Inhibition of thrombin-mediated factor V activation contributes to the anticoagulant activity of fibrinogen γ′

Background: Besides its role in blood clotting, fibrinogen exerts a poorly understood anticoagulant function by binding thrombin and modulating its activity. In particular, the γA/γ′ fibrinogen isoform binds with high affinity to thrombin exosite II through the anionic carboxyl-terminal end of the γ′ chain. This interaction down-regulates thrombin-mediated factor VIII (FVIII) activation, but its effect on FV activation is unknown. Objectives: To investigate the overall anticoagulant activity of fibrinogen and particularly of fibrinogen γ′ in plasma, and to verify whether the fibrinogen γ′ carboxyl-terminal peptide affects thrombin-mediated FV activation. Methods: Thrombin generation was measured by calibrated automated thrombography in whole and defibrinated plasma and in plasma supplemented with the (sulfated) fibrinogen γ′ carboxyl-terminal peptide (0-500 μmol L-1). The effect of the peptide on thrombin-mediated FV activation was studied in model systems and in plasma. Results: Total fibrinogen prolonged the lag time of thrombin generation at low tissue factor (TF) concentrations. The fibrinogen γ′ peptide dose-dependently prolonged the lag time and decreased the peak height of thrombin generation at low TF, whereas a scrambled control peptide was ineffective. These effects persisted in the presence of an anti-FVIII antibody, suggesting that the peptide may also inhibit thrombin-mediated activation of FV. This was confirmed in model systems and in plasma. Conclusions: Total fibrinogen and the fibrinogen γ′ peptide have an overall anticoagulant effect on thrombin generation determined at low TF. Inhibition of thrombin-mediated FV activation by the fibrinogen γ′ peptide is a novel mechanism of the anticoagulant activity of fibrinogen γ′.

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