α2β1 integrin, GPVI receptor, and common FcRγ chain on mouse platelets mediate distinct responses to collagen in models of thrombosis.

Robin J Marjoram,Mary M Zutter,Douglas M Tollefsen,S Kent Dickeson,Thomas J Kunicki,Zhengzhi Li,Li He,Samuel A Santoro,Ching-Ping Tseng

doi:10.1371/journal.pone.0114035

Abstract

ObjectivePlatelets express the α2β1 integrin and the glycoprotein VI (GPVI)/FcRγ complex, both collagen receptors. Understanding platelet-collagen receptor function has been enhanced through use of genetically modified mouse models. Previous studies of GPVI/FcRγ-mediated collagen-induced platelet activation were perfomed with mice in which the FcRγ subunit was genetically deleted (FcRγ−/−) or the complex was depleted. The development of α2β1−/− and GPVI−/− mice permits side-by-side comparison to address contributions of these collagen receptors in vivo and in vitro.Approach and ResultsTo understand the different roles played by the α2β1 integrin, the GPVI receptor or FcRγ subunit in collagen-stimulated hemostasis and thrombosis, we compared α2β1−/−, FcRγ−/−, and GPVI−/− mice in models of endothelial injury and intravascular thrombosis in vivo and their platelets in collagen-stimulated activation in vitro. We demonstrate that both the α2β1 integrin and the GPVI receptor, but not the FcRγ subunit influence carotid artery occlusion in vivo. In contrast, the GPVI receptor and the FcRγ chain, but not the α2β1 integrin, play similar roles in intravascular thrombosis in response to soluble Type I collagen. FcRγ−/− platelets showed less attenuation of tyrosine phosphorylation of several proteins including RhoGDI when compared to GPVI−/− and wild type platelets. The difference between FcRγ−/− and GPVI−/− platelet phosphotyrosine levels correlated with the in vivo thrombosis findings.ConclusionOur data demonstrate that genetic deletion of GPVI receptor, FcRγ chain, or the α2β1 integrin changes the thrombotic potentials of these platelets to collagen dependent on the stimulus mechanism. The data suggest that the FcRγ chain may provide a dominant negative effect through modulating signaling pathways in platelets involving several tyrosine phosphorylated proteins such as RhoGDI. In addition, these findings suggest a more complex signaling network downstream of the platelet collagen receptors than previously appreciated.

Highlights

Hemostasis relies on the highly regulated balance of prothrombotic and antithrombotic components to prevent blood loss from the vasculature while at the same time maintaining blood fluidity
The findings reported in this study demonstrate novel requirements in vivo and in vitro for the platelet collagen receptors, the a2b1 integrin and the glycoprotein VI (GPVI)/Fc receptor-gamma (FcRc) receptor complex
The loss of GPVI receptor or the FcRc chain decreases intravascular thrombosis in response to soluble Type I collagen, but surprisingly the FcRc2/2 animals retain some thrombotic potential to collagen

Summary

Introduction

Hemostasis relies on the highly regulated balance of prothrombotic and antithrombotic components to prevent blood loss from the vasculature while at the same time maintaining blood fluidity. Platelets play a central role in this balance especially during arterial hemostasis and pathological thrombosis. Fibrillar collagens represent a potent prothrombotic stimulus for platelets at sites of vascular injury. A2b1 integrin and the glycoprotein VI (GPVI)/ Fc receptor-gamma (FcRc) complex, that together mediate platelet adhesion and activation in response to collagens [1,2,3]. GPVI, a single span transmembrane receptor with two immunoglobulin domains non-covalently associates with the FcRc chain that contains the immunoreceptor tyrosine-based activation motif (ITAM), which in complex form the primary collagen signaling receptor [4,5,6]

Methods

Results

Conclusion