The interaction of platelets with leukocytes (neutrophils and monocytes) is a well recognized phenomenon. However, the molecular mechanisms of this interaction are poorly understood. Substantial evidence has accumulated indicating that plateletleukocyte interactions play a role in the pathogenesis of thrombosis and atherosclerosis. Platelet modulation of leukocyte function has also been suggested to be a important regulatory component of the inflammatory response. Studies in this laboratory have shown that platelets form specific cell-cell contacts with leukocytes in vitro and these contacts occur concurrently with platelet-mediated enhancement of leukocyte function. The hypothesis of the present proposal is that one or more of four adhesive glycoproteins (fibrinogen, fibronectin, von Willebrand factor, and thrombospondin)mediate specific platelet-leukocyte cell contacts by binding to the GPIIbIIIa receptor complex on platelets and an adhesive glycoprotein receptor(s) on leukocytes. This hypothesis will be addressed using a human in vitro model system. In the proposed studies, the time course and distribution of platelet-neutrophil and platelet-monocyte cell contacts will be determined by transmission electron microscopy. The relative inhibitory effect of anti-adhesive glycoprotein Fab antibody fragments will be assessed. Gold- and heme-octapeptide-labeled Fab antibody fragments directed against the adhesive glycoproteins will then be used to identify the proteins at the cell contacts with the electron microscope. The effect and dose response of exogenously added adhesive glycoproteins on cell contact formation will be determined with the proteins added alone and in combination. In addition, exogenous fibrinogen, fibronectin, von Willebrand's factor and thrombospondin labeled with 5 nm colloidal gold and heme-octapeptide will be incorporated into the platelet-leukocyte contacts for direct visualization via transmission electron microscopy. To corroborate morphological findings, synthetic peptides of candidate binding domains will be assessed for their ability to inhibit platelet-leukocyte binding. The receptors involved in contact bridge formation will be identified by inhibition studies and electron microscopy using a panel of antibodies directed against platelet and leukocyte integrins such as GPIIbIIIa on the platelet and MAC-1 on the leukocyte.