Confidence in providing methadone maintenance treatment of primary care providers in Vietnam.

BACKGROUND: Delivering methadone treatment in community health facilities by primary care providers is a task-shifting strategy to expand access to drug use treatment, especially in rural mountainous areas. This study aims to investigate factors related to confidence in providing methadone treatment among primary care providers in Vietnam to inform good practice development. METHODS: We conducted a cross-sectional survey with 276 primary care providers who were physicians, physician assistants, nurses, pharmacists or dispensing staff from 67 communes in a mountainous province in Northern Vietnam. Using self-report scales, we measured providers' confidence in providing methadone treatment, beliefs in harm reduction, perceived work-related support, perceived stigma and risk in working with drug-using patients, and empathy towards this population. We used multiple linear regression analyses to explore factors associated with providers' confidence in providing methadone treatment in the whole sample and to compare two groups of providers who did and did not have experience providing methadone. Potential associated factors were measured at facility and provider levels. RESULT: 114 (41.3%) participants had previously experience in providing methadone treatment. Providers with methadone treatment experiences had higher confidence in and more accurate knowledge of methadone treatment, perceived less stigma of working with drug-using patients, and reported more work-related support than those without experiences. Higher medical education is associated with lower confidence in providing methadone treatment among providers without methadone experiences, but higher confidence among providers with methadone experiences. Better methadone knowledge was associated with greater confidence in providing methadone treatment among inexperienced providers but not among those with experiences. Receiving work-related support was associated with greater confidence in providing treatment in both groups, regardless of their past methadone experiences. CONCLUSION: In rural provinces where methadone treatment has been expanded to primary care clinics, interventions to improve primary care providers' confidence should benefit professionals with diverse experiences in providing methadone treatment. Continued training and support at work for providers is essential to ensuring quality in decentralized methadone treatment.

On a sugar high: Role of O-GlcNAcylation in cancer.

Recent advances in the understanding of the molecular mechanisms underlying cancer progression have led to the development of novel therapeutic targeting strategies. Aberrant glycosylation patterns and their implication in cancer have gained increasing attention as potential targets due to the critical role of glycosylation in regulating tumor-specific pathways that contribute to cancer cell survival, proliferation, and progression. A special type of glycosylation that has been gaining momentum in cancer research is the modification of nuclear, cytoplasmic, and mitochondrial proteins, termed O-GlcNAcylation. This protein modification is catalyzed by an enzyme called O-GlcNAc transferase (OGT), which uses the final product of the Hexosamine Biosynthetic Pathway (HBP) to connect altered nutrient availability to changes in cellular signaling that contribute to multiple aspects of tumor progression. Both O-GlcNAc and its enzyme OGT are highly elevated in cancer and fulfill the crucial role in regulating many hallmarks of cancer. In this review, we present and discuss the latest findings elucidating the involvement of OGT and O-GlcNAc in cancer.

Kruppel-like factor 8 regulates triple negative breast cancer stem cell-like activity.

Introduction: Breast tumor development is regulated by a sub-population of breast cancer cells, termed cancer stem-like cells (CSC), which are capable of self-renewing and differentiating, and are involved in promoting breast cancer invasion, metastasis, drug resistance and relapse. CSCs are highly adaptable, capable of reprogramming their own metabolism and signaling activity in response to stimuli within the tumor microenvironment. Recently, the nutrient sensor O-GlcNAc transferase (OGT) and O-GlcNAcylation was shown to be enriched in CSC populations, where it promotes the stemness and tumorigenesis of breast cancer cells in vitro and in vivo. This enrichment was associated with upregulation of the transcription factor Kruppel-like-factor 8 (KLF8) suggesting a potential role of KLF8 in regulating CSCs properties. Methods: Triple-negative breast cancer cells were genetically modified to generate KLF8 overexpressing or KLF8 knock-down cells. Cancer cells, control or with altered KLF8 expression were analyzed to assess mammosphere formation efficiency, CSCs frequency and expression of CSCs factors. Tumor growth in vivo of control or KLF8 knock-down cells was assessed by fat-pad injection of these cell in immunocompromised mice. Results: Here, we show that KLF8 is required and sufficient for regulating CSC phenotypes and regulating transcription factors SOX2, NANOG, OCT4 and c-MYC. KLF8 levels are associated with chemoresistance in triple negative breast cancer patients and overexpression in breast cancer cells increased paclitaxel resistance. KLF8 and OGT co-regulate each other to form a feed-forward loop to promote CSCs phenotype and mammosphere formation of breast cancer cells. Discussion: These results suggest a critical role of KLF8 and OGT in promoting CSCs and cancer progression, that may serve as potential targets for developing strategy to target CSCs specifically.

