Thiol isomerases orchestrate thrombosis and haemostasis

Since protein disulphide isomerase (PDI) was first described in 1963, researchers have shown conclusively that PDI and sibling proteins are quintessential for thrombus formation. PDI, ERp5, ERp57 and ERp72, which in most cells are located in the endoplasmic reticulum and function to assist the folding of nascent protein, are released from platelets and vascular cells and interact with integrin αIIbβ3 on the outer surface of platelets. At the cell surface they continue to influence protein folding and function, propagating thrombosis and maintaining haemostasis. TMX1, which is a transmembrane thiol isomerase, is the first family member shown to negatively regulate platelets known to date. Targets of thiol isomerases have been indentified including integrin α2β1, Von Willebrand Factor (VWF), GpIbα, Nox-1, Nox-2 and tissue factor, all of which are pro-thrombotic, and several of which are on the cell surface. In spite of this, PDI can paradoxically catalyse the delivery of nitric oxide to platelets, which inhibits their function and decreases thrombus formation. Although the overall effect of PDI is to positively regulate platelet activation, it is still unclear how thiol isomerases function in pro-thrombotic states, such as obesity, diabetes and cancer. In parallel, there has been a surge in the development of novel thiol isomerase inhibitors, which display selectivity, potency and modulate thrombosis and haemostasis. The availability of selective thiol isomerase inhibitors has culminated in clinical trials with promising outcomes for the prevention of cancer-associated thrombosis. Altogether, thiol isomerases are perceived as an orchestrating force that regulates thrombus development. In the current review we will explore the history of PDI in cardiovascular biology, detail known mechanisms of action and summarise known thiol isomerase inhibitors

Socioeconomic factors associated with reports of domestic violence in large Brazilian cities

Background: Domestic violence is a traumatic experience that can lead to physical consequences, mental disorders and financial damage. Over 18 cases per 100,000 inhabitants were reported in Brazil between 2013 and 2014. The ministry of health poses a mandatory notification of all cases of domestic violence, which is essential, bearing in mind its systemic relation to various social issues and the extensive regional differences and high socioeconomic inequalities present in Brazil. Aim: To analyze the characteristics of the notification rates of domestic violence and investigate the correlation of these with health and socioeconomic characteristics of large Brazilian cities. Methods: Retrospective data on notifications of domestic violence was collected from the National Information System for Notifiable Diseases for Brazil, 2017. Dependent variables were collected from the Brazilian Institute of Geography and Statistics and Ministry of Citizenship. Inclusion criteria were: cities larger than 100.000 habitants and that had at least 20 reports, totaling 68.313 reports in 259 cities. These were stratified by age, race and sex of victim, type of violence used, violence perpetrator, place of occurrence and means of aggression. Proportional number of notified cases was calculated for each city to expose different characteristics of reports. A multiple linear regression model was used to investigate the correlation between report rates and different socioeconomic and health variables. Results: The analysis showed a high proportion of repeated violence, use of body strength and over 50% were perpetrated by a partner or boyfriend. Report rates were higher for women, black individuals and children under 4, highlighting subgroups of the population that were more vulnerable. Indeed, these groups were correlated differently with socioeconomic variables. Poverty, assessed as Bolsa Família investment, was correlated with domestic violence report rates across vulnerable groups. Conclusion: The study showed that black women and children are more vulnerable to domestic violence, highlighting deleterious effects of patriarchy and structural racism within Brazilian society. Altogether, we suggest that reducing poverty, patriarchy and structural racism could lead to fewer cases of domestic violence

Platelet-derived extracellular vesicles express NADPH oxidase-1 (Nox-1), generate superoxide and modulate platelet function

Background: Platelets release platelet-derived extracellular vesicles (PDEVs) upon activation – in a process that is regulated by generation of reactive oxygen species (ROS). Platelet NADPH oxidase-1 (Nox-1) contributes to ROS generation and thrombus formation downstream of the collagen receptor GPVI. Objectives: We aimed to investigate whether PDEVs contain Nox-1 and whether this is relevant for PDEV-induced platelet activation. Methods: PDEVs were isolated through serial centrifugation after platelet activation with thrombin receptor agonist TRAP-6 (activated PDEVs) or in the absence of agonist (resting PDEVs). The physical properties of PDEVs were analysed through nanoparticle tracking analysis. Nox-1 levels, fibrinogen binding and P-selectin exposure were measured using flow cytometry, and protein levels quantified by immunoblot analysis. ROS were quantified using DCF fluorescence and electron paramagnetic resonance. Results: Nox-1 was found to be increased on the platelet outer membrane upon activation and was found to be present in PDEVs. PDEVs induced platelet activation, while co-addition of GPVI agonist collagen-related peptide (CRP) did not potentiate this response. PDEVs were shown to be able to generate superoxide in a process at least partially mediated by Nox- 1, while Nox-1 inhibition with ML171 (also known as 2-APT) did not influence PDEV production. Finally, inhibition of Nox-1 abrogated PDEV-mediated platelet activation. Conclusions: PDEVs are able to generate superoxide, bind to and activate platelets in a process mediated by Nox-1. These data provide novel mechanisms by which Nox-1 potentiates platelet responses, thus proposing Nox-1 inhibition as a feasible strategy to treat and prevent thrombotic diseases

Protein disulphide isomerase and NADPH oxidase 1 cooperate to control platelet function and are associated with cardiometabolic disease risk factors

Background: Protein disulphide isomerase (PDI) and NADPH oxidase 1 (Nox- 1) regulate platelet function and reactive oxygen species (ROS) generation, suggesting potentially interdependent roles. Increased platelet reactivity and ROS production have been correlated with cardiometabolic disease risk factors. Objectives: To establish whether PDI and Nox-1 cooperate to control platelet function. Methods: Immunofluo- rescence microscopy was utilised to determine expression and localisation of PDI and Nox-1. Platelet aggregation, fibrinogen binding, P-selectin exposure, spreading and cal- cium mobilization were measured as markers of platelet function. A cross-sectional population study (n=136) was conducted to assess the relationship between platelet PDI and Nox-1 levels and cardiometabolic risk factors. Results: PDI and Nox-1 co-localized upon activation induced by the collagen receptor GPVI. Co-inhibition of PDI and Nox-1 led to additive inhibition of GPVI-mediated platelet aggregation, activation and calcium flux. This was confirmed in murine Nox-1-/- platelets treated with PDI inhibitor be- pristat, without affecting bleeding. PDI and Nox-1 together contributed to GPVI signal- ling that involved the phosphorylation of p38 MAPK, p47phox, PKC and Akt. Platelet PDI and Nox-1 levels were upregulated in obesity, with platelet Nox-1 also elevated in hypertensive individuals. Conclusions: We show that PDI and Nox-1 cooperate to con- trol platelet function and are associated with cardiometabolic risk factors