Journal of Thrombosis and Haemostasis最新文献_第7页

The mitochondrial calcium uniporter regulates calcium dynamics to drive platelet function, bioenergetics, and thrombosis 线粒体钙单转运体调节钙动力学以驱动血小板功能、生物能量学和血栓形成。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.019

Madankumar Ghatge , Gagan D. Flora , Rakesh B. Patel , Manasa K. Nayak , Mariia Kumskova , Tam Nguyen , Yuriy M. Usachev , Anil K. Chauhan

Background

The mitochondrial calcium uniporter (MCU), a selective intracellular calcium (Ca²⁺) channel, mediates mitochondrial Ca²⁺ uptake, supporting Ca²⁺ homeostasis and mitochondrial bioenergetics. While cytosolic Ca²⁺ flux from the dense tubular system (DTS) and store-operated Ca²⁺ entry are known to drive platelet activation, the role of mitochondrial Ca²⁺ handling in platelet function and thrombosis is not well understood.

Objectives

This study examined whether targeting MCU-dependent Ca²⁺ flux could attenuate platelet activation and arterial thrombosis.

Methods

Susceptibility to arterial thrombosis was assessed using the FeCl₃-induced carotid injury model in wild-type and MCU^−/− mice. Mitochondrial and cytosolic Ca²⁺ levels were measured in Rhod-2– and Fura-2–loaded platelets by fluorometry, and platelet bioenergetics were analyzed using a Seahorse extracellular flux analyzer.

Results

Genetic ablation of MCU inhibited agonist-induced platelet functions, including aggregation, fibrinogen binding to integrin α_IIbβ₃, granule secretion, and spreading on fibrinogen. MCU^−/− mice were less susceptible to in vivo arterial thrombosis with unaltered tail bleeding time, suggesting normal hemostasis. Mechanistically, these effects were associated with disruption of Ca²⁺ homeostasis mediated by reduced mitochondrial Ca²⁺ uptake, altered release of Ca²⁺ from dense tubular system, and impaired store-operated Ca²⁺ entry in agonist-stimulated MCU^−/− platelets. Consistent with this, Ca²⁺-dependent glycoprotein VI signaling events such as phospholipase γ2 and protein kinase C substrate phosphorylation were significantly reduced in collagen-stimulated MCU^−/− platelets. Furthermore, disruption of mitochondrial Ca²⁺ uptake significantly impaired mitochondrial respiration and associated adenosine triphosphate production in agonist-stimulated MCU^−/− platelets.

Conclusion

MCU facilitates platelet activation and thrombosis by regulating calcium flux (mitochondrial and cytosolic), thereby establishing its potential as a target for antithrombotic therapeutic intervention.

背景：线粒体钙单转运体（MCU）是一种选择性Ca2+通道，介导线粒体Ca2+摄取，支持Ca2+稳态和线粒体生物能量学。虽然已知来自致密管系统（DTS）和储存操作的Ca2+进入（SOCE）的细胞质Ca2+通量驱动血小板活化，但线粒体Ca2+处理在血小板功能和血栓形成中的作用尚不清楚。目的：探讨靶向mcu依赖性Ca2+通量是否能减弱血小板活化和动脉血栓形成。方法：采用fecl3诱导的WT和MCU-/-小鼠颈动脉损伤模型，评估小鼠对动脉血栓形成的易感性。通过荧光法测定Rhod-2和fura -2负载血小板中的线粒体和胞质Ca2+水平，并使用海马细胞外通量分析仪分析血小板生物能量学。结果：基因消融MCU抑制激动剂诱导的血小板功能，包括聚集、纤维蛋白原与整合素α ib β3结合、颗粒分泌和纤维蛋白原扩散。MCU-/-小鼠在体内动脉血栓形成的易感程度较低，尾出血时间不变，提示止血正常。在机制上，这些影响与Ca2+稳态的破坏有关，这些破坏是由线粒体Ca2+摄取减少、DTS Ca2+释放改变以及激动剂刺激的MCU-/-血小板中SOCE受损介导的。与此一致，Ca2+依赖性GPVI信号事件，如plc γ - 2和PKC底物磷酸化，在胶原刺激的MCU-/-血小板中显着减少。此外，线粒体Ca2+摄取的破坏显著损害了激动剂刺激的MCU-/-血小板的线粒体呼吸和相关ATP的产生。结论：MCU通过调节钙通量（线粒体和细胞质）促进血小板活化和血栓形成，从而确立了其作为抗血栓治疗干预靶点的潜力。

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引用次数: 0

Engineering a membrane-independent human prothrombinase through parsimonious mutation of factor Xa 通过Xa因子的简约突变构建膜非依赖性人凝血酶原。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.015

Fatma Işık Üstok, James A. Huntington

Background

Thrombin is generated from its precursor prothrombin by sequential cleavage at Arg320 and Arg271 by the prothrombinase complex, composed of factor (F)Xa and FVa on phospholipid (PL) surfaces. The affinity of human FXa for FVa is low in the absence of PL. However, FXa orthologs from the venom of group D snakes bind to FVa with high affinity, forming an active complex without PL. We previously characterized the properties of the FXa ortholog hopsarin D (HopD) from Hoplocephalus stephensii.