Acceptability and feasibility of long-acting injectable antiretroviral therapy for HIV-infected persons who inject drugs in Vietnam: A qualitative study.

Background: In Vietnam, HIV prevalence among people who inject drugs (PWID) is several times higher than in the general population (15% versus 0.3%). PWID also experience higher rates of HIV-related mortality, driven by poor antiretroviral therapy (ART) adherence. Long-acting injectable ART (LAI) is a compelling opportunity to improve treatment outcomes, but acceptability and feasibility among HIV-infected PWID remains unexplored. Methods: We conducted key informant in-depth interviews in Hanoi, Vietnam (February-November 2021). Participants were purposively sampled and included policymakers, ART clinic staff, and HIV-infected PWID. We applied the Consolidated Framework for Implementation Research to guide study design and analysis, using thematic coding to develop and iteratively refine a codebook and characterize barriers and facilitators to LAI implementation. Findings: We interviewed 38 key stakeholders: 19 PWID, 14 ART clinic staff, and five policymakers. Participants were enthusiastic about LAI convenience, highlighting less frequent and more discreet dosing. However, contrasting providers, several policymakers suggested LAI was not needed given perceived exceptional oral ART outcomes and rare viral failure among PWID. Policymakers also criticized strategies prioritizing PWID for LAI, emphasizing equity, whereas providers identified PWID as an ideal population for LAI given adherence challenges. LAI complexity, including storage and administration logistics, were deemed surmountable with training and resources. Finally, providers and policymakers acknowledged that adding LAI to drug formularies was key, but an onerous process. Interpretation: Although anticipated to be resource-intensive, LAI was a welcome addition for interviewed stakeholders and likely an acceptable alternative to oral ART among PWID living with HIV in Vietnam. Despite enthusiasm among PWID and providers that LAI could improve viral outcomes, some policymakers-whose buy-in is critical to LAI implementation-opposed strategies that preferentially distributed LAI to PWID, highlighting values of equity and revealing differences in perceived HIV outcomes among PWID. Results provide a vital foundation for developing LAI implementation strategies. Funding: Supported by National Institutes of Health.

Role of O-GlcNAcylation on cancer stem cells: Connecting nutrient sensing to cell plasticity.

Tumor growth and metastasis can be promoted by a small sub-population of cancer cells, termed cancer stem-like cells (CSCs). While CSCs possess capability in self-renewing and differentiating, the hierarchy of CSCs during tumor growth is highly plastic. This plasticity in CSCs fate and function can be regulated by signals from the tumor microenvironment. One emerging pathway in CSCs that connects the alteration in microenvironment and signaling network in cancer cells is the hexosamine biosynthetic pathway (HBP). The final product of HBP, UDP-N-acetylglucosamine (UDP-GlcNAc), is utilized for glycosylating of membrane and secreted proteins, but also nuclear and cytoplasmic proteins by the post-translational modification O-GlcNAcylation. O-GlcNAcylation and its enzyme, O-GlcNAc transferase (OGT), are upregulated in nearly all cancers and been linked to regulate many cancer cell phenotypes. Recent studies have begun to connect OGT and O-GlcNAcylation to regulation of CSCs. In this review, we will discuss the emerging role of OGT and O-GlcNAcylation in regulating fate and plasticity of CSCs, as well as the potential in targeting OGT/O-GlcNAcylation in CSCs.

Chronic Immune Platelet Activation Is Followed by Platelet Refractoriness and Impaired Contractility.

