LAUSR

Deep vein thrombosis results from the cooperative action of leukocytes, platelets, and endothelial cells. The proline-rich tyrosine kinase Pyk2 regulates platelet activation and supports arterial thrombosis. In this study, we combined pharmacological and genetic approaches to unravel the role of Pyk2 in venous thrombosis. We found that mice lacking Pyk2 almost completely failed to develop deep venous thrombi upon partial ligation of the inferior vena cava. Pyk2-deficient platelets displayed impaired exposure of phosphatidylserine and tissue factor expression by endothelial cells and monocytes was completely prevented by inhibition of Pyk2. In HUVECs, inhibition of Pyk2 hampered IL-11-induced expression of VCAM and P-selectin, and VWF release. Pyk2-deficient platelets showed defective adhesion on VWF and reduced ability to bind activated HUVECs under flow. Moreover, inhibition of Pyk2 in HUVECs strongly reduced platelet adhesion. Similarly, Pyk2-deficient neutrophils were unable to efficiently roll and adhere to immobilized endothelial cells under venous flow conditions. Moreover, platelets and neutrophils from Pyk2-KO mice showed defective ability to form heterogeneous aggregates upon stimulation, while plateletmonocyte interaction occurred normally. Consequently, platelet-neutrophil aggregates, abundant in blood of WT mice upon inferior vena cava ligation, were virtually undetectable in Pyk2-KO mice. Finally, we found that expression of Pyk2 was required for NETosis induced by activated platelets. Altogether our results demonstrate a critical role of Pyk2 in the regulation of the coordinated thromboinflammatory responses of endothelial cells, leukocytes and platelets leading to venous thrombosis. Pyk2 may represent a novel promising target in the treatment of deep vein thrombosis.