Objectives

In this study, we set out to create a PL-independent human prothrombinase by making mutations to FXa, guided by a model of the prothrombinase complex and the sequence differences between human FXa and HopD.

Methods

We assessed the contribution of individual domains of FXa to its binding to FVa by swapping each with the corresponding domain of HopD. We then chose 3 loops in the serine protease domain predicted to be in contact with FVa to swap to those of HopD. Eventually, 10 residues from the 3 loops in the serine protease domain and 7 from the epidermal growth factor 2 domain were selected for mutation.

Results

The resulting M17 FXa variant bound to FVa with a dissociation constant of 21 nM, similar to HopD, and together efficiently processed prothrombin through the meizothrombin intermediate in the absence of PL.

Conclusion

We conclude that the role of PL membranes in prothrombinase assembly and function is limited to improving the affinity of FXa for FVa and that the M17-FVa complex is likely to be structurally equivalent to human prothrombinase.

背景：凝血酶是由其前体凝血酶原在磷脂（PL）表面上由因子(f) Xa和fVa组成的凝血酶原复合物在Arg320和Arg271处连续切割而产生的。在没有PL的情况下，人类fXa对fVa的亲和力较低。然而，D组蛇毒中的fXa同源物与fVa结合的亲和力很高，形成了一个没有PL的活性复合物。我们之前表征了来自stefan Hoplocephalus的fXa同源物Hopsarin D （HopD）的性质。目的：在这里，我们开始通过对fXa进行突变来创建一个pl独立的人类凝血酶原，在凝血酶原复合物模型和人类fXa和HopD之间的序列差异的指导下。方法：通过与HopD的相应结构域交换，我们评估了fXa的各个结构域对其与fVa结合的贡献。然后，我们选择了三个丝氨酸蛋白酶（SP）结构域中预测与fVa接触的环，以交换到HopD的环。最终，从SP结构域的3个环中选出10个残基，从EGF2结构域选出7个残基进行突变。结果：得到的M17 fXa变体与fVa结合，Kd值为21 nM，与HopD相似，并且在没有PL的情况下通过减数凝血酶中间体有效地加工凝血酶原。结论：我们得出PL膜在凝血酶原组装和功能中的作用仅限于提高fXa对fVa的亲和力，并且M17-fVa复合物可能在结构上与人凝血酶原相当。

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引用次数: 0

Additional oxerutin therapy to promote deep vein thrombus resolution (RESOLVE-DVT): a randomized controlled pilot trial 额外的奥施罗汀治疗促进深静脉血栓溶解（Resolve-DVT）：一项随机对照试验。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.018

Aaron F.J. Iding , Ruben D. Hupperetz , Rutger J.B. Brans , Hugo ten Cate , Daan J.L. van Twist , Arina J. ten Cate-Hoek

Background

Postthrombotic syndrome (PTS) is a chronic complication of deep vein thrombosis (DVT), with residual venous obstruction (RVO) as a key contributing factor. Oxerutin, a venoactive drug, has the potential to promote DVT resolution, thereby reducing RVO and possibly preventing PTS.

Objectives

This pilot study aimed to estimate the effect of oxerutin therapy on RVO.

Methods

A single-center, patient-unblinded, physician-blinded, assessor-blinded, randomized controlled trial was conducted in adults with acute proximal DVT. Patients were randomized within 48 hours to receive 2-month oxerutin therapy plus standard treatment or standard treatment alone. The primary outcome was RVO at 3 months, defined as a transversal diameter ≥2mm on compressive ultrasound, with sensitivity analysis at ≥4 mm. Secondary outcomes included PTS at 3 months and biomarkers at 5 timepoints. The study was approved by the ethics committee. All patients gave written informed consent.

Results

A total of 44 patients were randomized to oxerutin or standard treatment. At 3 months, the proportion of RVO was lower in the oxerutin group, both for ≥2 mm (42.9% vs 59.1%; odds ratio [OR], 0.34; 95% CI, 0.08-1.28) and ≥4 mm (28.6% vs 54.5%; OR, 0.21; 95% CI, 0.04-0.85), adjusted for DVT extent. A lower proportion of PTS was observed in the oxerutin group (28.6% vs 40.9%; OR, 0.38; 95% CI, 0.08-1.53). Biomarker levels of intercellular adhesion molecule-1 and interleukin-6 were persistently lower in the oxerutin group.

Conclusion

Oxerutin therapy might reduce RVO by promoting DVT resolution. These preliminary findings support further investigation in a large-scale trial to assess the long-term prevention of PTS.

