Journal of Thrombosis and Haemostasis最新文献

Background: Chronic thromboembolic pulmonary hypertension (CTEPH) presents challenges due to its complex pathobiology. Although numerous studies have reported heterogeneous cell types by single cell RNA sequencing (sc-RNA seq), the atlas and characteristics of plasma cells remain poorly understood.

Methods: We performed scRNA seq on pulmonary endarterectomy tissue from five patients and six normal pulmonary arteries. Serum immunoglobulins (Igs) were measured using protein electrophoresis among 273 CTEPH patients, 259 idiopathic pulmonary arterial hypertension (IPAH) patients and 251 healthy controls.

Results: The percentage of plasma cells was significantly increased from less than 1% in healthy controls to 15% in CTEPH patients. We identified one B cell cluster and five distinct mature plasma cell clusters, including IGHG1, HSPA1A, AHNAK, IGLC3 and IGKV4. Notably, the AHNAK and IGLC3 subclusters are newly identified. GeneSwitches analysis indicated early activation of IGHG1 and early deactivation of HLA-DPA1. The trajectory of AHNAK cluster was earlier than that of IGLC3 cluster, with an enrichment for pathways responsive to lipopolysaccharide. The IGLC3 cluster revealed lower differentiation potential and was predominantly associated with Igs production. Furthermore, Igα2 levels in CTEPH patients were lower than in controls but higher than in IPAH patients. Significantly, Igγ levels were markedly elevated in CTEPH patients compared to IPAH patients and controls, better distinguishing CTEPH patients from controls and IPAH patients.

Conclusions: Plasma cells of CTEPH had a distinctive landscape and heterogeneity. The newly identified clusters represented excessive Igs production, but lacking immune response function. These findings highlight targeted plasma cells to develop novel CTEPH treatments.

Background: The impact of central localization of pulmonary embolism (PE) on clinical outcomes is uncertain.

Objectives: To compare clinical presentation, risk factors, and outcomes between patients with central pulmonary embolism (cPE) and non-cPE.

Methods: We retrospectively analyzed 597 patients with acute PE from the prospective SWITCO65+ cohort between 09/2009-12/2013. cPE was defined as an embolus in the pulmonary trunk or the left or right pulmonary artery. We compared baseline clinical characteristics and outcomes at 3 months (recurrent venous thromboembolism [VTE], overall/PE-related mortality, PE-related quality of life) and over the entire follow-up (recurrent VTE, overall/PE-related mortality) between patients with cPE vs. non-cPE. We examined the association between PE localization and recurrent VTE and overall mortality, adjusting for multiple confounders including thrombolysis and periods of anticoagulation, and competing risk of non-VTE-related death if appropriate.

Results: Overall, 217 (36.3%) patients had cPE. Symptoms/signs of respiratory distress, right-ventricular dysfunction, and myocardial injury were more prevalent in those with cPE. VTE recurrence, overall/PE-related mortality, and PE-related quality of life at 3 months did not vary by PE localization. After a median follow-up of 29.6 months, patients with cPE had a higher risk of fatal PE (5.5% vs. 2.1%; P=0.033). After adjustment, cPE was associated with recurrent VTE (SHR 2.22, 95%CI 1.25-3.91) but not with overall mortality (HR 0.74, 95%CI 0.45-1.21) during follow-up.

Conclusion: cPE was associated with a 2.2-fold increased risk of recurrent VTE compared to non-cPE. Whether an extended anticoagulation duration could reduce the recurrence risk following cPE should be further examined.

Background: Patients with testicular germ cell tumors (TGCT) have a high cancer-specific survival rate. We aimed to determine the short- and long-term risk of arterial thromboembolic events (ATE), their impact on mortality, and risk factors for ATE in TGCT patients.

Methods: Patients with TGCT treated between 1994-2020 were included in a single-center retrospective cohort study. The primary outcome was ATE (i.e., acute coronary syndrome, ischemic stroke, acute peripheral arterial occlusion). Cumulative incidences were obtained in competing risk analysis. The impact of ATE on mortality was analyzed in a multi-state model. Cox-regression was used to explore short-and long term ATE-risk factors.

