The aims of this proposal cover 3 major objectives: 1. To characterize the nature of FVIII-mediated platelet adhesion to vascular subendothelium. We shall compare the binding of FVIII and desialylated FVIII (ASVIII) to deendothelialized rabbit and human vessels, cultured endothelial cells, endothelial cell extracellular matrix, collagen and microfibrils, and the effect of FVIII and ASVIII in modulating platelet adhesion in these systems. We shall also compare ASVIII binding to unstimulated platelets with FVIII binding to stimulated platelets from normals, aspirinized normals, and storage pool disease, Bernard-Soulier syndrome and thrombasthenic patients. We shall study the inhibition of FVIII and ASVIII binding by our monoclonal antibodies to platelet GpIb and the GpIIb-IIIa complex. 2. To determine the mechanism(s) of action of lupus anticoagulants, and the hemostatic significance of circulating antibodies reactive with anionic phospholipids. We shall continue studies of the anionic phospholipid specificity of lupus anticoagulants and compare them with anti-DNA antibodies from patients with lupus. We shall study inhibition of PGI2 synthesis in cultured endothelial cells produced by lupus anticoagulants purified by isoelectric focusing or affinity chromatography on PS,PA-Sepharose. We shall investigate the mechanism of hypoprothrombinemia, seen in 27% of patients with lupus anticoagulants by doing 125I-prothrombin turnover studies in lupus patients with and without anticoagulants. 3. To investigate the relationship between platelet and endothelial cell membrane proteins. We shall characterize the human endothelial cell membrane component reactive with monoclonal and polyclonal antibodies to platelet GpIIIa, determine whether this component has P1A1 antigenic specificity (like platelet GpIIIa), and study whether the binding of these antibodies alters endothelial cell function, particularly PGI2 production. We shall study IgG fractions from sera of patients with thrombotic thrombocytopenia purpura, who may have antibodies directed toward endothelial cells and/or platelets, for reactivity against endothelial cells, and against the GpIIIa component, in particular. We shall search for other membrane components in common between endothelial cells and platelets.
