γA/γ’ fibrinogen inhibits thrombin-induced platelet aggregation

Abstract

The minor gammaA/gamma' fibrinogen isoform contains a high affinity binding site for thrombin exosite II that is lacking in the major gammaA/gammaA fibrinogen isoform. We therefore investigated the biological consequences of the gamma' chain binding to thrombin. Thrombin-induced platelet aggregation was inhibited by gammaA/gamma' fibrinogen. Carboxyl terminal peptide fragment gamma'410-427 from the gamma' chain was also inhibitory, with an IC(50) of approximately 200 microM in whole plasma. Deletion of the peptide from either the amino or carboxyl end significantly decreased inhibition. In contrast to thrombin-induced platelet aggregation, aggregation induced by epinephrine, ADP, arachidonic acid, or SFLLRN peptide showed little inhibition by the gamma' peptide. The inhibition of thrombin-induced platelet aggregation was not due to direct inhibition of the thrombin active site, since cleavage of a small peptidyl substrate was 91% of normal even in the presence of 1 mM gamma'410-427. The gamma'410-427 peptide blocked platelet adhesion to immobilized thrombin under both static and flow conditions, blocked soluble thrombin binding to platelet GPIbalpha, and inhibited PAR1 cleavage by thrombin. These results suggest that the gamma' chain of fibrinogen inhibits thrombin-induced platelet aggregation by binding to thrombin exosite II. Thrombin that is bound to the gamma' chain is thereby prevented from activating platelets, while retaining its amidolytic activity.

The pleiotropic role of the fibrinogen γ′ chain in hemostasis

“…Lovely et al showed similar results in a study using synthetic ␥Ј peptides. 48 These studies indicate that variations in the ratio of ␥A and ␥Ј fibrinogen could significantly modify platelet activation in vivo. The effect of the ␥Ј chain on platelet function should be considered largely antithrombotic.…”

γ′ Fibrinogen as a novel marker of thrombotic disease

“…For example, thrombin binding to the γ ′ chain inhibits cleavage of factor VIII (36) , which slows the rate of coagulation. In addition, thrombin binding to the γ ′ chain inhibits thrombin activation of platelets through PAR-1 (37,38) , which would inhibit hemostasis. The effect of the γ ′ chain on thrombin cleavage of fi brinopeptides A and B has been controversial, with three different results reported from three different laboratories (18,25,26) .…”

“…This antithrombin I activity inhibits thrombin activation of coagulation under static conditions by inhibiting factor VIII cleavage (36) . In addition, this activity inhibits thrombin activation of platelets by inhibiting PAR1 cleavage (37,38) . Studies with transgenic mice containing the human fi brinogen thrombin-binding γ ′ chain sequence showed a decrease in thrombosis (53) .…”

The fibrinogen γA/γ′ isoform does not promote acute arterial thrombosis in mice

“…Additionally, the γ’ chain supports high affinity binding to thrombin exosite II [8, 9], and studies have shown that thrombin binding to the γ’ chain competitively inhibits thrombin-mediated platelet activation [10] and reduces thrombin-mediated FpB cleavage [7], and factor VIII [11] and V [12] activation. These properties suggest γA/γ’ fibrinogen has anticoagulant activity in vitro .…”