High Concentrations of Thrombospondin-1 Suppress Shear Stress Induced Cleavage of Von Willebrand Factor by ADAMTS13.

Abstract 3063

Poster Board II-1039

ADAMTS13 is a circulating metalloprotease that cleaves and down regulates the activity of von Willebrand factor (VWF) in the circulation. Deficiency of ADAMTS13 causes thrombotic thrombocytopenic purpura (TTP). VWF is critical for supporting platelet adhesion and aggregation at sites of microvascular injury. Nevertheless, the factors that protect VWF from cleavage by ADAMTS13 remain unknown. Thrombospondin-1 (Thbs-1), an adhesive protein that exhibit binding to multiple proteins such as collagen, fibronectin and fibrin, promotes thrombus adherence in animal models of vascular injury. To investigate the role of Thbs-1 in the regulation of VWF cleavage by ADAMTS13, we analyzed the cleavage of VWF flowing through a capillary tube. In the presence of physiological concentrations of ADAMTS13, VWF fragments were generated in a shear stress dependent manner. The cleavage of VWF in the capillary tube device was inhibited by Thbs-1 in a concentration dependent manner. At physiological concentrations (30 ng/mL) of Thbs-1, the cleavage of VWF was decreased insignificantly to 85% of control. The cleavage was decreased to 50% at 100 ng/mL Thbs-1, and to less than 10% at 1000 ng/mL Thbs-1 (P<0.0001). The suppressive effect of Thbs-1 was also observed with a truncated variant of ADAMTS13 that lacks the distal six thrombospondin type 1 repeats and the CUB domains, suggesting that the C-terminal part of ADAMTS13 is not necessary for the suppressive effect of Thbs-1. Furthermore, the presence of platelets did not negate the suppressive effect of Thbs-1 on VWF cleavage. Thbs-1 did not suppress the cleavage of FRET-VWF73 or guanidine hydrochloride treated VWF. It also did not affect the cleavage of VWF multimers induced by vortexing in the presence of ADAMTS13 and platelets. In summary, the suppressive effect of Thbs-1 on VWF cleavage is shear stress dependent. Thbs-1 is not expected to play a major role in regulating the size of circulating VWF multimers. Nevertheless, high concentrations of Thbs-1 released from platelets may protect sheared VWF from cleavage by ADAMTS13. This protective effect of Thbs-1 on VWF proteolysis may help stabilize the hemostatic plugs at sites of microvascular injury.