The Glycoprotein VI-Phospholipase Cγ2 Signaling Pathway Controls Thrombus Formation Induced by Collagen and Tissue Factor In Vitro and In Vivo

Abstract

Objective-Both collagen and tissue factor can be initiating factors in thrombus formation. We investigated the signaling pathway of collagen-induced platelet activation in interaction with tissue factor-triggered coagulation during the thrombus-forming process. Methods and Results-In murine blood flowing over collagen, platelet exposure of phosphatidylserine and procoagulant activity, but not adhesion, completely relied on each of the following signaling modules: glycoprotein VI (GPVI), FcR ␥-chain, Src kinases, adaptor protein LAT, and phospholipase C␥2 (PLC␥2). On flow in the presence of tissue factor, these signaling components were essential for platelet aggregation and greatly enhanced fibrin clot formation. Collagen-stimulated thrombin generation relied on the presence and activity of GPVI, FcR ␥-chain, Src kinase, LAT, and PLC␥2. The physiological importance of this GPVI pathway was shown in a FeCl 3 -induced in vivo murine thrombosis model. In both venules and arterioles, signaling through GPVI, FcR ␥-chain, and Src kinases enhanced the formation of phosphatidylserine-exposing and fibrin-rich thrombi. Key Words: glycoprotein VI Ⅲ LAT Ⅲ platelets Ⅲ Src kinase Ⅲ thrombin T hrombus formation can be initiated by both platelet-and coagulation-activating factors. Collagens in the extracellular matrix and other vascular layers are thought to act as principal platelet-activating components of the damaged vessel wall; they also provide a surface for von Willebrand factor adhesion. 1 Tissue factor, also exposed in damaged vessels, is a key trigger of the coagulation process. 2 Because of the proposed major role of platelets in arterial thrombosis and the importance of coagulation in venous thrombosis, the current understanding is that collagen/von Willebrand factor-mediated events are more important in arteries, whereas tissue factor plays a more prominent role in venous thrombus formation. Conclusions-TheFlow studies with human and mouse blood have established that the signaling receptor glycoprotein VI (GPVI) exclusively mediates collagen-induced platelet procoagulant activity, thus linking the processes of platelet activation and coagulation. [3][4][5] This procoagulant platelet response is mediated by a prolonged and potent rise in cytosolic [Ca 2ϩ ] i , which results in exposure of procoagulant phosphatidylserine (PS) at the platelet outer surface. 6 PS exposure is a key regulating factor in the coagulation process. For instance, PS-containing membrane surfaces dramatically increase the formation of factor Xa and thrombin. 7 However, several authors have argued that this platelet response has only an assistant role in coagulation and that, in vivo, other platelet reactions may play important roles as well. 2,8 Thus, although there is no doubt that platelets enhance thrombin generation (coagulation) in plasma or whole blood, the precise mechanism is still a matter of debate. The platelet immunoreceptor GPVI is coexpressed with the Fc receptor (FcR) ␥-chain, although the latter also interacts with other plat...

PDK1 Determines Collagen-Dependent Platelet Ca 2+ Signaling and Is Critical to Development of Ischemic Stroke In Vivo

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