Journal of Thrombosis and Haemostasis最新文献

Background: Polyphosphate (polyP) released from activated platelets acts as a procoagulant, promoting thrombin generation and fibrin formation. Like heparin, polyP binds to thrombin and fibrin(ogen).

Objectives: To determine whether, like heparin, polyP promotes thrombin binding to fibrin, thereby forming a ternary polyP-thrombin-fibrin complex that modulates thrombin activity.

Methods: The affinity of fluorescein isothiocyanate-labeled polyP or heparin for fibrin was determined by measuring unbound polyanion in the supernatants of fibrin clots. Likewise, the affinity of thrombin for fibrin in the absence or presence of polyanion was determined by quantifying unbound thrombin by chromogenic assay. Rate constants of thrombin inhibition by antithrombin-heparin were determined in the absence or presence of polyanion and fibrin. Clots formed with polyanions were incubated with labeled fibrinogen to monitor clot accretion.

Results: PolyP and heparin bind fibrin with Kd values of 1.7 ± 0.1 μM and 1.1 ± 0.1 μM, respectively. In the presence of polyP or heparin, thrombin binds to fibrin with Kd values of 1.2 ± 0.2 μM or 0.9 ± 0.1 μM, respectively, whereas the Kd is 5.6 ± 0.4 μM in the absence of a polyanion. Thrombin within the ternary polyP-thrombin-fibrin complex is protected from inhibition by the antithrombin-heparin complex but has reduced capacity to induce fibrin accretion.

Conclusions: Like heparin, polyP promotes the formation of a ternary polyP-thrombin-fibrin complex, which protects thrombin from inhibition by antithrombin-heparin. However, the ternary complex reduces the ability of clot-bound thrombin to generate fibrin. These findings highlight the interplay among polyP, thrombin, and fibrin in regulating coagulation.

Background: Elevated plasma mannose-binding lectin (MBL) levels are associated with increased risk of venous thromboembolism (VTE). Although MBL levels are affected by environmental factors, it is mainly genetically regulated.

Objectives: We aimed to investigate the association between genetically predicted MBL and VTE risk in a population-based cohort.

Methods: The study comprised 68 999 individuals from the Trøndelag Health Study (HUNT). Six genetic variants were used to classify individuals in low, medium, and high genetically predicted MBL. Linear regression was applied to estimate whether genetically predicted MBL explained plasma variability of MBL. Cox regression was used to estimate hazard ratios (HRs) with 95% CIs for VTE across genetically predicted MBL categories stratified by sex (men and women) and age groups (reference: low category).

Results: There were 2043 incident VTEs during 12 years' median follow-up. Genetically predicted MBL explained 52.8% of the plasma variability of MBL, and women had lower plasma MBL than men. Genetically predicted MBL was associated with a significantly increased risk of VTE in individuals <60 to 65 years and displayed sex differences. In sex- and age-stratified analysis, men <50 years with high genetically predicted MBL had elevated risk of overall VTE (HR, 2.13; 95% CI, 1.27-3.55), unprovoked VTE (HR, 2.87; 95% CI, 1.28-6.43), and pulmonary embolism (HR, 2.66; 95% CI, 1.11-6.33). In older men and in women, no associations were found.

Conclusion: We found a moderate association between genetically predicted MBL and VTE in young- and middle-aged men. Sex differences and accumulation of environmental factors with age might preclude associations in older men and in women.

Caplacizumab has been shown to reduce time to platelet normalization, reduce thrombotic thrombocytopenic purpura (TTP) exacerbations, and prevent refractory acute disease. We report the first case of caplacizumab "resistance" in a patient with immune TTP with a failure of suppression of in vivo von Willebrand factor (VWF) activity by the drug and persistence of VWF-mediated platelet capture in a flow-based in vitro assay. Genetic analysis revealed a missense variant, P1266L, in exon 28 of the VWF gene that affects the A1-domain binding site of glycoprotein Ib, but it is associated with normal platelet counts and the missense variant V1279I. Failure of caplacizumab therapy resulting in TTP exacerbation or relapse has to date been related to premature stopping of therapy associated with severe ADAMTS-13 (adisintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) deficiency. This case presents a previously unreported phenomenon involving a VWF variant that affects the A1 domain at the caplacizumab-binding site, impairing its effect.

