A genetically-engineered von Willebrand disease type 2B mouse model displays defects in hemostasis and inflammation

Abstract

von Willebrand disease (VWD)-type 2B is characterized by gain-of-function mutations in the von Willebrand factor (VWF) A1-domain, leading to increased affinity for its platelet-receptor, glycoprotein Ibα. We engineered the first knock-in (KI) murine model for VWD-type 2B by introducing the p.V1316M mutation in murine VWF. Homozygous KI-mice replicated human VWD-type 2B with macrothrombocytopenia (platelet counts reduced by 55%, platelet volume increased by 44%), circulating platelet-aggregates and a severe bleeding tendency. Also, vessel occlusion was deficient in the FeCl3-induced thrombosis model. Platelet aggregation induced by thrombin or collagen was defective for KI-mice at all doses. KI-mice manifested a loss of high molecular weight multimers and increased multimer degradation. In a model of VWF-string formation, the number of platelets/string and string-lifetime were surprisingly enhanced in KI-mice, suggesting that proteolysis of VWF/p.V1316M is differentially regulated in the circulation versus the endothelial surface. Furthermore, we observed increased leukocyte recruitment during an inflammatory response induced by the reverse passive Arthus reaction. This points to an active role of VWF/p.V1316M in the exfiltration of leukocytes under inflammatory conditions. In conclusion, our genetically-engineered VWD-type 2B mice represent an original model to study the consequences of spontaneous VWF-platelet interactions and the physiopathology of this human disease.

LIM kinase/cofilin dysregulation promotes macrothrombocytopenia in severe von Willebrand disease-type 2B

“…The presence of giant platelets in type 2B mutant vWF/p.V1316M (hereafter referred to simply as 2B) mice has been previously associated with reduced platelet counts (17). Platelet parameters of 2B mice are depicted in Table 1: the mean platelet volume (MPV) was increased by 44% compared with WT.…”

“…The VWD-type 2B knockin mouse model (p.V1316M) has been described elsewhere (17). Mice homozygous for the p.V1316M mutation are referred to as 2B mice and their control littermates as WT mice.…”

“…However, we also demonstrated that the hemorrhagic phenotype of these mice was not corrected, suggesting that elevation of platelet count was not sufficient to restore a normal hemostasis. Furthermore, we previously demonstrated that platelets from 2B patients (29) and 2B mice (17) are defective in integrin αIIbβ3 activation. After LIMK inhibition, the activation of integrin αIIbβ3 of 2B platelets was not restored.…”

“…Interestingly, in a model of platelet formation upregulated by exposure of cultured MKs on a vWF surface to high shear forces, we have previously established a defect in platelet release from normal MKs exposed to immobilized p.V1316M vWF (16). We have recently engineered a knockin murine model for severe VWD-type 2B by introducing the p.V1316M mutation in the A1 domain of the Vwf gene, and we have demonstrated that these mice display a bleeding phenotype that we attributed to both macrothrombocytopenia and thrombopathy (17). Given the crucial role of the cytoskeletal changes to allow megakaryocytopoiesis, we used this mouse model of VWD-type 2B to investigate the hypothesis that a disturbed regulation of actin remodeling contributes to impaired proplatelet formation in VWD-type 2B.…”

Targeting von Willebrand Factor–Mediated Inflammation

“…[4][5][6][7][8] Moreover, although the underlying biology remains unexplained, the animal data further suggest that the relative importance of these discrete VWF-mediated pathways varies between different types of inflammation. Further insights into the role played by VWF in regulating leukocyte extravasation and endothelial cell permeability and the complex cross-talk that exists in vivo between hemostasis and inflammation will undoubtedly emerge in the near future.…”