Journal of Thrombosis and Haemostasis最新文献

Background: Thrombotic thrombocytopenic purpura (TTP) is a rare but life-threatening thrombotic microangiopathy, with a substantial risk of relapse despite advances in therapy. Robust biomarkers to predict relapse are urgently needed to inform risk-adapted management. This study represents the first application of bone marrow proteomic profiling in TTP to explore relapse-associated biomarkers.

Objectives: To identify clinical risk factors and novel protein biomarkers for relapse in TTP through integrated clinical and bone marrow proteomic analyses.

Methods: We conducted a retrospective cohort study involving 123 patients diagnosed with TTP at a single center. Clinical and laboratory variables at initial presentation were analyzed to identify potential predictors of relapse using Cox proportional hazards models. To explore molecular correlates of relapse, bone marrow paraffin-embedded samples from 18 patients were subjected to quantitative proteomic profiling. Machine learning algorithms were applied to identify candidate relapse-associated proteins. Selected proteins were subsequently validated by immunohistochemistry of corresponding tissue sections.

Results: Age ≤30 years and severe neuropsychiatric symptoms were independently associated with increased relapse risk, but demonstrated only moderate predictive performance (area under the curve = 0.706). Proteomic profiling revealed TMEM109 as a relapse-associated protein with superior discriminative capacity (area under the curve = 0.929). Immunohistochemistry analysis confirmed reduced TMEM109 expression in relapsed patients (P < .0001).

Conclusion: TMEM109 is a promising biomarker for predicting TTP relapse. While clinical factors such as age and neuropsychiatric symptoms provide valuable prognostic information, TMEM109 may enhance relapse-risk stratification, offering potential for individualized monitoring and therapeutic decision-making. These findings underscore the value of proteomic profiling in complementing clinical evaluation in TTP.

Background: Apolipoprotein C3 (apoC3) circulates primarily on triglyceride-rich lipoproteins and promotes atherosclerosis by fostering lipidemia and inflammation. Thus, apoC3 represents an important cardiovascular risk factor and is associated with cardiovascular events and mortality. Due to their dual nature as hemostatic and immunomodulatory effector cells, platelets play an important role in the development and progression of atherosclerosis and are responsible for thrombotic/thromboembolic events upon plaque rupture.

Objectives: We aimed to elucidate the impact of apoC3 on prothrombotic platelet functions.

Methods: Platelets were isolated from healthy volunteers and the effect of apoC3 on their activation and aggregation was assessed by flow cytometry, ELISA, and light transmission and multiple electrode aggregometry. Platelet spreading and cytoskeletal remodeling were examined by immunofluorescence microscopy. In vivo relevance was confirmed in a murine model of FeCl₃-induced thrombosis.

Results: ApoC3 strongly reduced platelet aggregation independently of the presence of plasma or other blood cells and impaired both α_IIbβ₃ activation and degranulation. While agonist-induced calcium mobilization, hyperpolarization, and membrane fluidity were not affected, apoC3 slightly reduced AKT phosphorylation and increased vasodilator-stimulated phosphoprotein (VASP) phosphorylation. Furthermore, apoC3-treated platelets displayed impaired lamellipodia formation, which was accompanied by aberrant actin cytoskeleton remodeling. Finally, transfusion of apoC3-treated platelets into mice delayed thrombus formation in vivo.

Conclusion: We identified apoC3 as lipoprotein-derived inhibitor of prothrombotic platelet functions, mediating antiaggregatory effects, likely via modulating AKT and VASP signaling and interfering with actin cytoskeleton remodeling to impair lamellipodia formation. Thus, apoC3 may counterbalance its proatherogenic properties on lipid metabolism and inflammation by dampening thrombotic risk in hyperlipidemia.

Background: Venous thromboembolism (VTE) prevention in obstetrics remains a major public health concern. Precise data on timing of occurrence and on women at risk are necessary to improve thromboprophylaxis strategies.

