Journal of Thrombosis and Haemostasis最新文献

Background: Fibrinogen is a critical coagulation factor that plays an essential role in thrombosis and is elevated in individuals with chronic inflammation. Here, we used fibrinogen as a representative quantitative measure of pro-coagulant risk and evaluated metabolites associated with fibrinogen levels through non-targeted plasma metabolomic profiling (Broad and Metabolon platforms).

Methods: Our analysis included 10,533 individuals across six U.S. based cohorts representing diverse population groups. The cross-sectional relationship between each of 789 tested metabolites and plasma fibrinogen concentration was assessed with adjustment for relevant covariates such as age, sex, body mass index, and circulating lipoprotein levels.

Results: Meta-analysis of per-cohort results revealed 270 metabolites significantly associated with fibrinogen level (FDR adjusted p-value < 0.05). Lipid species such as glycerophospholipids, sphingolipids, and fatty acyls were prevalent among significantly associated metabolites; some of these may capture effects of inflammation, as supported by sensitivity analyses adjusted for C-reactive protein. Significant associations between fibrinogen levels and serotonin, thyroxine, and sex-hormone derivatives may capture endogenous influences on fibrinogen levels. Exogenous compounds and microbial co-metabolites significantly associated with fibrinogen also implicate lifestyle and microbiome risk-factors. Only a portion of fibrinogen-associated metabolites (30%) have been associated with a cardiovascular disease outcome in a prior study, suggesting the associations discovered here may provide insights on vascular biology which case-control studies may not yet be powered to detect.

Conclusions: These findings contribute to a growing list of metabolite biomarkers that may influence coagulation and inflammation pathways and may thereby contribute to vascular risk.

Background: Thrombotic and bleeding complications in Myelodysplastic Syndromes (MDS) can affect therapy, but how they impact overall survival remains unclear.

Objective: Evaluating the impact of venous thromboembolism (VTE) and bleeding on survival in older MDS patients.

Methods: Incident MDS cases were collected from SEER-Medicare database (2007-2017) and followed from diagnosis until death. Baseline demographics, comorbidities, and SEER-Medicare MDS Risk Score (SMMRS) were assessed. VTE and bleeding events were identified using validated algorithms and modeled as time-varying covariates. Cox regression, adjusted for age, sex, race, NCI Comorbidity Index, and SMMRS, estimated the hazard ratio (HR) for death.

Results: Among 13,995 MDS patients (median age 82, 46% female), 1,114 VTE, and 1,905 bleeding events occurred. VTE was associated with increased mortality <3 months (HR 3.21; 95% CI 2.84-3.62) and 3-6 months (HR 1.51; 95% CI1.23-1.85), but not for 6-12 months (HR 1.18; 95% CI 0.98, 1.42) or >12 months (HR 1.00; 95% CI 1.91, 1.11). Bleeding was associated with mortality for all post-event time-periods assessed; <3 months (HR 4.25; 95% CI 3.89, 4.65), 3-6 months (2.11; 95% CI 1.82, 2.45), 6-12 months (1.60; 95% CI 1.39, 1.85) and 12+ months (1.43; 95% CI 1.31, 1.57). Case-crossover analysis confirmed these associations for bleeding but revealed potential unmeasured confounders for VTE.

Conclusion: Bleeding was associated with subsequent mortality in MDS. For VTE this risk seems to be affected by potential unmeasured confounders. These findings underscore the need for strategies to mitigate bleeding risk in this population.

Background: Pregnant women with von Willebrand Disease (VWD) receive prophylactic von Willebrand factor (VWF) concentrate based on third-trimester VWF/FVIII levels to reduce the risk of severe postpartum hemorrhage (PPH, ≥1000 mL). Due to high severe PPH rates, Dutch guidelines were revised in 2018. Consensus was reached to increase the third-trimester threshold for prophylaxis from <50 IU/dL to <80 IU/dL, and peak target levels during childbirth from ≥100 IU/dL to ≥150 IU/dL.

Methods: Pregnant Dutch women with VWD were prospectively enrolled (2018-2024) to assess the severe PPH incidence after guideline revision. VWF/FVIII activity levels, hematologic and obstetric outcomes were compared to a historical cohort (2012-2017). Statistics included descriptives and logistic regression to correct for confounders.

Results: Severe PPH occurred in 18.1% (n=29/160)without thrombosis or exsanguinations. Prophylaxis in those with third trimester levels <80 IU/dL led to PPH rates similar to those with spontaneous a rise >80 IU/dL. Compared to the historical cohort (prophylaxis cut-off <50 IU/dL), severe PPH incidence did not decrease (n=20/151 vs. n=29/160, OR 1.45 95%CI 0.78-2.69). Also in the third-trimester 50-80 IU/dL subgroup and third-trimester <50 IU/dL subgroup, the risk for severe PPH was similar (n=31/160 vs n=23/151, OR 0.86, 95%CI 0.23-3.28 and n=64/160 vs n=48/151, OR 2.59, 95%CI 0.78-8.60, respectively), despite increased peak target levels of 150 IU/dL.

Conclusion: Increasing the third-trimester VWF and FVIII cut-off to <80 IU/dL and aiming for ≥150 IU/dL at delivery did not decrease severe PPH. More research is needed on optimal peripartum hemostatic prophylaxis in VWD.

Background: Hemostasis is maintained through a delicate balance between procoagulant and anticoagulant mechanisms. Protein S (PS), a multidomain, vitamin K-dependent glycoprotein, contributes to this balance not only as a cofactor for activated protein C and tissue factor pathway inhibitor but also by directly inhibiting activated factor IX (FIXa). However, the structural determinants underlying FIXa inhibition remain unclear.

