladesh and India. Mud plastering did not reduce sand fly density (Bangladesh study); lime plastering in India and one Nepali site, resulted in a significant reduction of sand fly density but not in the second Nepali site.</p> <p>Conclusion</p> <p>Sand fly control can contribute to the regional VL elimination programme; IRS should be strengthened in India and Nepal but in Bangladesh, where vector control has largely been abandoned during the last decades, the insecticide treatment of existing bed nets (coverage above 90% in VL endemic districts) could bring about an immediate reduction of vector populations; operational research to inform policy makers about the efficacious options for VL vector control and programme performance should be strengthened in the three countries.</p