Methods: From June 2015 to January 2019, the prospective cohort "HEMOTHEPP" study included all pregnant women aged ≥16 admitted for delivery after 15 weeks of gestation, in the Finistère region, France (6 maternity units), and followed during three-month postpartum. Ethics committee approved the study. The primary outcome was symptomatic, documented VTE - deep vein thrombosis (DVT), pulmonary embolism (PE), unusual site VTE - adjudicated by an independent committee.

Findings: Among the 20,238 included women, VTE incidence was 4·9‰ person-years (95% confidence interval 3·3-7·2) during postpartum and 1·3‰ (0·8-2·0) during pregnancy. Postpartum VTE included 40·0% isolated PE, 12·0% PE with DVT, 12·0% isolated DVT, and 36·0% unusual site VTE. 32·0% occurred during days 0 to 7, 60·0% during days 8 to 42, and 8·0% during days 43 to 84. According to the Royal College of Obstetricians and Gynaecologists risk score, 7,532 women (37·3%) were classified at intermediate or high risk of postpartum VTE and eligible for thromboprophylaxis: among them, only 3,179 (42·2%) received it. Among intermediate-risk women, VTE incidence was similar with or without thromboprophylaxis: 12·2‰ and 13·1‰ person-years, respectively.

Interpretation: Postpartum VTE incidence was twice higher than expected. Thromboprophylaxis during postpartum was under-prescribed. The high VTE incidence rate after the first week postpartum and in intermediate-risk women, regardless of thromboprophylaxis, highlights the need for randomised trials to evaluate thromboprophylaxis benefit.

Background: Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant disorder with an overwhelming hemorrhagic phenotype. It is mainly caused by variants in the ENG and ACVRL1 genes. HHT prevalence is currently estimated to be 1 in 5000 individuals, but the disease is likely underdiagnosed due to variable clinical presentation, misdiagnosis, and delayed recognition.

Objectives: To estimate the global genetic prevalence of HHT-associated variants in ENG and ACVRL1.

Methods: We analyzed 3 large population-scale genomic databases: gnomAD, All of Us, and Regeneron Genetics Center-Million Exome. We considered known pathogenic and likely pathogenic variants of ENG and ACVRL1 and extended the analysis to potentially pathogenic variants passing the pathogenic criteria established by the guidelines for HHT of the American College of Medical Genetics and Genomics/Association for Molecular Pathology.

Results: The genetic prevalence of HHT ranged from 1.753 to 2.555 in 5000 individuals, when considering only pathogenic and likely pathogenic variants, and from 2.874 to 4.327 in 5000 individuals, when also potentially pathogenic variants were considered.

Conclusion: This study assesses the prevalence of HHT-associated variants in the general population. Our unbiased approach demonstrates that the genetic prevalence of the disease is substantially higher than currently estimated.

Background: Acute myocardial infarction (AMI) remains a major cause of morbidity and mortality. Besides traditional cardiovascular risk factors, autoimmune components such as antiphospholipid antibodies (aPL) have been implicated, but their prognostic role in AMI is unclear.

Objectives: This study investigated the prognostic significance of aPL in patients with AMI and explored potential mechanisms.

Methods: This study consecutively enrolled 777 patients with AMI and detected aPL. Cox proportional hazard models assessed cardiovascular event risk. For mechanistic insights, an antiphospholipid syndrome (APS) myocardial infarction mouse model was developed, and bulk RNA sequencing was performed on human coronary artery endothelial cells (HCAECs) stimulated with APS immunoglobulin G.