Autoimmune diseases, including systemic lupus erythematosus (SLE), have a high risk of thrombotic and hemorrhagic complications associated with altered platelet functionality. We studied platelets from the blood of SLE patients and their reactivity. The surface expression of phosphatidylserine, P-selectin, and active integrin αIIbß3 were measured using flow cytometry before and after platelet stimulation. Soluble P-selectin was measured in plasma. The kinetics of platelet-driven clot contraction was studied, as well as scanning and transmission electron microscopy of unstimulated platelets. Elevated levels of membrane-associated phosphatidylserine and platelet-attached and soluble P-selectin correlated directly with the titers of IgG, anti-dsDNA-antibodies, and circulating immune complexes. Morphologically, platelets in SLE lost their resting discoid shape, formed membrane protrusions and aggregates, and had a rough plasma membrane. The signs of platelet activation were associated paradoxically with reduced reactivity to a physiological stimulus and impaired contractility that revealed platelet exhaustion and refractoriness. Platelet activation has multiple pro-coagulant effects, and the inability to fully contract (retract) blood clots can be either a hemorrhagic or pro-thrombotic mechanism related to altered clot permeability, sensitivity of clots to fibrinolysis, obstructiveness, and embologenicity. Therefore, chronic immune platelet activation followed by secondary platelet dysfunction comprise an understudied pathogenic mechanism that supports hemostatic disorders in autoimmune diseases, such as SLE.

Stigma as a barrier to integrated substance use and HIV care in Vietnam: A qualitative examination of patient and provider perspectives.

Integration of substance use disorder (SUD) treatment and HIV care can increase antiretroviral therapy coverage among people with opioid use disorder (OUD). However, implementation of integrated treatment models remains limited. Stigma towards people with OUD poses a barrier to initiation of, and adherence to, HIV treatment. We sought to understand the extent of stigma towards SUD and HIV among people with OUD in Vietnam, and the effect of stigma on integrated OUD and HIV treatment services utilization. Between 2013 and 2015, we conducted in-depth interviews with 43 patients and 43 providers at 7 methadone clinics and 8 HIV clinics across 4 provinces in Vietnam. We used thematic analysis with a mixed deductive and inductive approach at the semantic level to analyze key topics. Two main themes were identified: (1) Confidentiality concerns about HIV status make patients reluctant to receive integrated care at HIV clinics, given the requirements for daily buprenorphine dosing at HIV clinics. (2) Provider stigma existed mostly toward people with OUD and seemed to center on the belief that substance use causes a deterioration in one's morals, and was most frequently manifested in the form of providers' apprehensive approach towards patients. Concerns regarding stigmatization may cause patients to feel reluctant to receive treatment for both OUD and HIV at a single integrated clinic. Interventions to reduce stigma at the clinic and policy levels may thus serve to improve initiation of and adherence to integrated care.

Familial support in integrated treatment with antiretroviral therapy and medications for opioid use disorder in Vietnam: A qualitative study.

Background: Patients report that familial support can facilitate initiation and maintenance of antiretroviral therapy (ART) and medications for opioid use disorder (MOUD). However, providing such support can create pressure and additional burdens for families of people with opioid use disorder (OUD) and HIV. We examined perspectives of people with HIV receiving treatment for OUD in Vietnam and their family members. Methods: Between 2015 and 2018, we conducted face-to-face qualitative interviews with 44 patients and 30 of their family members in Hanoi, Vietnam. Participants were people living with HIV and OUD enrolled in the BRAVO study comparing HIV clinic-based buprenorphine with referral to methadone treatment at 4 HIV clinics and their immediate family members (spouses or parents). Interviews were professionally transcribed, coded in Vietnamese, and analyzed using a semantic, inductive approach to qualitative thematic analysis. Results: Family members of people with OUD and HIV in Vietnam reported financially and emotionally supporting MOUD initiation and maintenance as well as actively participating in treatment. Family members described the burdens of supporting patients during opioid use, including financial costs and secondary stigma. Conclusions: Describing the role of family support in the lives of people living with OUD and HIV in the context of Vietnam enriches our understanding of their experiences and will support future treatment efforts targeting the family unit.

O-GlcNAc transferase regulates glioblastoma acetate metabolism via regulation of CDK5-dependent ACSS2 phosphorylation.