Results: Overall, 1,277 patients were included (median age: 35 years; seminoma: 56%, 44% cisplatin-based chemotherapy). Cumulative ATE-incidences at 1-, 10-, and 25-years were 0.6% (95% confidence interval [CI]: 0.3-1.1), 2.6% (1.8-3.7), and 12.0% (8.7-15.9). ATE diagnosis was independently associated with increased all-cause mortality (age-adjusted transition hazard ratio: 4.61 [95%CI: 2.40-8.85], p<0.001). Cisplatin-based chemotherapy was associated with ATE-risk within 1 year after TGCT diagnosis (1.4% vs 0%, p<0.001), whereas no differences were observed thereafter. Regarding long-term ATE-risk, a point-based risk score was derived (age ≥35, smoking, LDH ≥250IU/L), which efficiently stratified ATE risk (Harrel´s C: 0.71 [95% CI: 0.63-0.78]), with cumulative ATE-incidences in low-, intermediate- and high-risk patients of 3.9%, 11.4%, and 22.7%, respectively.

Conclusions: ATE represent a common complication in TGCT survivors and are associated with increased mortality. A simple point-based score efficiently stratifies long-term ATE-risk, whereas cisplatin-based chemotherapy increased short-term ATE risk.

Background: Body height is associated with venous thromboembolism (VTE) and may contribute to differences in VTE risk in men versus women.

Aims: To investigate the risk of VTE according to body height in men and women, and assess VTE risk in men versus women after adjustment for height in young, middle-aged, and elderly individuals.

Methods: Participants of the Tromsø Study (1994-2020) and the Nord-Trøndelag Health Study (1995-2019) formed the study cohort (n=114,567). Cox regression was used to estimate hazard ratios (HR) with 95% confidence intervals (CI) of VTE per 10 cm increase in body height in men and women. VTE risk in men versus women was estimated in the age groups 19-49, 50-74 and ≥75 years before and after adjustment for height.

Results: Taller stature was associated with increased VTE risk in both men (HR: 1.34, CI: 1.25-1.44) and women (HR: 1.23, CI: 1.13-1.33). In the middle-aged, the risk was higher in men than in women (HR: 1.45, CI: 1.32-1.60), but diminished after adjustment for height (HR 0.98, CI 0.85-1.13). In the young (HR: 0.87, CI 0.70-1.08) and elderly (HR: 1.08, CI: 0.97-1.20) there was no difference in VTE risk in men versus women before adjustment, while the risk was higher in women after height adjustment (HR: 0.60, CI: 0.44-0.83 and HR: 0.83, CI: 0.71-0.97, respectively).

Conclusions: Taller stature was a risk factor for VTE in men and women. The risk of VTE in men versus women was substantially affected by adjustment for body height in all age groups.

Background: Thrombotic thrombocytopenic purpura (TTP) is a potentially fatal blood disorder, resulting from severe deficiency of plasma ADAMTS13 activity. Current treatment for immune-mediated TTP includes daily therapeutic plasma exchange (TPE), plus caplacizumab and immunosuppressives. For hereditary TTP, resulting from mutations of ADAMTS13, plasma infusion or recombinant ADAMTS13 is the treatment of choice. However, there are still unmet needs for an effective alternative therapy for TTP.

Objective: The present study aims to evaluate therapeutic efficacy of a novel humanized antibody Fab fragment against platelet glycoprotein Ibα (CA1001) in a murine model of TTP.

Methods: Platelet agglutination profiles, microfluidic shear-based assay, and intravital microscopy thrombosis model, as well as lysine histone-induced murine "TTP-like" model were employed.

Results: CA1001 exhibits potent inhibition of botrocetin-induced murine platelet agglutination in a dose and time-dependent manner. CA1001 also significantly inhibits shear-dependent adhesion and aggregation of murine platelets to endothelial von Willebrand factor (VWF) released from calcium ionophore activated cremaster venules in Adamts13^-/- mice and blocks the formation of platelet-VWF rich thrombosis. More importantly, CA1001 appears to be efficacious in preventing and treating a histone-induced "TTP-like" syndrome in Adamts13^-/- mice, demonstrated by the allievation of thrombocytopenia, prerenal injury, and the formation of microvascular thrombosis in major organ tissues.