Background: Venous thromboembolism (VTE) is associated with approximately 100 000 deaths annually in the United States (U.S.). Progress in the prevention, diagnosis, treatment, and recovery from VTE depends on research funding. The National Institutes of Health (NIH), the world's largest funder of biomedical research, does not currently report VTE-specific funding in its annual Categorical Spending Report.

Objectives: This study aimed to provide a descriptive analysis of NIH funding for VTE research over the past decade.

Methods: We conducted a search of the NIH Research Portfolio Online Reporting Tools Expenditures and Results database from 2015 to 2024 using a string of VTE-related search terms. Grants were categorized as VTE research (yes/no) using a large language model prompted with predefined classification criteria. We tabulated annual NIH funding amounts, the number of VTE-related grants, and the number of unique principal investigators. For 2023, VTE research investment was compared with that for heart disease and stroke, the leading causes of vascular mortality in the U.S.

Results: The search yielded 2130 grants with complete data, of which 1114 were classified as VTE research. When excluding renewal awards, 490 unique VTE grants were identified. Total inflation-adjusted NIH funding for VTE research was $42 million in 2015, peaked at $73 million in 2021, and totaled $67.1 million in 2024. In 2023, NIH funding per annual deaths was $2765 for heart disease, $2724 for stroke, and $639 for VTE.

Conclusion: NIH investment in VTE research has increased over the past decade, but remains disproportionately low relative to other major causes of vascular mortality in the U.S.

Background: Extracorporeal life support (ECLS) can cause bleeding via excessive cleavage of multimeric protein von Willebrand Factor (VWF) by protease ADAMTS-13, a bleeding diathesis termed acquired von Willebrand syndrome. VWF high-molecular-weight multimers (HMWM) are more procoagulant than smaller multimers. Acute-phase responses generate high levels of VWF-HMWM and reduced ADAMTS-13 activity.

Objectives: Assess whether VWF-HMWM degradation is less pronounced and VWF:collagen binding activity (CB)/VWF:antigen (Ag) ratios are lower during ECLS in acute trauma vs noninjury settings.

Methods: Anesthetized, mechanically ventilated swine (45-60 kg) were randomized to polytrauma (INJ CTRL, n = 14) managed with mechanical ventilation, uninjured receiving veno-venous extracorporeal carbon dioxide removal (ECLS CTRL, n = 20), or polytrauma managed with extracorporeal carbon dioxide removal (ECLS INJ, n = 22). In ECLS groups, 50% of animals received heparin anticoagulation, while 50% received no systemic anticoagulation. VWF:Ag, VWF:CB, VWF multimer size distribution, and ADAMTS-13 activity were evaluated at baseline, post-injury/intervention, and at 6, 24, 48, and 72 hours.

Results: VWF-HMWM were depleted at 72 hours in ECLS CTRL (-19 ± 3%, P < .0001), but not in ECLS INJ (-10 ± 5%, P = .09). VWF:CB/VWF:Ag ratios relative to baseline were reduced at 48 to 72 hours in ECLS CTRL (0.84 ± 0.04, P = .002) and ECLS INJ (0.69 ± 0.05, P < .001) but not INJ CTRL. ADAMTS-13 activity decreased in all groups. Animals treated with heparin showed less pronounced VWF-HMWM degradation, higher VWF:CB/VWF:Ag ratios, and higher ADAMTS-13 activity vs matched non-anticoagulated animals.

Conclusion: Trauma managed with ECLS was associated with reduced VWF:CB/VWF:Ag ratios and lower ADAMTS-13 activity, but VWF-HMWM depletion was minimal. Thus, potential for acquired von Willebrand syndrome is a concern for trauma patients undergoing ECLS. Heparin partially countered effects of ECLS on VWF and ADAMTS-13.

Background: Venous thromboembolism (VTE) is a common complication of cancer. Complex interactions between tumor biology and the hemostatic system contribute to development of cancer-associated thrombosis.

Objectives: This study examined associations of somatic mutations with VTE in a large multicancer cohort.