Glioblastomas (GBMs) preferentially generate acetyl-CoA from acetate as a fuel source to promote tumor growth. O-GlcNAcylation has been shown to be elevated by increasing O-GlcNAc transferase (OGT) in many cancers and reduced O-GlcNAcylation can block cancer growth. Here, we identify a novel mechanism whereby OGT regulates acetate-dependent acetyl-CoA and lipid production by regulating phosphorylation of acetyl-CoA synthetase 2 (ACSS2) by cyclin-dependent kinase 5 (CDK5). OGT is required and sufficient for GBM cell growth and regulates acetate conversion to acetyl-CoA and lipids. Elevating O-GlcNAcylation in GBM cells increases phosphorylation of ACSS2 on Ser-267 in a CDK5-dependent manner. Importantly, we show that ACSS2 Ser-267 phosphorylation regulates its stability by reducing polyubiquitination and degradation. ACSS2 Ser-267 is critical for OGT-mediated GBM growth as overexpression of ACSS2 Ser-267 phospho-mimetic rescues growth in vitro and in vivo. Importantly, we show that pharmacologically targeting OGT and CDK5 reduces GBM growth ex vivo. Thus, the OGT/CDK5/ACSS2 pathway may be a way to target altered metabolic dependencies in brain tumors.

Family-based intervention for adolescents with substance use disorders in Vietnam.

BACKGROUND: Adolescent substance use is a leading risk factor of medical and social problems in adults. However, evidence-based interventions for substance use disorders (SUD) among youth in resource-limited countries are lacking. Treatnet Family (TF), developed by United Nations Office on Drugs and Crime (UNODC), aims to make youth SUD care more affordable and accessible in low- and middle-income countries. This study explores the suitability of TF in Vietnam. METHOD: Twenty interviews were conducted with eight adolescents and their family members who participated in TF, and four practitioners who delivered TF. Questions centred on their experiences with the intervention and suggestions for improvement. Thematic analysis was used to evaluate the data. RESULTS: All adolescents were male with an average age of 19.3. Seven of them had left school. Most caregivers were female. Both family members and adolescents expressed a great demand for support, and both groups appreciated the immediate improvement in parent-child communication. However, the impact of TF could be compromised due challenges in recruiting families, possibly arising from the novelty of a family-based intervention in Vietnam and drug-related stigma. The perception of drug use as an acute condition instead of a chronic disorder, and the lack of a continuing care system, also made it difficult to retain participants. CONCLUSION: Vietnamese adolescents with SUD and their family members were in great need of support and access to evidence-based interventions. Building a comprehensive, health-centred substance use disorder treatment and care system would enhance treatment impact.

What Shapes Late HIV Diagnosis in Vietnam? A Qualitative Investigation of Multilevel Factors.

Late HIV treatment remains a global public health issue despite significant efforts. To better understand what shapes this issue, we interviewed 36 Vietnamese ART-naive patients who came to HIV treatment in 2017. Half of them had intake CD4 counts fewer than 100 cells/mm3, the others had intake CD4 counts of 350 cells/mm3 and above. Late diagnosis was the reason of late treatment in our sample. Most late presenters were not members of the key populations at increased risk of HIV (e.g., people who inject drugs, commercial sex workers, and men who have sex with men). Individual-level factors included low risk appraisal, habit of self-medication, and fear of stigma. Network and structural-level factors included challenges to access quality health care, normalization of HIV testing in key populations and inconsistent provider-initiated HIV testing practices. Structural interventions coupled with existing key population-targeted strategies would improve the issue of late HIV diagnosis.

Effects of Hyperhomocysteinemia on the Platelet-Driven Contraction of Blood Clots.

Hyperhomocysteinemia (HHcy) is associated with thrombosis, but the mechanistic links between them are not understood. We studied effects of homocysteine (Hcy) on clot contraction in vitro and in a rat model of HHcy. Incubation of blood with exogenous Hcy for 1 min enhanced clot contraction, while 15-min incubation led to a dose-dependent suppression of contraction. These effects were likely due to direct Hcy-induced platelet activation followed by exhaustion, as revealed by an increase in fibrinogen-binding capacity and P-selectin expression determined by flow cytometry. In the blood of rats with HHcy, clot contraction was enhanced at moderately elevated Hcy levels (10-50 µM), while at higher Hcy levels (>50 µM), the onset of clot contraction was delayed. HHcy was associated with thrombocytosis combined with a reduced erythrocyte count and hypofibrinogenemia. These data suggest that in HHcy, platelets get activated directly and indirectly, leading to enhanced clot contraction that is facilitated by the reduced content and resilience of fibrin and erythrocytes in the clot. The excessive platelet activation can lead to exhaustion and impaired contractility, which makes clots larger and more obstructive. In conclusion, HHcy modulates blood clot contraction, which may comprise an underappreciated pro- or antithrombotic mechanism.