背景：血栓形成后综合征（Post-thrombotic syndrome， PTS）是深静脉血栓形成（deep vein thrombosis， DVT）的一种慢性并发症，其中残余静脉阻塞（residual venous梗阻，RVO）是主要致病因素。Oxerutin是一种静脉活性药物，具有促进DVT消退的潜力，从而降低RVO并可能预防PTS。目的：本初步研究旨在评估奥施罗汀治疗对RVO的影响。方法：对成人急性近端深静脉血栓形成患者进行单中心、非盲法、医师盲法、评估盲法随机对照试验。患者在48小时内随机接受两个月的奥施罗汀治疗加标准治疗或单独标准治疗。主要终点是3个月时的RVO，定义为压缩超声的横径≥2mm，敏感性分析≥4mm。次要终点包括3个月时的PTS和5个时间点的生物标志物。这项研究得到了伦理委员会的批准。所有患者均给予书面知情同意。结果：44例患者随机分为奥施罗汀组和标准组。在3个月时，奥赛芦丁组的RVO比例较低，≥2mm （42.9% vs 59.1%；比值比[OR] 0.34； 95%可信区间[CI] 0.08-1.28）和≥4mm (28.6% vs 54.5%; OR 0.21; 95% CI 0.04-0.85)，根据DVT程度进行调整。奥赛芦丁组PTS发生率较低（28.6% vs 40.9%; OR 0.38; 95% CI 0.08-1.53）。oxerutin组细胞间粘附分子-1和白细胞介素-6的生物标志物水平持续降低。结论：奥施罗汀治疗可能通过促进DVT消退而降低RVO。这些初步发现支持在大规模试验中进一步调查以评估PTS的长期预防。

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引用次数: 0

Evaluation of the updated ABC-AF-bleeding score 2.0 in patients with atrial fibrillation treated with a direct oral anticoagulant or warfarin 直接口服抗凝剂或华法林治疗的心房颤动患者最新abc - af出血评分2.0的评估

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.09.032

Ziad Hijazi , Johan Lindbäck , Jonas Oldgren , John H. Alexander , Alexander P. Benz , David D. Berg , Anthony P. Carnicelli , John W. Eikelboom , Robert P. Giugliano , Shinya Goto , Christopher B. Granger , Renato D. Lopes , Christian T. Ruff , Agneta Siegbahn , David A. Morrow , Lars Wallentin

Background

Oral anticoagulation (OAC) reduces stroke in patients with atrial fibrillation (AF), but increases bleeding.

Objectives

This study aimed to evaluate an updated version of the Age, Biomarkers, and Clinical history of bleeding in AF (ABC-AF)-bleeding score (2.0) including consideration of OAC type (direct oral anticoagulant [DOAC] or warfarin) and compare its performance with other bleeding risk scores in 25 962 patients from the COMBINE AF cohort.

Methods

The COMBINE AF biomarker cohort contains individual participant data from patients with AF enrolled in 3 pivotal randomized trials comparing DOACs with warfarin. The biomarkers in the ABC-AF-bleeding score (growth differentiation factor 15, hemoglobin, and troponin-T) were analyzed in baseline samples. The biomarker-based ABC-AF-bleeding score was updated (version 2.0) by incorporating OAC type into the model (DOAC or warfarin). Discrimination was assessed by Harrell C-index and compared with clinically based bleeding scores; HAS-BLED (Hypertension, Abnormal renal/liver function, Stroke, Bleeding history or predisposition, Labile INR, Elderly, Drugs/alcohol), DOAC, and ORBIT (Older age, Reduced haemoglobin/haematocrit or history of anaemia, Bleeding history, Insufficient renal function, Treatment with antiplatelet agents).

Results

During follow-up, 1321 patients (5.1%) had an International Society on Thrombosis and Haemostasis major bleeding event, including 480 gastrointestinal, and 248 intracranial hemorrhages. The ABC-AF-bleeding 2.0 risk score showed better discrimination and calibration than the original version and provided superior discrimination than clinical risk scores for all outcomes. The ABC-AF-bleeding score 2.0 C-indices for major bleeding were 0.69 (95% CI, 0.68-0.71); gastrointestinal bleeding, 0.72 (95% CI, 0.69-0.74); and intracranial bleeding, 0.66 (95% CI, 0.63-0.70). The ABC-AF-bleeding score 2.0 also provided consistent superior discrimination in clinically relevant subgroups.

Conclusion

The updated ABC-AF-bleeding score 2.0 provided better discrimination and calibration for the risk of major bleeding than clinical risk scores, which was consistent across multiple subgroups. These findings support the utility of the ABC-AF-bleeding score for advancing precision medicine in AF.

背景：腹侧抗凝（OAC）可减少房颤（AF）患者的卒中，但会增加出血。本研究旨在评估AF患者年龄、生物标志物和出血的临床史(ABC-AF)-出血评分（2.0）的更新版本，包括OAC类型（直接口服抗凝剂[DOAC]或华法林），并将其与其他出血风险评分进行比较。方法：COMBINE房颤生物标志物队列包含了3项比较DOACs与华法林的关键随机试验中房颤患者的个体参与者数据。在基线样本中分析abc - af出血评分中的生物标志物（生长分化因子15、血红蛋白和肌钙蛋白- t）。通过将OAC类型（DOAC或华法林）纳入模型，更新了基于生物标志物的abc - af出血评分（2.0版）。采用Harrell c指数评估辨别性，并与临床出血评分进行比较；ha -出血（高血压、肾/肝功能异常、中风、出血史或易感、INR不稳定、老年人、药物/酒精）、DOAC和ORBIT（年龄较大、血红蛋白/红细胞压积降低或贫血史、出血史、肾功能不足、抗血小板药物治疗）。结果随访期间，1321例（5.1%）患者发生国际血栓与止血学会大出血事件，其中胃肠道出血480例，颅内出血248例。与原始版本相比，abc - af -出血2.0风险评分具有更好的辨别和校准能力，并且在所有结果上都比临床风险评分具有更好的辨别能力。abc - af -出血评分2.0 c -大出血指数为0.69 (95% CI, 0.68-0.71)；胃肠道出血，0.72 (95% CI, 0.69-0.74)；颅内出血，0.66 （95% CI, 0.63-0.70）。abc - af -出血评分2.0在临床相关亚组中也提供了一致的优势区分。结论更新后的abc - af -出血评分2.0比临床风险评分能更好地区分和校准大出血风险，且在多个亚组中是一致的。这些发现支持abc -AF出血评分在推进房颤精准医学方面的应用。

