Mural thrombus generation in type 2A and 2B von Willebrand disease under flow conditions

Sugimoto, Mitsuhiko; Matsui, Hideto; Mizuno, Tomohiro; Tsuji, Shizuko; Miyata, Shigeki; Matsumoto, Masanori; Matsuda, Michio; Fujimura, Yoshihiro; Yoshioka, Akira

doi:10.1182/blood-2002-03-0944

Cited by 46 publications

(46 citation statements)

References 25 publications

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“…This enhanced thrombogenesis most likely reflects a block in ADAMTS13 activity rather than nonspecific effects of antibody on platelets, since immunostaining of thrombi with N10 antibody, which reacts only with VWF cleaved by ADAMTS13, visually confirmed the reduced VWF cleavage within thrombi by the anti-ADAMTS13 antibody (Figure 2A). Thus, these results clearly point to the regulatory role of ADAMTS13 during thrombus generation.While the preceding observations were made under a much higher shear rate than the high shear rate typically used in platelet functional studies (ie, 1500 s Ϫ1 in our laboratory [14][15][16] ), similar observations, although less pronounced, were confirmed under lower shear rates ( Figure 2D). In addition, longitudinal views of thrombi revealed the preferential VWF-cleavage activity of ADAMTS13 at the surface portions of forming thrombi during thrombogenesis ( Figure 2B,C).…”

supporting

confidence: 77%

Functional imaging of shear-dependent activity of ADAMTS13 in regulating mural thrombus growth under whole blood flow conditions

Shida¹,

Nishio

Sugimoto³

et al. 2008

Blood

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The metalloprotease ADAMTS13 is assumed to regulate the functional levels of von Willebrand factor (VWF) appropriate for normal hemostasis in vivo by reducing VWF multimer size, which directly represents the thrombogenic activity of this factor. Using an in vitro perfusion chamber system, we studied the mechanisms of ADAMTS13 action during platelet thrombus formation on a collagen surface under whole blood flow conditions. Inhibition studies with a functionblocking anti-ADAMTS13 antibody, combined with immunostaining of thrombi with an anti-VWF monoclonal antibody that specifically reflects the VWFcleaving activity of ADAMTS13, provided visual evidence for a shear ratedependent action of ADAMTS13 that limits thrombus growth directly at the site of the ongoing thrombus generation process. Our results identify an exquisitely specific regulatory mechanism that prevents arterial occlusion under high shear rate conditions during mural thrombogenesis. IntroductionThe adhesive protein von Willebrand factor (VWF) plays a major role in platelet thrombogenesis, a process crucial for hemostasis. However, the excessive function of VWF is thought to increase the risk of fatal arterial thrombosis. 1,2 The thrombogenic activity of VWF is strictly dependent upon its multimeric structure, which is thought to be regulated in vivo by the metalloprotease ADAMTS13 through its cleavage of the A2 domain of the VWF subunit. 3,4 Indeed, patients with congenital deficiency of ADAMTS13 suffer repeated thrombotic complications attributed to excessive function of the ultra-large VWF (ULVWF) multimer, which is not found in normal blood circulation. [3][4][5][6] This concept was recently confirmed by knock-out mouse studies, in which ADAMTS13 Ϫ/Ϫ mice exhibited enhanced thrombogenicity in the ex vivo or in vitro experimental blood flow conditions tested. 7,8 The mechanisms by which ADAMTS13 regulates VWF remain poorly understood. However, recent studies showing that ADAMTS13 under flow conditions can rapidly cleave ULVWF secreted from and anchored to cultured endothelial cell layers 9,10 have raised the possibility that blood flow is critical in activating ADAMTS13. 11 Indeed, the VWF-cleaving activity of ADAMTS13 cannot be reproduced in vitro under static conditions unless the substrate VWF molecule is somewhat modified (eg, denatured by guanidine-HCl or urea). 3,4 Further, the question arises of whether ADAMTS13, in addition to its known action on ULVWF freshly released from endothelial cells, might also act directly at the local sites of thrombus generation to regulate thrombus growth.To address these issues, we analyzed the role and mechanisms of ADAMTS13 action in mural platelet thrombogenesis on a collagen-coated glass surface in an in vitro perfusion chamber system. Our visual evidence demonstrates that ADAMTS13 cleaves VWF and down-regulates mural thrombus growth at the site of ongoing thrombus generation in a shear rate-dependent manner under whole blood flow conditions. Methods Blood collectionThe present work was approved by the i...