Psychosocial Interventions for Amphetamine Type Stimulant Use Disorder: An Overview of Systematic Reviews.

Introduction: Amphetamine-type stimulants (ATS) use is a global concern due to increased usage and the harm to physical, mental, and social well-being. The objective of this overview of systematic reviews is to summarise trial results of psychosocial interventions and describe their efficacy and safety. Methods: We searched seven bibliographic databases to November 2020 for systematic reviews examining ATS misuse treatment by psychosocial interventions. Given the apparent incompleteness of the included reviews, we undertook a supplemental meta-analysis of all eligible primary studies. Results: We included 11 systematic reviews of moderate to high quality and 39 primary studies which assessed the outcomes of psychosocial interventions on people who use ATS. The key findings include: (1) There were conflicting results about the effectiveness of psychosocial interventions among reviews, which may confuse decision-makers in selecting treatment. (2) In the supplemental meta-analysis, relative to usual care (only counselling or self-help materials), membership of a psychological intervention group was associated with an important reduction in drug usage [risk ratio (RR) 0.80, 95% CI: 0.75 to 0.85]. Patients in psychological interventions used injectables substantially less [odds ratio (OR) 0.35, 95% CI: 0.24 to 0.49]. The risk of unsafe sex in the psychosocial intervention group was lower than in the control group (RR 0.49, 95% CI: 0.34 to 0.71). The combination of therapies reduced 1.51 day using drugs in the preceding 30 days (95% CI: -2.36 to -0.67) compared to cognitive behavioural therapy intervention alone. (3) Compared to usual care, cognitive behavioural therapy was less likely to be retained at follow-up (RR 0.89, 95% CI: 0.82 to 0.97; high-quality evidence). However, the additional of contingency management strategy can make an important improvement upon retention (RR 1.42, 95%CI: 1.25 to 1.62). Authors' Conclusions: Integrated models are more effective than a single-treatment strategy. Comprehensive and sustained psychosocial interventions can help to reduce use of ATS and other drugs, risk behaviours and mental disorders, and significantly improve treatment adherence.

Substantial Need for Preexposure Prophylaxis Among Men Who Have Sex With Men in Hanoi, Vietnam.

ABSTRACT: We used data from the Health in Men-Hanoi cohort to determine the proportion of HIV-negative men who have sex with men with PrEP indications in Hanoi. Among 717 men who have sex with men, 537 (72.2% [66.6%-77.3%]) had ≥1 PrEP indication, signaling a substantial need for PrEP scale-up. Condomless anal intercourse was the most frequent indication (68.7% [60.3%-76.1%]), followed by previous/current sexually transmitted infection (59.4% [51.0%-67.2%]).

O-GlcNAc Transferase Regulates Cancer Stem-like Potential of Breast Cancer Cells.

Breast tumors are heterogeneous and composed of different subpopulation of cells, each with dynamic roles that can change with stage, site, and microenvironment. Cellular heterogeneity is, in part, due to cancer stem-like cells (CSC) that share properties with stem cells and are associated with treatment resistance. CSCs rewire metabolism to meet energy demands of increased growth and biosynthesis. O-GlcNAc transferase enzyme (OGT) uses UDP-GlcNAc as a substrate for adding O-GlcNAc moieties to nuclear and cytoplasmic proteins. OGT/O-GlcNAc levels are elevated in multiple cancers and reducing OGT in cancer cells blocks tumor growth. Here, we report that breast CSCs enriched in mammosphere cultures contain elevated OGT/O-GlcNAcylation. Inhibition of OGT genetically or pharmacologically reduced mammosphere forming efficiency, the CD44H/CD24L, NANOG+, and ALDH+ CSC population in breast cancer cells. Conversely, breast cancer cells overexpressing OGT increased mammosphere formation, CSC populations in vitro, and also increased tumor initiation and CSC frequency in vivo. Furthermore, OGT regulates expression of a number of epithelial-to-mesenchymal transition and CSC markers including CD44, NANOG, and c-Myc. In addition, we identify Krüppel-like factor 8 (KLF8) as a novel regulator of breast cancer mammosphere formation and a critical target of OGT in regulating CSCs. IMPLICATIONS: These findings demonstrate that OGT plays a key role in the regulation of breast CSCs in vitro and tumor initiation in vivo, in part, via regulation of KLF8, and thus inhibition of OGT may serve as a therapeutic strategy to regulate tumor-initiating activity.