Conclusions: CA1001 can effectively inhibit VWF-platelet interaction and thrombus formation under various (patho)physiological conditions. Thus, CA1001 may be a potential candidate for further development as a novel therapeutic for immune-mediated and hereditary TTP and perhaps for other inflammatory thrombotic disorders such as ischemic stroke.

Background: Both protein disulfide isomerase (PDI) and SARS-CoV-2 main protease (M^pro) are reliant on active site cysteines stabilized by adjacent amino acids. We reasoned that redox active compounds might interfere with both enzymes by acting in the vicinity of these reactive sites thus interfering with viral replication and thrombus formation. Our previous screen of 1019 flavonoids identified several compounds that inhibit SARS-CoV-2 M^pro.

Aims: Our goal was to identify phytochemical inhibitors of SARS-CoV-2 M^pro that block thiol isomerases and are antithrombotic.

Methods: PDI, ERp57, ERp5, ERp46, isolated domains of PDI, and PDI mutants were used to evaluate effects of galloylated polyphenols and their analogs on thiol isomerase reductase activity. Laser-injury and FeCl₃ models of thrombus formation and a tail snip assay were used to assess effects on thrombosis and hemostasis.

Results: Pinocembrin 7-O-(3''-galloyl-4'',6''-(S)-hexahydroxydiphenoyl)-beta-D-glucose (PGHG) inhibited both PDI and SARS-CoV-2 M^pro. Evaluation of isolated PDI fragments and active site cysteine mutants showed that PGHG acts at the catalytic domains. Structure-function studies showed that PGHG interacts with histidines within the CGHC motifs of PDI. PGHG was equally active against other thiol isomerases, including ERp57, ERp5, ERp72, and ERp46. Screening numerous galloylated polyphenols demonstrated a class effect on thiol isomerase inhibition. Structure-activity relationships indicated that the galloyl moieties within large galloylated polyphenols were important for their inhibitory activity. PGHG and punicalagin were antithrombotic in murine models of thrombus formation.

Conclusions: Galloylated polyphenols represent a large class of antithrombotic compounds with broad activity against thiol isomerases. Many of these compounds also inhibit SARS-CoV-2 M^pro and viral replication.

Over the past three decades, omics technologies have revolutionized our understanding of platelet molecular content and organization, enabling the systematic analyses of platelet physiology. Among these approaches, proteomics has been especially significant in discovering as well as validating molecular mechanisms of platelet function in health and disease. However, several conceptual and practical challenges continue to limit the full utility of platelet proteomics tools and data. Methodological and analytical inconsistencies remain a key concern, with biological and technical variables exerting substantial influence on study outcomes and interpretation. These issues are compounded by the rapid pace of proteomics tool development and dataset collection, outstripping efforts to standardize best practices and ensure consensus as platelet proteomics consolidates itself as a tool for research even outside the Thrombosis and Hemostasis field. In this communication from the International Society on Thrombosis and Haemostasis (ISTH) Scientific and Standardization Committee (SSC), we highlight recent advances in platelet proteomics studies, and we identify where collective efforts can strengthen experimental design, execution, and analysis. As a practical recommendation, we encourage platelet biologists to recognize current discrepancies and advance efforts to standardize and customize methods and reporting practices, including blood collection, platelet isolation, data acquisition, and data interpretation. By aligning protocols and ensuring detailed reporting, the field can more effectively integrate proteomics findings and accelerate our understanding of platelet biology.

Background: Neuraxial anaesthesia is used for pain management in surgical and non-surgical settings. Spinal/epidural haematomas likely occur in between 1:10,000 and 1:200,000 procedures. Risk is thought to be greater in patients with bleeding disorders/thrombocytopenia and there are no existing comprehensive recommendations to guide neuraxial anaesthesia in these patients.