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引用次数: 0

Preclinical safety and bleeding evaluation in swine for a small interfering RNA-lipid nanoparticle that prevents excess fibrinogen synthesis sirna -脂质纳米颗粒预防过量纤维蛋白原合成的临床前安全性和猪出血评估。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.09.012

Monica S. Seadler , Francesca Ferraresso , Halen M. Turner , Laura M. Ketelboeter , Youjie Zhang , Katherine Badior , William G. Hayssen , Taylor Chen , Amber L. Haugen , Massimo F. Cau , Madelaine Robertson , Chad Skaer , Muskan Bansal , Patrick B. Murphy , Marc de Moya , Mitchell R. Dyer , Christian J. Kastrup

Background

Fibrinogen levels can drastically increase from inflammation, trauma, or surgery, which increases risk of venous thromboembolism (VTE). We developed small interfering RNA (siFibrinogen) that decreased fibrinogen production and thrombosis in rodent models, which became effective hours after administration and knockdown lasted over a week. Here, we tested whether knockdown of fibrinogen was feasible and safe in a preclinical large animal model. We hypothesized that fibrinogen could be controllably knocked down to levels that still enable hemostasis and avoid limitations of the current standard of care agent, low-molecular-weight heparin (LMWH).

Objectives

This study evaluated the preclinical safety and feasibility of a novel therapy for VTE prophylaxis for complications in trauma and surgical patients.

Methods

Female Yorkshire/cross swine (7-15 kg) were infused with small interfering RNA targeting fibrinogen (siFibrinogen), LMWH, or a vehicle control. Hemostasis was assessed in an established model of hemorrhagic shock. Reversibility was evaluated by administering fibrinogen concentrate.

Results

Circulating fibrinogen concentrations decreased in swine in a dose-dependent manner, lasting over a week from 1 injection of siFibrinogen. Fibrinogen reduction to levels > 0.4 g/L did not impair hemostasis during hemorrhagic shock when compared with LMWH or vehicle control. The effects of siFibrinogen were reversed by the administration of fibrinogen concentrate. No infusion-related reactions or toxicity was observed.

Conclusion

siFibrinogen represents a promising novel approach for VTE prophylaxis, avoiding limitations of LMWH. siFibrinogen administered early in patient care could decrease fibrinogen in a sustained and predictable manner to prevent thrombosis while preserving hemostasis.

背景：纤维蛋白原水平可急剧增加炎症，创伤，或手术，这驱动静脉血栓栓塞（VTE）的风险增加。我们开发了小干扰RNA (siFibrinogen)，它可以减少啮齿动物模型中纤维蛋白原的产生和血栓形成，在给药后几小时内生效，敲除持续超过一周。在这里，我们在临床前大型动物模型中测试了敲除纤维蛋白原是否可行和安全。我们假设纤维蛋白原可以被控制地降低到仍然能够止血的水平，并避免当前标准护理药物低分子肝素（LMWH）的局限性。目的：为了预防创伤和外科患者的并发症，本研究评估了一种预防静脉血栓栓塞（VTE）的新疗法的临床前安全性和可行性。方法：雌性约克郡/杂交猪（7- 15kg）注射sifibrin原、低分子肝素或对照。在建立的失血性休克模型中评估止血情况。通过给予纤维蛋白原浓缩物来评估可逆性。结果：猪体内循环纤维蛋白原呈剂量依赖性下降，注射一次纤维蛋白原后持续一周以上。与低分子肝素或对照组相比，纤维蛋白原降低到0.4 g/L以上的水平不会损害失血性休克期间的止血。sifibrin原的作用随着纤维蛋白原浓缩物的施用而逆转。未观察到输注相关反应或毒性。结论：siFibrinogen代表了一种很有前途的静脉血栓栓塞预防新方法，避免了低分子肝素的局限性。在患者护理早期给予纤维蛋白原可以持续和可预测地降低纤维蛋白原，以防止血栓形成，同时保持止血。

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引用次数: 0

Frequency of, and prognostic factors for, residual thrombosis and complete veno-occlusion following anticoagulation for provoked venous thromboembolism in patients <21 years: findings from the Kids-DOTT Multinational Trial <21岁患者诱发性静脉血栓栓塞（VTE）抗凝后残留血栓形成和完全静脉闭塞的频率和预后因素：来自儿童dott多国试验的发现

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.007

Rukhmi Bhat , Anjali Sharathkumar , Riten Kumar , Fernando F. Corrales-Medina , Heleen van Ommen , Mihir Bhatt , Smita Dandekar , Maua Mosha , Neil A. Goldenberg

Background

Prognostic factors for residual thrombosis (RT) and complete veno-occlusion (CVO) after 6 weeks of anticoagulation in pediatric acute VTE remain unknown.

