Certain components of the extracellular (ECM) matrix have a major effect on phagocytic cell function. Laminin (Lam), fibronectin (Fn), and serum anyloid P component all enhance the ability of blood derived phagocytes to ingest particles opsonized with either IgG or complement. This phenomenon may play an important role in phagoctye activation at extravascular sites of infection and inflammation. The molecular mechanisms by which these proteins enhance opsonin mediated phagocytosis are unknown. Moreover, very little is known about the interaction of the ECM proteins with phagocytes. We propose to investigate the biology of phagocytosis enhancement in great detail. We will investigate the function of IgG-Fc receptors, C3b receptors (CR1), and iC3b receptors (CR3) before and after phagocyte stimulation with Fn and Lam, using defined multivalent ligands as probes for the function of each receptor. The effect of Lam and Fn on three important aspects of receptor function will be investigated: 1) the extent of binding and the affinity of the probes for phagocytes will be determined before and after Fn or Lam stimulation; 2) the rate and extent of internalization of the probes by the phagocytes will be determined and evaluated as a function of Fn or Lam stimulation; 3) the effects of Fn and Lam on the process of coendocytosis (the internalization of monomeric ligand in the presence of multimeric ligand) will be assessed. This project will also characterize the interactions of phagocytes with defined domains of Fn and Lam. Using this information we will isolate and raise antibodies to the cellular receptors for Fn and Lam. We will investigate the roles of phospholipid hydrolysis and protein phosphorylation in transduction of the signal for phagocytosis enhancement provided by Fn and Lam. Finally, we will use electron microscopy to confirm and extend the findings of these biochemical and physiologic studies. It is expected that the data obtained from these studies will suggest a molecular mechanism for the enhancement of binding and phagocytosis of opsonized particles by phagocytes exposed to ECM proteins. These data will enhance our understanding the cell biological mechanisms of phagocyte activation after emigration to extravascular sites of inflammation, infection or injury. This will be relevant to the physiology and pathophysiology of wound healing and many connective tissue, immune complex mediated, and infectious diseases.
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