Identification of an endothelial cell cofactor for thrombin-catalyzed activation of protein C.

TLDR

Perfusion of the myocardium with protein C and thrombin generates activated protein C, demonstrating a potent anticoagulant activity. The study aims to determine whether the vascular endothelial surface supplies a cofactor that boosts thrombin‑mediated protein C activation. The endothelial surface acts as a cofactor, markedly accelerating protein C activation by thrombin. In myocardium and cultured endothelial monolayers, thrombin‑catalyzed protein C activation exceeds that in solution by >20,000‑fold, persists for at least an hour with thrombin present, decays rapidly without thrombin, and is further inhibited by diisopropyl‑phospho‑thrombin, which also reduces activation when co‑perfused.

Abstract

Perfusion of the myocardium with protein C in the presence of thrombin (EC 3.4.21.5) elicits a potent anticoagulant activity, which is identified as activated protein C on the basis of synthetic substrate hydrolysis and anticoagulant properties. The rate of activated protein C formation during the transit through the myocardium is at least 20,000 times that of thrombin-catalyzed activation of protein C in the perfusion solution. The capacity of the heart to activate protein C is maintained for at least 1 hr when thrombin is present in the perfusate, but decays (half-life approximately 30 min) once thrombin is omitted. Addition of diisopropyl-phospho-thrombin increases this decay rate more than 10-fold. Coperfusing diisopropylphospho-thrombin with active thrombin lowers the amount of protein C activation in the myocardium. Cultured monolayers of human endothelium enhance the rate of thrombin-catalyzed protein C activation. As with myocardium, the activation rate is inhibited by including diisopropylphospho-thrombin in the medium. It is proposed that the surface of vascular endothelium provides a cofactor that enhances the rate of protein C activation by thrombin.