Objectives

This study assessed the frequency and predictors of RT and CVO through a prespecified secondary analysis of the Kids-DOTT randomized clinical trial.

Methods

Per trial protocol, RT and CVO were radiologically assessed 6 weeks after acute VTE diagnosis. Univariate logistic regression analyses were performed for both outcomes, and variables with a P value of .05 were included in a multivariable models, where a P value of <.05 denoted statistical significance.

Results

Among 532 enrolled patients, 28.8% demonstrated RT and 12.6% CVO after 6 weeks of treatment. Age and gender distributions did not differ significantly by RT and CVO status. In multivariable analysis, cerebral venous sinus location of VTE (odds ratio [OR], 2.52; 95% CI, 1.22-5.21; P = .01) and comorbid infection (OR, 1.61; 95% CI, 1.00-2.58; P = .049) were independently associated with RT. Internal jugular vein (OR, 3.97; 95% CI, 1.26-12.48; P = .02) and lower extremity VTE (OR, 2.28; 95% CI, 1.01-5.15; P = .046) were independently associated with CVO.

Conclusion

Approximately 29% of young patients developed RT, and 13% CVO, after 6 weeks of anticoagulation. VTE location and comorbid infection were identified as predictors for RT and CVO. These findings highlight the need for further studies on long-term outcomes of RT and CVO, particularly their impact on postthrombotic syndrome development and quality-of-life measures in pediatric VTE.

背景：儿童急性静脉血栓栓塞（VTE）抗凝治疗6周后残余血栓形成（RT）和完全静脉闭塞（CVO）的预后因素尚不清楚。本研究通过预先指定的Kids-DOTT随机临床试验的二次分析来评估RT和CVO的频率和预测因素。方法：根据试验方案，在急性静脉血栓栓塞诊断后6周，对RT和CVO进行放射学评估。对两种结果进行单因素logistic回归分析，并将p值为0.05的变量纳入多变量模型，其中p值为结果：在532名入组患者中，治疗6周后，28.8%的患者显示RT， 12.6%的患者显示CVO。年龄和性别分布在RT和CVO状态上无显著差异。在多变量分析中，VTE的脑静脉窦位置（OR 2.52, 95% CI=1.22-5.21, p=0.01）和共病感染（OR 1.61, 95% CI=1.00-2.58, p=0.049）与rt独立相关。颈内静脉（OR 3.97, 95% CI=1.26-12.48, p=0.02）和下肢VTE （OR 2.28, 95% CI=1.01-5.15, p= 0.046）与CVO独立相关。结论：大约29%的年轻患者在抗凝治疗6周后出现RT， 13%出现CVO。静脉血栓栓塞位置和合并症感染被确定为RT和CVO的预测因素。这些发现强调需要进一步研究RT和CVO的长期结果，特别是它们对儿科静脉血栓栓塞综合征后发展和生活质量措施的影响。

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引用次数: 0

von Willebrand factor-T441N, a novel missense variant in the von Willebrand factor D2 domain, exhibits impaired multimerization and deformed Weibel-Palade bodies VWF- t441n是VWF D2结构域的一种新型错义变体，表现出多聚性受损和韦贝尔-帕拉德体变形。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.017

Shuichi Okamoto , Shogo Tamura , Atsuo Suzuki , Nobuaki Suzuki , Takeshi Kanematsu , Naruko Suzuki , Masashi Tomita , Fumihiko Hayakawa , Akira Katsumi , Hitoshi Kiyoi , Tetsuhito Kojima , Tadashi Matsushita

Background

von Willebrand factor (VWF) propeptide (VWFpp) located in the D1-D2 domain of pro-VWF, is essential for multimerization and intracellular storage of VWF. Pathogenic variants of VWFpp cause various subtypes of von Willebrand disease, such as type 1, type 2A, type 2N, and type 3. In heterozygotes of these variants, VWFpp derived from the normal allele can rescue the abnormalities observed in the homozygote. This rescue effect may highlight the phenotypic difference between heterozygotes and homozygotes.

Objectives

We investigated the molecular pathogenesis of a novel missense variant of VWFpp, VWF c.1322 C>A (p.T441N), identified in a heterozygous 20-year-old woman.

Methods

The VWF gene was analyzed using whole-exome sequencing and Sanger sequencing. Recombinant VWF (rVWF) and VWF propeptide (rVWFpp) were overexpressed in HEK293 cells. rVWF production was measured using an in-house enzyme-linked immunosorbent assay and multimer analysis. The morphology of pseudo-Weibel–Palade bodies (WPBs) and WPBs in endothelial colony-forming cells were observed by immunocytochemistry.

Results

In the patient’s plasma, the VWF antigen level was in the normal range, but VWF ristocetin cofactor activity was slightly decreased, by 0.38 IU/mL, and high-molecular-weight and intermediate-molecular-weight multimers were decreased. Pseudo-WPBs were also shortened and rounded, corresponding with patient-derived endothelial colony-forming cells. These abnormalities were partially rescued by coexpression of wild-type rVWFpp on separate vectors.

