Grant: Operating Grant

von Willebrand factor (VWF) is a large pro-coagulant adhesive protein that is required for two key functions in blood clotting: the initial attachment of blood platelets to sites of vessel damage, and the carriage of clotting factor VIII in plasma. Inherited quantitative or qualitative abnormalities of VWF result in the mucocutaneous bleeding disease, von Willebrand disease (VWD). The aim of the studies proposed in this research project is to investigate the molecular basis of these variant forms of VWF. The first group of studies will characterize quantitative abnormalities of VWF relating to changes in the genetic sequence regulating expression of the VWF gene. Sequence variation in this upstream region of the VWF gene has already been identified, and we will now evaluate the molecular mechanisms responsible for this reduction in VWF expression. In aims 2 and 3 of the project, we will address a number of questions relating to qualitative variants of VWF. These studies will focus on two principal areas of interest: cleavage of VWF by the processing enzyme ADAMTS13, and the interaction of VWF with the platelet receptor GpIb. In each instance, we will assess variant forms of VWF known, or hypothesized, to influence these activities. After the initial in vitro confirmation of functional changes, studies will be performed using two in vivo strategies in mice with severe VWF deficiency. Comprehensive testing of the hemostatic potential of the variant forms of VWF will be performed following initial recombinant protein infusion studies. These studies will be followed by more long-term protein replacement approaches involving variant VWF transgene delivery by hydrodynamic delivery to assess the contribution of plasma VWF function. The in vivo functional characterization of these variants will include a detailed analysis of the VWF/FVIII complex, the effects of ADAMTS13-mediated VWF cleavage, and involvement in primary hemostasis and arteriolar thrombogenesis.