Objectives: The study's objective was to develop recommendations to advise clinicians on treatment thresholds for neuraxial anaesthesia in patients with platelet disorders/coagulation defects.

Methods: A four-round electronic modified Delphi consensus study was conducted. A steering committee generated the original Delphi statements and refined them based on panelist feedback. Consensus was achieved if ≥70% of participants agreed/strongly agreed or disagreed/strongly disagreed with a statement. This project was endorsed by the International Society on Thrombosis and Hemostasis Scientific and Standardization Committee Subcommittee on von Willebrand factor.

Results: Forty-five experts participated (42% response rate) with an essentially equal number of haematologists and anaesthesiologists. Thirty consensus statements were developed for 11 disorders ranging from various causes of thrombocytopenia, inherited platelet function disorders (IPFD), and single or multiple coagulation defects in obstetrical and non-obstetrical patients. Risk of sampling bias is present due to a predominantly North American sample, attrition (common in Delphi studies), and steering committee participation in the Delphi rounds.

Conclusions: This is the first set of consensus recommendations for neuraxial anaesthesia in adult patients with an array of platelet disorders/coagulation defects. These recommendations, based on the best available evidence and expert opinion, provide a decision framework for clinicians when faced with this challenging scenario.

Background: Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by the presence of antiphospholipid antibodies (aPL) in patients with thromboembolic/-inflammatory events and/or obstetric complications.

Objectives: The aim of this study was to examine whether there are alterations in the platelet lipidome of APS patients in comparison to patients affected by thromboembolism without APS (control) and healthy volunteers.

Methods: We applied quantitative mass spectrometry-based lipidomics to investigate the platelet lipidome of isolated resting and thrombin-stimulated platelets as well as platelet release in patients with APS, controls and healthy volunteers.

Results: Lipidomic data revealed an increase in lysophospholipids (LPL) in platelets from APS patients, specifically in lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE) species. As LPL are cleavage products generated by phospholipase A (PLA) from the corresponding phospholipid (PL) precursor, LPL/PL ratios may be employed as surrogates for PLA1 and PLA2 activities. The surrogate ratios for PLA2, which participates in the release of arachidonic acid (AA) during platelet activation, were significantly increased in APS in both resting platelets and upon thrombin-induced activation for phosphatidylcholine (PC) and phosphatidylethanolamine (PE). The PC-PLA2 surrogate ratio was found to correlate with serum levels of anti-β2-glycoprotein I and anti-cardiolipin IgG. Finally, receiver operator characteristic (ROC) analysis demonstrated excellent discrimination of patients with APS from controls and healthy volunteers.

Conclusion: These findings provide substantial evidence that platelet activation is enhanced in APS in vivo, involving the activation of PLA2.

Transparent reporting is key to improve reproducibility of scientific research. In 2023, the International Society for Extracellular Vesicles (ISEV) updated the "Minimal information for studies of extracellular vesicles" (MISEV) reporting guidelines, and published new recommendations for blood EV research entitled "MIBlood-EV: Minimal information to enhance the quality and reproducibility of blood extracellular vesicle research". The MIBlood-EV recommendations are part of MISEV2023, and promote reporting not only the protocols used for blood collection and handling, but also the composition of the prepared samples that are used to measure extracellular vesicles (EVs). Plasma and serum are commonly used starting materials for EV research; reporting their composition can help to improve reproducibility, comparison of measurement results, and support evidence-based guideline development. We conducted an online survey amongst ISTH EV researchers. Of the twenty respondents, 95% are familiar with MISEV, but 35% are unaware of the 2023 update, and only 65% apply these guidelines to their reports. With regard to MIBlood-EV, 40% are unaware of this reporting tool, and 20% do not follow its recommendations. This is surprising, because most respondents agree that preanalytical variables of blood EV research are not satisfactorily described (75%), confirm that having a standardized reporting tool is beneficial for blood EV research (90%), and consider MIBlood-EV applicable to other fields of ISTH research (80%). In this SSC communication, we summarize the survey results, as well as the background and goals of MISEV, and how MIBlood-EV can be useful to improve the reproducibility of blood research within the ISTH community.