Conclusion

VWF-T441N results in impaired multimerization and deformed WPBs. In heterozygous VWF-T441N, these abnormalities are partially rescued by VWFpp derived from the normal allele. The observed rescue effect provides further insights for understanding the phenotypic heterogeneity of VWFpp variants.

背景：血管性血友病因子（VWF）前肽（VWFpp）位于VWF前体的D1-D2结构域，对VWF的多聚和细胞内储存至关重要。VWFpp的致病变异可导致血管性血友病（VWD）的各种亚型，如1型、2A型、2N型和3型。在这些变异的杂合子中，来自正常等位基因的VWFpp可以挽救纯合子中观察到的异常。这种拯救效应可能突出了杂合子和纯合子之间的表型差异。目的：研究一种新的VWFpp错义变异VWF c.1322的分子发病机制C >a (p.T441N)，在一名杂合的20岁女性中发现。方法：采用全外显子组测序和Sanger测序对VWF基因进行分析。重组VWF （rVWF）和VWF前肽（rVWFpp）在HEK293细胞中过表达。rVWF的产生采用内部酶联免疫吸附试验和多点分析来测量。采用免疫细胞化学方法观察内皮细胞集落形成细胞（ECFCs）中伪韦贝尔-帕拉德小体（WPBs）和伪韦贝尔-帕拉德小体的形态。结果：患者血浆中VWF抗原水平在正常范围内，但VWF里斯托肽辅因子活性略有下降，下降幅度为0.38 IU/mL，高、中分子量多聚体降低。伪wpb也缩短和变圆，与患者来源的ecfc相对应。野生型rVWFpp在不同载体上的共表达部分挽救了这些异常。结论：VWF-T441N可导致细胞多化功能受损和小波细胞变形。在杂合子VWF-T441N中，这些异常被来自正常等位基因的VWFpp部分挽救。观察到的拯救效应为理解VWFpp变异的表型异质性提供了进一步的见解。

{"title":"von Willebrand factor-T441N, a novel missense variant in the von Willebrand factor D2 domain, exhibits impaired multimerization and deformed Weibel-Palade bodies","authors":"Shuichi Okamoto , Shogo Tamura , Atsuo Suzuki , Nobuaki Suzuki , Takeshi Kanematsu , Naruko Suzuki , Masashi Tomita , Fumihiko Hayakawa , Akira Katsumi , Hitoshi Kiyoi , Tetsuhito Kojima , Tadashi Matsushita","doi":"10.1016/j.jtha.2025.10.017","DOIUrl":"10.1016/j.jtha.2025.10.017","url":null,"abstract":"<div><h3>Background</h3><div>von Willebrand factor (VWF) propeptide (VWFpp) located in the D1-D2 domain of pro-VWF, is essential for multimerization and intracellular storage of VWF. Pathogenic variants of VWFpp cause various subtypes of von Willebrand disease, such as type 1, type 2A, type 2N, and type 3. In heterozygotes of these variants, VWFpp derived from the normal allele can rescue the abnormalities observed in the homozygote. This rescue effect may highlight the phenotypic difference between heterozygotes and homozygotes.</div></div><div><h3>Objectives</h3><div>We investigated the molecular pathogenesis of a novel missense variant of VWFpp, <em>VWF</em> c.1322 C>A (p.T441N), identified in a heterozygous 20-year-old woman.</div></div><div><h3>Methods</h3><div>The <em>VWF</em> gene was analyzed using whole-exome sequencing and Sanger sequencing. Recombinant VWF (rVWF) and VWF propeptide (rVWFpp) were overexpressed in HEK293 cells. rVWF production was measured using an in-house enzyme-linked immunosorbent assay and multimer analysis. The morphology of pseudo-Weibel–Palade bodies (WPBs) and WPBs in endothelial colony-forming cells were observed by immunocytochemistry.</div></div><div><h3>Results</h3><div>In the patient’s plasma, the VWF antigen level was in the normal range, but VWF ristocetin cofactor activity was slightly decreased, by 0.38 IU/mL, and high-molecular-weight and intermediate-molecular-weight multimers were decreased. Pseudo-WPBs were also shortened and rounded, corresponding with patient-derived endothelial colony-forming cells. These abnormalities were partially rescued by coexpression of wild-type rVWFpp on separate vectors.</div></div><div><h3>Conclusion</h3><div>VWF-T441N results in impaired multimerization and deformed WPBs. In heterozygous VWF-T441N, these abnormalities are partially rescued by VWFpp derived from the normal allele. The observed rescue effect provides further insights for understanding the phenotypic heterogeneity of VWFpp variants.</div></div>","PeriodicalId":17326,"journal":{"name":"Journal of Thrombosis and Haemostasis","volume":"24 2","pages":"Pages 483-497"},"PeriodicalIF":5.0,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145452109","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Protein S as a therapeutic target 蛋白S作为治疗靶点。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.10.024

