Function of glycoprotein VI and integrin alpha(2)beta(1) in the procoagulant response of single, collagen-adherent platelets

Various collagen-based materials were used to assess the structural require ments of collagen for inducing the procoagulant response of adhering platel ets, as well as the collagen receptors involved. Crosslinked or monomeric c ollagen-related peptide (CRP), Gly-Cys-Hyp-(Gly-Pro-Hyp)(10)-Gly-Cys-Hyp-Gl y was highly adhesive for platelets in a glycoprotein VI- (GpVI-)dependent manner. Adhesion was followed by a prolonged increase in cytosolic [Ca2+](i ) formation of membrane blebs, exposure of phosphatidylserine (PS) and gene ration of prothrombinase-stimulating activity. Fibrils of type-I collagen w ere less adhesive but, once adhered, many of the platelets presented a full procoagulant response. Monomeric type-I collagen was unable to support adh esion, unless Mg2+-dependent integrin alpha(2)beta(1) interactions were fac ilitated by omission of Ca2+ ions. With all surfaces, however, post-additio n of CaCl2 to adhering platelets resulted ina potent Ca2+-influx signal, fo llowed by PS exposure and bleb formation. The procoagulant response elicite d by binding to CRP was inhibited by anti-GpVI Fab fragments, but not by im peding integrin alpha(2)beta(1)-mediated events. With fibrillar collagen, i t was inhibited by blocking either the GpVI- or integrin alpha(2)beta(1)-me diated interactions. This suggests that the triple-helical Gly-Pro-Hyp repe at in CRP and analogous sequences in fibrillar collagen stimulate the proco agulant response of adherent platelets by acting as ligands for GpVI. Influ x of Ca2+ is required for this response, and adhesion via integrin alpha(2) beta(1) serves to potentiate the signaling effects of GpVI.