Results: Compared with the aPL-negative group, patients in the aPL-positive group were significantly younger (62.0 [54.0-69.0] vs 52.0 [44.0-60.0] years; P < .001). aPL-positive AMI group had higher incidence of thrombosis (199 [27.3%] vs 22 [46.8%]; P = .007) and fewer traditional risk factors such as hypertension and hyperlipidemia. During a mean follow-up of 18.8 months, aPL positivity was significantly associated with future cardiovascular events (adjusted hazards ratio, 2.28; 95% CI, 1.26-4.12; P = .006). In the mouse model, the APS group exhibited accelerated disease progression and higher thrombosis rates. Neutrophil-related chemokines were upregulated in APS immunoglobulin G-stimulated HCAECs, consistent with increased neutrophil infiltration and neutrophil extracellular traps formation in vivo.

Conclusion: Our findings suggest that patients with aPL-positive AMI represent a distinct AMI subset with worse outcomes. They may benefit from additional therapies beyond standard treatment. Neutrophil-driven inflammatory pathways could be a potential underlying mechanism.

Background: Neutrophil extracellular traps (NETs) are important in host defense but implicated in pathological thrombosis in acute ischemic stroke (AIS). NET quantification is typically done with immunostaining for NET-specific markers, but molecular techniques like mass spectrometry can provide another angle into thrombo-physiology.

Objectives: This study leverages both histological analysis of NETs, and molecular insights from proteomics, to validate using proteomics for evaluating stroke thrombus NET burden.

Patients/methods: Sections of 30 AIS thrombi were stained for myeloperoxidase (MPO), citrullinated histone H3 (H3Cit), and DNA. ImageJ measured average staining area. Mass spectrometry of sample thrombi fragments detected 5,784 proteins. Spearman correlations and Mann Whitney U-tests compared NET burden from staining to proteomic composition. Dimensional reduction of H3Cit and MPO staining created a single NET marker variable (PC1), used to compare thrombus composition between thrombi with NET marker greater (NET-rich) or lesser (NET-poor) than median.

Results: Percent coverage for the three stains was interrelated (Spearman's ρ > 0.6, p < 5E-4 for all). 531 neutrophil degranulation proteins were positively associated with all stains. PC1 explained ∼92% of the variance. Ubiquitin-proteasome degradation pathways were negatively correlated with the NET markers (q-val < 0.05 for all relevant pathways). Older patient age (median, IQR: 79, 73-85 vs 61, 57-73; Mann Whitney p = 0.03) and atrial fibrillation-associated stroke (Fisher's Exact p = 0.03) thrombi were NET-rich.

Conclusions: This study marries immunostaining with proteomics for quantifying NET burden, pointing to molecular phenotypes within NET-rich and NET-poor thrombi that may underpin NET formation and maintenance in stroke thrombi.

Background: Renal ischemia-reperfusion injury (IRI) contributes to acute kidney injury (AKI) by inducing oxidative stress, inflammation, and thromboinflammatory responses. Conventional heparins mitigate IRI, but their use is limited by bleeding and thrombocytopenia.

Objectives: To evaluate octaparin, a structurally defined synthetic sulfated octasaccharide with anticoagulant activity and minimal bleeding risk, as a novel therapeutic for IRI.

Methods: Murine renal IRI models and hypoxia/reoxygenation (H/R)-injured renal tubular epithelial cells (TCMK-1) were treated with octaparin. Renal function, histopathological changes, fibrosis, apoptosis, oxidative stress, inflammation, and coagulation parameters were systematically evaluated. DC-SIGN engagement and NF-κB signaling were analyzed through molecular docking, receptor blockade experiments, and immunoblotting.

Results: Octaparin significantly improved renal function in vivo, attenuated tubular necrosis and fibrosis, and suppressed apoptosis. It also markedly reduced oxidative stress, restored antioxidant enzyme activity, and downregulated pro-inflammatory cytokines, with reduced neutrophil infiltration. In terms of coagulation, octaparin inhibited tissue factor expression and fibrin deposition, alleviating thromboinflammatory responses. In vitro, octaparin similarly suppressed excessive ROS bursts, restored antioxidant defenses, reduced inflammatory cytokine release, and inhibited apoptosis in TCMK-1 cells. Mechanistically, octaparin bound to the DC-SIGN receptor on renal epithelial cells, inhibiting NF-κB phosphorylation and nuclear translocation, thereby disrupting ROS-driven inflammatory cascades-via a pathway distinct from classical heparin anticoagulation.

