The long term goals of this proposal are: 1) to elucidate the mechanisms and the regulation of the fusion on mononuclear phagocytes (MNP) leading to the formation of both osteoclasts (OC) and giant cells (GC) and 2) to characterize the functional consequences of multinucleation in both normal and pathological bone resorption and in the host defense mechanisms involved in inflammatory reactions and tumors. Two approaches will be used: The first approach focuses upon the cellular and molecular basis for the particular tendency of cells of the MNP lineage to form polykaryons. This approach will more specifically involve kinetic studies of plasma membrane proteins of MNPs induced to form multinucleated cells in vitro, looking for a potential fusogenic protein(s). Combining the specific iodination of plasma membrane proteins, the metabolic labeling of membrane proteins and their subsequent immunoprecipitation with anti-mouse macrophage antibodies will compare the membrane protein composition of MNPs and GCs, i.e., before and after fusion. The second approach concentrates upon the role of the local microenvironment in the induction and regulation of the formation of multinucleated OC and GC from precursors of the MNP lineage. This approach will more specifically involve studies on the role of soluble factor(s) resulting from lymphocyte-macrophage interactions (Macrophage Fusion Factor). Attempts will be made to purify this putative factor and to analyze which cell(s) and cell interaction(s) lead to its secretion in vitro using monoclonal antibodies directed against specific lymphocyte subsets and macrophages. The effects of local factors such as acidification, proteases and calcium concentration on fusion of MNPs will also be studied in vitro. These two approaches closely interact in that the effects of local factors on the plasma membrane protein composition will also be studied. Parallel studies will be made on in vivo induced OCs and GCs, studing the kinetics of the formation of these cells as well as the molecular composition of their membranes using immunoelectron microscopic techniques and known surface antigen markers. In the long run, these studies may be of critical importance in the understanding of the functional implications of fusion in bone resorption and host defense mechanisms.