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supporting

confidence: 77%

Functional imaging of shear-dependent activity of ADAMTS13 in regulating mural thrombus growth under whole blood flow conditions

Shida¹,

Nishio

Sugimoto³

et al. 2008

Blood

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“…A single study published 25 years ago showed that reduced platelet aggregation and secretion were correlated to impaired granule contents in response to thrombin, collagen, and ADP in 2 patients with vWDtype 2B (9). Along the same line, an in vitro study using perfusion assays on a collagen matrix reported a heterogeneous defect in thrombus growth in vWD-type 2B (10).…”

Section: Introductionmentioning

confidence: 68%

von Willebrand factor mutation promotes thrombocytopathy by inhibiting integrin αIIbβ3

Casari¹,

Berrou²,

Lebret³

et al. 2013

J. Clin. Invest.

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von Willebrand disease type 2B (vWD-type 2B) is characterized by gain-of-function mutations in von Willebrand factor (vWF) that enhance its binding to the glycoprotein Ib-IX-V complex on platelets. Patients with vWD-type 2B have a bleeding tendency that is linked to loss of vWF multimers and/or thrombocytopenia. In this study, we uncovered evidence that platelet dysfunction is a third possible mechanism for bleeding tendency. We found that platelet aggregation, secretion, and spreading were diminished due to inhibition of integrin αIIbβ3 in platelets from mice expressing a vWD-type 2B-associated vWF (vWF/p.V1316M), platelets from a patient with the same mutation, and control platelets pretreated with recombinant vWF/p.V1316M. Impaired platelet function coincided with reduced thrombus growth. Further, αIIbβ3 activation and activation of the small GTPase Rap1 were impaired by vWF/p.V1316M following exposure to platelet agonists (thrombin, ADP, or convulxin). Conversely, thrombin-or ADP-induced Ca 2+ store release, which is required for αIIbβ3 activation, was normal, indicating that vWF/p.V1316M acts downstream of Ca 2+ release and upstream of Rap1. We found normal Syk phosphorylation and PLCγ2 activation following collagen receptor signaling, further implying that vWF/p.V1316M acts directly on or downstream of Ca 2+ release. These data indicate that the vWD-type 2B mutation p.V1316M is associated with severe thrombocytopathy, which likely contributes to the bleeding tendency in vWD-type 2B.

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“…7 This results in acquired type 2 vWS, which is characterized by the loss of the largest von Willebrand multimers, which are most effective in platelet-mediated hemostasis. 8,9 The incidence of bleeding events in patients with aortic stenosis within 6 months before valve replacement was 21%. 10 It has also been shown that the severity of the stenosis, indicated by higher mean transvalvular gradients as well as higher wall shear stress, was negatively correlated with the percentages of the highest-molecular-weight multimers.…”

Section: Editorial See P 647 Clinical Perspective On P 681mentioning

confidence: 99%

Acquired von Willebrand Syndrome in Patients With an Axial Flow Left Ventricular Assist Device

Meyer

Malehsa²,

Bara³

et al. 2010

Circ: Heart Failure

257

145

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Background-Rotary blood pumps used as left ventricular assist devices (LVADs) allow for long-term support and may become suitable alternatives to heart transplantation. Effects of this technology on the coagulation system are not completely understood, leading to controversial anticoagulation protocols. Thus, we investigated the primary hemostasis in patients with chronic LVAD therapy. Methods and Results-Twenty-six outpatients received axial flow LVAD (HeartMate II; Thoratec) for a median support time of 4.5 months. In a cross-sectional protocol, platelet aggregation in response to ADP and epinephrine, von Willebrand antigen (vWF:AG), and collagen-binding capacity (vWF:CB) were obtained. Von Willebrand factor (vWF) multimer analyses were performed, and patients were screened for bleeding events. This analysis was repeated after removal of the device for transplantation or recovery (nϭ12) and after a median of 15.5 months in ongoing patients (nϭ11). In all patients on devices, severe impairment of platelet aggregation as well as a loss of large vWF multimers were found. In 10 patients, a decreased vWF:CB/vWF:AG ratio was observed. Bleeding episodes occurred with an incidence of 0.17 per patient-year. After removal of the device, normal patterns of platelet aggregation, multimer analysis, and vWF:CB/vWF:AG ratio were recorded. In the second analysis of ongoing patients, impairment of platelet aggregation and loss of large vWF multimers were verified. Conclusions-A diagnosis of von Willebrand syndrome type 2 was established in all patients after LVAD implantation, and bleeding events confirmed this finding. Reversibility of this condition was found after removal of the device. (Circ Heart Fail. 2010;3:675-681.)

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Mural thrombus generation in type 2A and 2B von Willebrand disease under flow conditions

Cited by 46 publications

References 25 publications

Functional imaging of shear-dependent activity of ADAMTS13 in regulating mural thrombus growth under whole blood flow conditions

Functional imaging of shear-dependent activity of ADAMTS13 in regulating mural thrombus growth under whole blood flow conditions

von Willebrand factor mutation promotes thrombocytopathy by inhibiting integrin αIIbβ3

Acquired von Willebrand Syndrome in Patients With an Axial Flow Left Ventricular Assist Device

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