Methods: We analyzed paired tumor and germline whole genome sequence data and electronic health records from 12 507 cancer patients recruited to the Genomics England National Genomic Research Library, to evaluate associations of somatic mutations across 608 genes, tumor mutational burden (TMB), and 25 single-base substitution (SBS) mutational signatures with VTE.

Results: In multivariable Cox regressions adjusted for age, sex, and genetic ancestry, somatic mutations in 4 genes associated with higher rates of VTE at a false-discovery rate of <0.1: CDKN2A (hazard ratio [HR], 1.62; 95% CI, 1.23-2.13), KRAS (HR, 1.31; 95% CI, 1.12-1.53), PCDH15 (HR, 1.48; 95% CI, 1.24-1.76), and TP53 (HR, 1.55; 95% CI, 1.38-1.73). TMB of ≥20 mutations/Mb and 2 DNA mismatch repair signatures (SBS6 and SBS26) associated with lower rates of VTE. Evidence for these associations remained robust after additional adjustment for tumor type, stage, and systemic anticancer treatment. Assessment of interactions between somatic mutations and germline genetic risks for VTE demonstrated that heterozygous carriers of factor V Leiden (rs6025) with somatic PCDH15 mutations had a 6-month VTE incidence of 13% (95% CI, 6%-19%) compared with 4% (95% CI, 3%-5%) in rs6025 carriers without PCDH15 mutations.

Conclusions: These findings support the hypothesis that tumor somatic mutations influence risk of VTE. This may provide insights into the pathophysiology of cancer-associated thrombosis and inform future efforts to improve clinical risk prediction.

Background: Oral anticoagulants are commonly used for venous thromboembolism (VTE), but the optimal treatment duration remains uncertain, highlighting the need for personalized strategies based on individual risk.

Objectives: We developed predictive models to assess VTE recurrence and major bleeding risks during and after anticoagulant treatment, incorporating time-varying data.

Methods: This study included 14,653 patients with VTE who received oral anticoagulants between July 2018 and December 2020 using a claims database. Two cohorts were constructed: an on-treatment cohort, segmented into 90-day intervals for dynamic updates, and an off-treatment cohort. VTE and major bleeding were identified using diagnosis codes, hospitalization records, and procedure data. Three predictive models were developed: models 1 and 2 predicted 90-day risks of VTE recurrence and major bleeding during treatment, and model 3 predicted VTE recurrence after discontinuation. Time-varying effects were identified through interaction-term testing. Models were developed using least absolute shrinkage and selection operator regression, with optimism-corrected areas under the receiver operating characteristic curves (AUROCs) assessed via bootstrapping.

Results: The incidence proportions were 2.7% for on-treatment VTE recurrence, 1.3% for major bleeding, and 0.8% for VTE recurrence within 90 days after discontinuation. The final models showed optimism-corrected AUROCs of 0.715, 0.828, and 0.679 for models 1, 2, and 3, respectively. Calibration was suboptimal for models 2 and 3.

Conclusion: These models may provide supportive information for individualized treatment decisions to dynamically assess VTE recurrence and bleeding risks, but their clinical value must be confirmed through external validation and recalibrated in larger cohorts before clinical use.

Thrombotic thrombocytopenic purpura (TTP) is a thrombotic microangiopathy caused by severe deficiency of the metalloprotease ADAMTS13. TTP has 2 subtypes: immune (iTTP) and congenital (cTTP). Autoimmune mechanisms underlie iTTP; however, the genetic factors influencing disease risk, relapse risk, and response to treatment are incompletely understood. In cTTP, although variants are described throughout the ADAMTS13 gene, the effect of many of these variants is uncertain. We conducted a comprehensive literature review of both iTTP and cTTP, incorporating findings from genome-wide association studies, case-control studies, registry publications, and population-level databases. In iTTP, 9 studies have investigated the effect of HLA alleles, with HLA-DRB1∗11 most consistently associated with an increased risk, and HLA-DRB1∗04 with protection. However, effect sizes varied across ancestral populations, as we highlight in a meta-analysis of available studies. Beyond the HLA locus, a genome-wide association study in iTTP patients with European ancestry identified a locus on chromosome 3 (rs9884090) associated with a reduced iTTP risk. In cTTP, we identified 364 variants within the literature, the majority (199, 54.7%) being missense variants within coding regions. Although variants are located across the gene, the highest density of variants was observed within exon 7 (32 variants, 8.8% of the total), corresponding to the metalloprotease domain. Finally, analysis of population-level constraint data from gnomAD offered additional insight into the tolerance of ADAMTS13 to variation. Together, these findings highlight the complexity of the genetic factors influencing TTP, and the value of combining clinical information and population data to increase understanding of the disease.

