The procoagulant effect of thrombin on fibrin(ogen)-bound platelets

In a final stage of activation, platelets become procoagulant because of th e appearance of phosphatidylserine (PS) at the membrane outer surface. This PS exposure requires a rise in cytosolic [Ca2+](i), is accompanied by form ation of membrane blebs, and stimulates the formation of thrombin from its precursor prothrombin. Here, we investigated whether thrombin, as a potent platelet agonist, can induce this procoagulant response in plasma-free plat elets interacting with fibrin or fibrinogen through their integrin alpha(ll b)beta(3) receptors. First, in platelets that were stimulated to spread ove r fibrin or fibrinogen surfaces with adrenaline, addition of thrombin and C aCl2 caused a potent Ca2+ signal that in about 30% of the cells was accompa nied by exposure of PS. At low doses, integrin alpha(llb)beta(3) receptor a ntagonist (RGD peptide) inhibited platelet spreading as well as thrombin-ev oked PS exposure. Second, in platelet-fibrinogen microaggregates that were preformed in the presence of adrenaline, thrombin/CaCl2 induced PS exposure and bleb formation of about 35% of the cells. Third, a potent, thrombin-de pendent stimulation of prothrombinase activity was measured in platelet sus pensions that were incubated with a fibrin clot. These results indicate tha t, in the absence of coagulating plasma, thrombin is a moderate inducer of the procoagulant response of platelets, once integrin alpha(llb)beta(3)-med iated interactions are stimulated (by adrenaline) and CaCl2 is present.