In systemic lupus erythematosus anti-dsDNA antibodies can promote thrombosis through direct platelet activation.

Systemic lupus erythematosus (SLE) is associated with a high risk of venous and arterial thrombosis, not necessarily associated with prothrombotic antiphospholipid antibodies (Abs). Alternatively, thrombosis may be due to an increased titer of anti-dsDNA Abs that presumably promote thrombosis via direct platelet activation. Here, we investigated effects of purified anti-dsDNA Abs from the blood of SLE patients, alone or in a complex with dsDNA, on isolated normal human platelets. We showed that anti-dsDNA Abs and anti-dsDNA Ab/dsDNA complexes induced strong platelet activation assessed by enhanced P-selectin expression and dramatic morphological and ultrastructural changes. Electron microscopy revealed a significantly higher percentage of platelets that lost their discoid shape, formed multiple filopodia and had a shrunken body when treated with anti-dsDNA Abs or anti-dsDNA Ab/dsDNA complexes compared with control samples. In addition, these platelets activated with anti-dsDNA Ab/dsDNA complexes typically contained a reduced number of secretory α-granules that grouped in the middle and often merged into a solid electron dense area. Many activated platelets released plasma membrane-derived microvesicles and/or fell apart into subcellular cytoplasmic fragments. Confocal microscopy revealed that platelets treated with anti-dsDNA Ab/dsDNA complex had a heterogeneous distribution of septin2 compared with the homogeneous distribution in control platelets. Structural perturbations were concomitant with mitochondrial depolarization and a decreased content of platelet ATP, indicating energetic exhaustion. Most of the biochemical and morphological changes in platelets induced by anti-dsDNA Abs and anti-dsDNA Ab/dsDNA complexes were prevented by pre-treatment with a monoclonal mAb against FcγRIIA. The aggregate of data indicates that anti-dsDNA Abs alone or in a complex with dsDNA strongly affect platelets via the FcγRIIA receptor. The immune activation of platelets with antinuclear Abs may comprise a prothrombotic mechanism underlying a high risk of thrombotic complications in patients with SLE.

Differential sensitivity of various markers of platelet activation with adenosine diphosphate.

A number of techniques have been available to assess platelet activation, but their relative sensitivity is unknown and their usage is variable and not based on any rational criteria. Here, we compared the ability of several techniques based on morphological and biochemical markers to detect the first signs of ADP-induced platelet activation. Scanning electron microscopy of platelets was performed in parallel with flow cytometry to quantify the surface expression of P-selectin (marked by labeled anti-CD62P antibodies), active αIIbß3-intergrin (assessed by the binding of labeled fibrinogen) and phosphatidylserine (assessed by the binding of labeled Annexin V). When expressed as a fraction of activated platelets, shape changes were the most sensitive to a low ADP concentration compared to the biochemical markers in the following order of sensitivity: morphological changes>fibrinogen binding capacity>P-selectin expression> phosphatidylserine exposure. These results suggest the greater sensitivity of platelet microscopy and the importance of its combination with flow cytometry used to detect surface expression of the molecular markers of platelet activation.

Platelet factor 4-containing immune complexes induce platelet activation followed by calpain-dependent platelet death.