Objectives: This study aimed to (1) identify the FIXa binding interface on PS and (2) investigate the role of its laminin G (LG) domains in mediating FIXa binding and inhibition.

Methods: Molecular docking was used to predict the FIXa binding interface on PS. Fluorescence-based binding assays were performed to determine binding affinities, and functional coagulation assays were conducted to measure inhibitory constants. Finally, site-directed mutagenesis was carried out to generate specific PS mutants.

Results: Molecular docking and in vitro binding assays demonstrated that both the LG1 and LG2 domains interact with FIXa. Quantitative fluorescence-based binding analyses revealed that the LG1+2 tandem domains exhibited the highest affinity for FIXa (K_d ≈ 52.15 nM). Functional coagulation assays showed that LG1+2 effectively blocked FIXa-mediated factor X activation and suppressed thrombin generation. Furthermore, site-directed mutagenesis confirmed that residues E435 and E437 within the LG domains are critical for FIXa binding and inhibition.

Conclusion: These findings identify the LG domains of PS as essential structural elements for direct FIXa inhibition, independent of its cofactor function. By elucidating this domain-specific mechanism, our work provides a structural framework for the rational design of selective FIXa inhibitors and FIXa-binding peptides as novel antithrombotic strategies.

Background: Inversions involving intron 22 (Inv22) of F8 are detected in approximately 45% of all severe hemophilia A (HA) patients. Digital droplet PCR using mile-post assays and MLPA allows robust diagnosis of inversions and copy number variations as well as more complex rearrangements in F8.

Objectives: 350 HA patients and 280 of their mothers from the HIGS cohort were analyzed to identify additional F8 mutations.

Methods: Digital droplet PCR and MLPA were used to investigate archival samples.

Results: Out of 350 patients analyzed, 13 were found to harbor previously unidentified mutations, three with inversions (Inv22 type 1) and 10 with large deletions. In addition, 7 patients had complex rearrangements with duplications in addition to their Inv22 type 1 inversions. Out of 138 mothers to patients with inversions, 7 were non-carriers. Five of these were found to have the same duplications that were detected in their 7 sons, indicating that most, if not all, non-carrier mothers have other rearrangements in the F8 region. MLPA showed that these duplications had breakpoints in intron 22 with either a duplication of the first part of the gene (exons 1-22) or the last part of the gene (exons 23-26).

Conclusions: Mutation detection in F8 can be improved by dedicated analysis for inversions and duplications/deletions using mile-post analysis. Non-carrier mothers of hemophilia A patients with Inv22 inversions often have other rearrangements.

Background: Several illicit drugs have been reported to be associated with ischemic stroke and myocardial infarction. However, the underlying mechanisms remain poorly investigated. In this study, we focused on N-Ethylpentylone (NEP), a synthetic cathinone with potent hallucinogenic property and explored its role in platelet activation.

Methods: Platelet function assays and flow cytometry were used to evaluate the effects of NEP on platelet aggregation, spreading, clot retraction, and integrin αIIbβ3 activation. The role of the 5-Hydroxytryptamine2A receptor (5-HT2AR) in NEP-mediated platelet responses was investigated using the selective 5-HT2AR antagonist M100907 in both ex vivo and in vivo. Phosphoproteomics identified NEP-regulated phosphorylation sites, and Western blotting validated mitogen-activated protein kinase (MAPK) pathway activation by targeting key phosphorylation nodes.

Results: NEP significantly potentiated platelet aggregation, spreading, clot retraction, and integrin αIIbβ3 activation in a concentration-dependent manner. The NEP potentiated platelet responses were suppressed by M100907 in both ex vivo and in vivo models, confirming the critical regulatory role of 5-HT2AR. Phosphoproteomics revealed NEP-triggered phosphorylation upregulation, enriched in MAPK pathways. Western blotting confirmed selective ERK and p38 phosphorylation, with no effect on JNK.

Conclusion: NEP directly enhances agonist-induced platelet activation by targeting 5-HT2AR on platelet membranes. The findings of this study provide valuable insights for elucidating the hematotoxic mechanisms of NEP and other illicit drugs.

Thrombosis, when considering all its manifestations including myocardial infarction, stroke, and venous thromboembolism, constitutes the leading cause of death world-wide. Treatment of thrombotic diseases has been revolutionized by direct oral anticoagulants (DOACs) targeting thrombin and factor (F)Xa. However, the therapeutic or prophylactic use of DOACs is not without limitations, including persistent and significant bleeding risks. In this review, we summarize and discuss current state-of-the-art of novel and experimental anticoagulation and fibrinolytic/thrombolytic therapies beyond DOACs. In particular, we review studies investigating contact pathway inhibition, including in vitro and in vivo studies of FXIIa and FXIa inhibition, and clinical trials of contact pathway inhibition. We review in vitro and in vivo studies investigating inhibition of common pathway coagulation targets, including FV, FVIII, and FIX. This is followed by analysis of options for the therapeutic targeting of fibrin or fibrinogen and FXIII. Next, we explore opportunities for the therapeutic harnessing of naturally occurring anticoagulant pathways as well as fibrinolytic mechanisms. The current state-of-the-art research for each of these mechanisms is summarized, including whether studies have progressed from in vitro to in vivo experimentation and whether clinical trials have been performed. We highlight particularly novel areas of interest and include evaluation of the relative preclinical and clinical trial progress for the selected targets. The increased understanding of mechanisms driving thrombosis holds promise for future developments in novel anticoagulants that may contribute to reducing the impact and burden of thrombotic diseases while improving safety of current therapeutic options.