Complementary roles for fibrin(ogen), thrombospondin and vWF in mediating shear-dependent aggregation of platelets stimulated at threshold thrombin concentrations

We have evaluated the relative contribution of the adhesive ligands, von Wi llebrand factor (vWF), fibrinogen (Fg) arid thrombospondin (TSP), all surfa ce-expressed on washed platelets (WP) activated with a threshold thrombin c oncentration (similar to 0.04 U/ml), to platelet microaggregation (PA) at s hear rates (G) from 300-2000 s(-1). In suspensions of thrombin-activated WP sheared immediately (tau (0)), all three ligands were required for optimal aggregation at all G, as shown by a 50-70% inhibition of capture efficienc ies of PA (measured from initial rates of PA), by antibodies (Abs) directed against each protein. This aggregation involved both GPIb and GPIIbIIIa, a s indicated by similar to 80% and 100% inhibition by Ab 6D1 and Ab 10E5, re spectively. For WP preexposed to thrombin for 10 min to ensure maximal surf ace expression of secreted ligands and activated GPIIbIIIa (tau (10)), vWT was predominantly required at all G (63-75% inhibition by anti-vWF Ab), tog ether with TSP (35-50% inhibition by anti-TSP Ab). Under these conditions, Fg was extensively converted to fibrin, so that fibrin, rather than Fg, cou ld participate in microaggregation, with GPIb less required than GPIIbIIIa as indicated by a 30-60% inhibition by Ab 6D1 as compared to 100% inhibitio n by Ab 10E5. Our results show that interactions between multiple ligands a nd receptors favour micro aggregation depending on shear and thrombin activ ation conditions.