The Procoagulant Effect of Thrombin on Fibrin(ogen)-Bound Platelets

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In a final stage of activation, platelets become procoagulant because of the appearance of phosphatidylserine (PS) at the membrane outer surface. This PS exposure requires a rise in cytosolic [Ca²⁺]_i, is accompanied by formation 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 platelets interacting with fibrin or fibrinogen through their integrin α_IIbβ₃ receptors. First, in platelets that were stimulated to spread over fibrin or fibrinogen surfaces with adrenaline, addition of thrombin and CaCl₂ caused a potent Ca²⁺ signal that in about 30% of the cells was accompanied by exposure of PS. At low doses, integrin α_IIbβ₃ receptor antagonist (RGD peptide) inhibited platelet spreading as well as thrombin-evoked 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-dependent stimulation of prothrombinase activity was measured in platelet suspensions that were incubated with a fibrin clot. These results indicate that, in the absence of coagulating plasma, thrombin is a moderate inducer of the procoagulant response of platelets, once integrin α_IIbβ₃-mediated interactions are stimulated (by adrenaline) and CaCl₂ is present.