Activation of Protein C and Thrombin Activable Fibrinolysis Inhibitor on Cultured Human Endothelial Cells

Essentials It is unknown if thrombin activatable fibrinolysis inhibitor (TAFI) and protein C compete on cells. TAFI and protein C activation on endothelial cells was simultaneously quantified. TAFI and protein C do not compete for activation on endothelial cells. TAFI and protein C are independently recognized by the thrombin-thrombomodulin complex. Summary Background When bound to thrombomodulin (TM), thrombin is a potent activator of protein C (PC) and thrombin activable fibrinolysis inhibitor (TAFI). By binding PC and presenting it to the thrombin-TM complex, endothelial cell PC receptor (EPCR) enhances PC activation. It is unknown whether PC and TAFI compete for the thrombin-TM complex on endothelial cells. Objective To compare PC and TAFI activation on the surface of cultured human endothelial cells in the absence or presence of JRK1535 and/or CTM1009, inhibitory antibodies directed against EPCR and TM, respectively, and to determine whether PC and TAFI compete with each other for activation. Methods PC and TAFI activation on endothelial cells were compared, and the effect of PC on TAFI activation and TAFI on PC activation was determined in the absence or presence of JRK1535 and/or CTM1009. Results In the absence of antibodies, activation of PC was four-fold faster than that of TAFI. Blocking EPCR with JRK1535 resulted in a 53-fold decrease in PC activation and no effect on TAFI activation. Blocking TM with CTM1009 inhibited both TAFI and PC activation. Neither TAFI nor PC competed with each other in the absence or presence of JRK1535. Conclusions PC and TAFI are concurrently activated in a TM-dependent manner and do not compete for the thrombin-TM complex, raising the possibility that they interact with distinct activation complexes. EPCR selectively enhances PC activation so that PC and TAFI activation kinetics become comparable on endothelial cells.