Raja Prince Eladnani , Rim Diab , Anne Angelillo-Scherrer

Protein S (PS) is a key anticoagulant with additional roles in cell signaling through its interaction with Tyro3-Axl-MerTK family receptor tyrosine kinases Tyro3 and MerTK. Therapeutic targeting of PS—either enhancing or inhibiting its activity—has emerged as a promising strategy to modulate coagulation and inflammation. Approaches include nanobodies, monoclonal antibodies, and RNA interference, aiming to restore hemostatic balance or prevent bleeding, depending on the clinical context. Severe PS deficiency (homozygous or compound heterozygous) causes life-threatening conditions such as disseminated intravascular coagulation and purpura fulminans. Heterozygous deficiency is associated with an elevated venous thromboembolic risk. These insights highlight the need for tailored modulation of PS activity. A PS-enhancing nanobody has demonstrated potent antithrombotic effects in murine models, establishing proof-of-concept for a novel, safe anticoagulant strategy. Conversely, PS inhibition is being pursued to treat bleeding disorders such as von Willebrand disease and hemophilia. Both monoclonal antibodies and small-interfering RNAs targeting PS have shown safety and efficacy in preclinical models. Notably, a monoclonal antibody has demonstrated safety and preliminary efficacy in a phase 1/2 trial in patients with von Willebrand disease. Beyond coagulation, PS exerts protective effects on joint and bone tissues via Tyro3 and MerTK receptor signaling, further broadening its therapeutic potential. These findings support the continued development of PS-targeted therapies, with implications for both thrombotic and bleeding disorders, as well as for mitigating tissue damage in chronic bleeding conditions.

蛋白S （PS）是一种关键的抗凝剂，通过与TAM家族受体酪氨酸激酶Tyro3和MerTK相互作用，在细胞信号传导中发挥额外作用。靶向治疗ps -增强或抑制其活性-已成为一种有前途的策略来调节凝血和炎症。方法包括纳米体、单克隆抗体和RNA干扰，旨在恢复止血平衡或防止出血，具体取决于临床情况。严重的PS缺乏（纯合子或复合杂合子）会导致危及生命的疾病，如弥散性血管内凝血和暴发性紫癜。杂合子缺乏与静脉血栓栓塞风险升高有关。这些见解强调了对PS活动进行量身定制的调制的必要性。一种ps增强纳米体在小鼠模型中显示出强大的抗血栓作用，为一种新的、安全的抗凝策略建立了概念验证。相反，PS抑制正被用于治疗出血性疾病，如血管性血友病和血友病。针对PS的单克隆抗体和小干扰rna在临床前模型中均显示出安全性和有效性。值得注意的是，一种单克隆抗体已经在血管性血友病患者的1/2期试验中证明了安全性和初步有效性。除凝血外，PS还通过Tyro3和MerTK受体信号传导对关节和骨组织产生保护作用，进一步拓宽了其治疗潜力。这些发现支持ps靶向治疗的持续发展，对血栓性和出血性疾病以及减轻慢性出血条件下的组织损伤都有意义。

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引用次数: 0

Human leukocyte antigen alleles associated with inhibitor development in severe hemophilia A: analysis of the “My Life, Our Future” hemophilia A cohort 人类白细胞抗原等位基因与严重血友病A抑制剂发展相关：“我的生活，我们的未来”血友病A队列分析

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.08.040

Sara Contente , Anwar E. Ahmed , Barbara A. Konkle , Jill M. Johnsen , Andrew Frank , Kathleen P. Pratt

Background

An earlier analysis of the “My Life, Our Future” Repository indicated the risk of individuals with severe hemophilia A (HA) developing neutralizing antifactor (F)VIII antibodies (inhibitors) was (1) higher in Black and Hispanic cohorts and (2) similar in intron-22 inversion mutations vs other large structural changes in the F8 gene.

Objectives

This study determined whether specific human leukocyte antigen (HLA) alleles or haplotypes indicate either risk of or protection from inhibitor development in severe HA and if such associations correlate with race/ethnicity or F8 variant type.

Methods

HLA haplotypes were obtained from whole genome sequence data for 1841 subjects with severe HA and their racial/ethnic categories verified by principal component analysis. Pairwise and multivariable logistic regression analyses were performed for subcohorts of HLA alleles/haplotypes defined by HA mutation type and race/ethnicity.

Results

HLA-DRB1∗15:01, DQB1∗06:02, and DQA1∗01:02 and B∗07:02 were higher-risk alleles for the White/non–Hispanic cohort, while HLA-DRB∗14:01, DQB1∗05:03, DQB1∗03:01, B∗38:01, B∗44:02, and C∗05:01 were lower-risk alleles. HLA-B∗15:03 and B∗58:01 were higher-risk alleles for the Black/non–Hispanic cohort, while HLA-A∗33:03 was a lower-risk allele. Additional alleles were associated with inhibitor risk across the following cohorts: all (N = 1841), intron-22 inversion (n = 836), missense (n = 288), not-missense (n = 1553). No associations of HLA with inhibitor risk were seen for the Hispanic/White cohort. Analysis of subjects carrying both higher-risk and lower-risk alleles revealed combinations affecting the risks associated with individual alleles.

Conclusion

Our results implicate both innate and adaptive immune mechanisms, possibly including multiallelic HLA-associated effects on FVIII processing/presentation or signaling pathways in the development of inhibitors vs peripheral tolerance to FVIII.