Heparin-induced thrombocytopenia (HIT) is a complication of heparin therapy sometimes associated with thrombosis. The hallmark of HIT is antibodies to the heparin/platelet factor 4 (PF4) complex that cause thrombocytopenia and thrombosis through platelet activation. Despite the clinical importance, the molecular mechanisms and late consequences of immune platelet activation are not fully understood. Here, we studied immediate and delayed effects of the complexes formed by human PF4 and HIT-like monoclonal mouse anti-human-PF4/heparin IgG antibodies (named KKO) on isolated human platelets in vitro. Direct platelet-activating effect of the KKO/PF4 complexes was corroborated by the overexpression of phosphatidylserine (PS) and P-selectin on the platelet surface. The immune platelet activation was accompanied by a decrease of the mitochondrial transmembrane potential (ΔΨm), concurrent with a significant gradual reduction of the ATP content in platelets, indicating disruption of energy metabolism. A combination of PS expression and mitochondrial depolarization induced by the PF4-containing immune complexes observed in a substantial fraction of platelets was considered as a sign of ongoing platelet death, as opposed to a subpopulation of activated live platelets with PS on the plasma membrane but normal ΔΨm. Both activated and dying platelets treated with KKO/PF4 formed procoagulant extracellular microvesicles bearing PS on their surface. Scanning and transmission electron microscopy revealed dramatic morphological changes of KKO/PF4-treated platelets, including their fragmentation, another indicator of cell death. Most of the effects of KKO/PF4 were prevented by an anti-FcγRII monoclonal antibody IV.3. The adverse functional and structural changes in platelets induced by the KKO/PF4 complexes were associated with strong time-dependent activation of calpain, but only trace cleavage of caspase 3. The results indicate that the pathogenic PF4-containing HIT-like immune complexes induce direct prothrombotic platelet activation via FcγRIIA receptors followed by non-apoptotic calpain-dependent death of platelets, which can be an important mechanism of thrombocytopenia during HIT development.

Struggling to achieve a 'normal life': A qualitative study of Vietnamese methadone patients.

BACKGROUND: Methadone maintenance treatment, initially introduced in Vietnam for HIV harm reduction, has marked a significant switch in the country's drug policy - from addiction as a moral issue to addiction as a brain disease. After the some initial outstanding achievements, the programme is facing a high dropout rate that threatens both goals of HIV prevention and drug treatment. This sociological study, as part of an HIV intervention research project, explores the challenges and opportunities that individuals who use drugs are faced with in relation to addiction treatment. METHODS: A qualitative study among drug users with and without methadone maintenance treatment experiences recruited by peer outreach workers. We conducted 58 in-depth interviews and 2 focus groups between 2016 and 2017. RESULTS: The start of treatment brought about significant feelings of success as heroin use was no longer compulsive. However, being in treatment programmes is also challenging with respect to continuing the recovery process. Barriers to retention include a popular fear of methadone as another harmful drug, a feeling of dependence related to the current practices of methadone treatment programmes and a poor therapeutic relationship. In the face of such challenges, the two major motivations that keep patients in care come from the desire to completely break up with heroin and the pursuit of family happiness. CONCLUSION: The current practices of methadone programmes pose challenges to patients' recovery efforts from addiction and threaten treatment retention. Prompt interventions are needed to help Vietnam attain its objective of providing better care for larger vulnerable populations.

Fatal dysfunction and disintegration of thrombin-stimulated platelets.

Platelets play a key role in the formation of hemostatic clots and obstructive thrombi as well as in other biological processes. In response to physiological stimulants, including thrombin, platelets change shape, express adhesive molecules, aggregate, and secrete bioactive substances, but their subsequent fate is largely unknown. Here we examined late-stage structural, metabolic, and functional consequences of thrombin-induced platelet activation. Using a combination of confocal microscopy, scanning and transmission electron microscopy, flow cytometry, biochemical and biomechanical measurements, we showed that thrombin-induced activation is followed by time-dependent platelet dysfunction and disintegration. After ~30 minutes of incubation with thrombin, unlike with collagen or ADP, human platelets disintegrated into cellular fragments containing organelles, such as mitochondria, glycogen granules, and vacuoles. This platelet fragmentation was preceded by Ca2+ influx, integrin αIIbß3 activation and phosphatidylserine exposure (activation phase), followed by mitochondrial depolarization, generation of reactive oxygen species, metabolic ATP depletion and impairment of platelet contractility along with dramatic cytoskeletal rearrangements, concomitant with platelet disintegration (death phase). Coincidentally with the platelet fragmentation, thrombin caused calpain activation but not activation of caspases 3 and 7. Our findings indicate that the late functional and structural damage of thrombin-activated platelets comprise a calpain-dependent platelet death pathway that shares some similarities with the programmed death of nucleated cells, but is unique to platelets, therefore representing a special form of cellular destruction. Fragmentation of activated platelets suggests that there is an underappreciated pathway of enhanced elimination of platelets from the circulation in (pro)thrombotic conditions once these cells have performed their functions.