Conclusion: Octaparin exerts multifaceted renoprotective effects in IRI by modulating DC-SIGN/NF-κB signaling. Its ability to simultaneously suppress thromboinflammation, oxidative stress, and apoptosis-without the risks associated with conventional heparins-represents a transformative therapeutic strategy for AKI.

Background: The incidence of cancer-associated venous thromboembolism (CAT) has risen in recent decades, underscoring the need for novel risk biomarkers and mechanistic insights. MicroRNAs (miRNAs) have emerged as promising biomarkers for various diseases, with elevated miRNA-145-5p (miR-145-5p) levels linked to reduced venous thromboembolism (VTE) risk. We aimed to investigate the association between plasma miR-145-5p levels and risk of future VTE related to (CAT) and unrelated to cancer (non-CAT).

Methods: Plasma miR-145-5p were measured in a case-cohort derived from the Trøndelag Health Study (HUNT3; n=50,807). The study included 455 VTEs (95 CATs), occurring during 9 years of follow-up, and 1,740 randomly sampled age-weighted subcohort participants. VTEs were classified as CAT if they occurred within 1 year before or 2 years after a cancer diagnosis. Multivariable-adjusted hazard ratios (HRs) for CAT and non-CAT were estimated using weighted Cox regression, with cancer modeled as a time-varying covariate.

Results: Increasing miR-145-5p levels were associated with lower risk of CAT (HR per 1 standard deviation: 0.70, 95% CI: 0.52-0.96) and non-CAT (HR: 0.81, 95% CI: 0.72-0.91). Individuals in the highest miR-145-5p quartile had lower risk of both CAT (HR: 0.55, 95% CI: 0.25-1.20) and non-CAT (HR: 0.48, 95% CI: 0.33-0.69) compared to those in the lowest quartile.

Conclusions: High plasma miR-145-5p levels were associated with lower risk of CAT and non-CAT, suggesting that miR-145-5p has the potential to serve as a risk biomarker for CAT.

Background: Human platelets express 2 thrombin-activated G protein-coupled receptors (GPCRs), protease activated receptor (PAR)1 and PAR4. Previous studies have demonstrated PAR1 activation to be fast on-fast off while PAR4 activation was slower but more sustained. However, how PARs regulate second messengers such as cyclic adenosine monophosphate (cAMP), a negative regulator of platelet activation, remain unknown.

Objectives: This study investigated how PAR1 and PAR4 activation by thrombin regulates cAMP levels and platelet activation.

Methods: PAR1 and PAR4 on washed human platelets were inhibited with vorapaxar or BMS-986120, respectively, in the presence or absence of prostaglandin (PG)I₂. Platelets were stimulated with various thrombin concentrations, and calcium flux and platelet aggregation were measured. PAR regulation of cAMP in megakaryocytes (MKs) was also studied by genetic disruption of PAR1 or PAR4 using CRISPR/Cas9 editing.

Results: In the presence of PGI₂, cAMP was significantly reduced after thrombin stimulation when platelets or MKs were activated through PAR1, but not PAR4. Calcium flux was highly associated with cAMP level changes. High concentrations of cAMP were associated with low calcium flux with PAR4 stimulation. In contrast, stimulating PAR1 decreased cAMP levels, and this was associated with a more pronounced calcium flux. Phosphodiesterase (PDE)3A is involved in regulating cAMP levels. Blocking PDE3A with a pharmacological inhibitor in platelets or genetic knockout in MKs reversed PAR1-induced decrease of cAMP level, while PAR4 was not affected by PDE3A inhibition.

Conclusion: The data suggest differences between PAR1 and PAR4 induced thrombin activation are due, in part, to cAMP regulation by PDE3A.