早期对“我的生活，我们的未来”数据库的分析表明，严重血友病A （HA）患者产生中和性抗因子(F)VIII抗体（抑制剂）的风险(1)在黑人和西班牙裔人群中较高，(2)与F8基因的其他大结构变化相比，内含子22倒置突变的风险相似。目的：本研究确定特异性人类白细胞抗原（HLA）等位基因或单倍型是否表明严重HA患者抑制剂发展的风险或保护，以及这种关联是否与种族/民族或F8变异类型相关。方法从1841例重度HA患者的全基因组序列数据中获得shla单倍型，并采用主成分分析对其人种分类进行验证。对由HA突变类型和种族/民族定义的HLA等位基因/单倍型亚群进行两两和多变量logistic回归分析。结果hla - drb1∗15:01、DQB1∗06:02、DQA1∗01:02和B∗07:02为白人/非西班牙裔人群的高危等位基因，HLA-DRB∗14:01、DQB1∗05:03、DQB1∗03:01、B∗38:01、B∗44:02和C∗05:01为低风险等位基因。HLA-B∗15:03和B∗58:01是黑人/非西班牙裔队列的高风险等位基因，而HLA-A∗33:03是低风险等位基因。在以下队列中，其他等位基因与抑制剂风险相关：所有（N = 1841），内含子-22倒置（N = 836），错义（N = 288），非错义（N = 1553）。在西班牙裔/白人队列中，HLA与抑制剂风险没有关联。对携带高风险和低风险等位基因的受试者的分析揭示了影响个体等位基因相关风险的组合。我们的研究结果暗示了先天和适应性免疫机制，可能包括多等位hla相关的FVIII加工/呈递或信号通路在抑制剂与外周对FVIII耐受的发展中的作用。

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引用次数: 0

A comparative analysis of HAS-BLED, ORBIT, DOAC, and AF-BLEED bleeding-risk scores in anticoagulated patients with atrial fibrillation: a report from the prospective Murcia atrial fibrillation project III (MAFP-III) cohort 房颤抗凝患者的hs - bled、ORBIT、DOAC和AF- bleed出血风险评分的比较分析：来自前瞻性Murcia AF项目III （mfp -III）队列的报告。

IF 5 2区医学 Q1 HEMATOLOGY

Journal of Thrombosis and Haemostasis

Pub Date : 2026-02-01 DOI: 10.1016/j.jtha.2025.07.040

Eva Soler-Espejo , María Pilar Ramos-Bratos , José Miguel Rivera-Caravaca , Eduardo González-Lozano , Francisco Marín , Vanessa Roldán , Gregory Y.H. Lip

Background

Patients with atrial fibrillation (AF) often require oral anticoagulation (OAC), but predicting bleeding risk remains challenging. Classic risk scores such as HAS-BLED and ORBIT, along with newer models such as DOAC and AF-BLEED, exhibit only modest predictive performance.

Objectives

In this study, we compared these 4 scores in patients with AF on OAC to evaluate their predictive accuracy and clinical utility.

Methods

We included patients with AF starting OAC between January 2016 and November 2021. Endpoints assessed were major bleeding, major bleeding/clinically relevant nonmajor bleeding (CRNMB), and intracranial hemorrhage (ICH). Score performance was assessed through discrimination, calibration, reclassification (net reclassification improvement), decision curve analysis, and accuracy over a 2-year follow-up period.

Results

In total, 3259 patients with AF were included (median age, 77 years; IQR, 70-83 years; 52.8% female). Over 2 years’ follow-up, 196 patients (6.35%/year) experienced major bleeding, 413 (10.3%/year) major bleeding/CRNMB, and 32 (0.5%/year) experienced ICH. All risk scores performed modestly for major bleeding (c-indexes, <0.7), with ORBIT (c-index, 0.664) and HAS-BLED (c-index, 0.651) performing the best. All 4 scores demonstrated good discriminatory ability (log-rank P < .001). The ORBIT score showed a slight improvement in net reclassification improvement for major bleeding compared with HAS-BLED, with a modest increase in discrimination, while no other score outperformed HAS-BLED. Calibration showed that HAS-BLED and ORBIT outperformed the DOAC and AF-BLEED scores for major bleeding and CRNMB. The Brier index indicated high accuracy across all scores.

Conclusion

All bleeding-risk scores only had modest predictive ability. None demonstrated clear superiority in predicting major bleeding, major bleeding/CRNMB, or ICH.

房颤（AF）患者通常需要口服抗凝（OAC），但预测出血风险仍然具有挑战性。经典的风险评分，如HAS-BLED和ORBIT，以及较新的模型，如DOAC和AF-BLEED，仅表现出适度的预测性能。在此，我们比较了房颤患者OAC的这四种评分，以评估其预测准确性和临床实用性。方法：我们纳入了2016年1月至2021年11月间开始OAC治疗的房颤患者。评估的终点为大出血、大出血/临床相关非大出血（CRNMB）和颅内出血（ICH）。在两年的随访期间，通过区分、校准、重新分类（NRI）、决策曲线分析（DCA）和准确性来评估得分表现。结果：纳入3259例房颤患者（中位年龄77 [IQR 70-83]岁，女性占52.8%）。随访2年，大出血196例（6.35%/年），大出血/CRNMB 413例（10.3%/年），脑出血32例（0.5%/年）。所有风险评分对大出血的预测能力一般。结论：所有出血风险评分仅具有一般的预测能力。在预测大出血、大出血/CRNMB或脑出血方面没有明显的优势。

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引用次数: 0