Background: Obstructive prosthetic valve thrombosis (PVT) is a life-threatening complication. The first-line therapy remains uncertain. While surgery has historically been preferred, advances in low-dose, slow-infusion fibrinolysis protocols have improved outcomes.

Objectives: To evaluate the efficacy and safety of fibrinolytic therapy versus urgent valve surgery in PVT.

Methods: Following a systematic search of 5 databases, a meta-analysis compared fibrinolysis with urgent valve surgery for obstructive PVT. The primary outcomes were in-hospital all-cause mortality and complete restoration of valve function. Secondary outcomes included stroke, systemic embolism, major bleeding, recurrent PVT, and all-cause mortality during follow-up. Data were pooled as risk ratio (RR) using random-effects models, with sensitivity and meta-regression analyses.

Results: Across 12 observational studies and 1 randomized controlled trial, 1300 patients (fibrinolysis/surgery: 714/586) were included. No significant difference was observed in in-hospital mortality (RR: 0.59 [0.27-1.30]; I²=57.1%). However, fibrinolysis was associated with lower complete restoration of valve function (RR: 0.82 [0.70-0.96]; I²=71.6%) and higher risks of stroke (RR: 3.19 [1.30-7.85]; I²=0%), systemic embolism (RR: 3.88 [1.44-10.4]; I²=0%), and recurrent PVT (RR: 2.44 [1.18-5.05]; I²=58.3%). No differences were found in major bleeding or all-cause mortality during follow-up. Sensitivity analyses restricted to alteplase-based regimens favored fibrinolysis, showing lower in-hospital mortality (RR: 0.12 [0.05-0.29]; I²=0%), consistent with meta-regression findings, and efficacy comparable to surgery.

Conclusion: Our study suggests that surgery offers higher immediate success with fewer embolic or recurrent events. However, contemporary alteplase protocols may be associated with improved safety outcomes relative to surgery, with no apparent loss of efficacy. Given the low certainty of available evidence, treatment should be individualized according to patient risk profile, and institutional resources.

Background: The thrombin generation assay (TGA) is a global coagulation test able to determine the amount of thrombin formed in plasma samples. TGA has been shown to be useful to monitor the effect of procoagulant and anticoagulation therapies in patients with bleeding diatheses or an increased thrombosis risk.

Objectives: This study describes a new miniaturized platform to measure chemiluminescent-based microfluidic thrombin generation assay (TGA_lum).

Methods: In TGA_lum, generated thrombin cleaves a β-Ala-Gly-Arg tripeptide from a caged aminoluciferin moiety, which almost instantaneously results in the generation of a photon by luciferase. The number of photons generated is proportional to the activity of thrombin in the microfluidic TGA_lum system. By printing the assay components, a microfluidic cartridge was functionalized for TGA_lum. By only applying 20 μL of citrated plasma to the microfluidic cartridges, thrombin generation was monitored successfully with the characteristic peak profile in 110 nL detection chambers using a prototype readout device.

Results: The TGA_lum was shown to be sensitive to therapeutic levels of anticoagulants (direct oral anticoagulants [DOACs]) to prevent thrombosis, as well as to the reversal effects of the antidotes thereof. Furthermore, TGA_lum was able to detect the defect in the plasma of patients with hemophilia A (pwHA), as well as the hemostatic replacement effect of factor VIII infusion in pwHA.

Conclusion: This study provides proof-of-concept that the present microfluidic TGA_lum is applicable for monitoring therapy of pwHA at a global level and monitoring anticoagulant therapy with DOACs using a portable readout device.