The mechanisms of osteoclast recruitment, differentiation, and action at the bone surface are poorly understood. Recent observations support the hypothesis that bone resorbing osteoclasts are of vascular origin. In addition many osteoclast responses to humoral and microenvironmental cues are mediated by cell surface components. Little is known about osteoclast-specific cell surface proteins or about their acquisition and turnover during osteoclast development and regulation. The proposed studies are intended to test the hypothesis that specific osteoclast cell surface phenotypic markers exist. Furthermore we plan to investigate the emergence of the osteoclast phenotype from their precursor cells. Finally, we shall begin to investigate the function of these cell surface components as related to osteoclast differentiation. Such results will help understand normal and inflammatory bone loss and aid in devising therapies for arthritic, diabetic, and periodontal disease associated osteopenia. We propose to accomplish this by utilizing an osteoclast isolation procedure that results in osteoclast populations enriched for between 75-90% and quantities ranging between 15 and 30 million cells per preparation. These cells may be cultured for up to two weeks maintaining osteoclast morphology and enzymatic profiles. We have also described a cell culture system in which circulating monocytes transform into multinucleated giant cells. Monocytes when cultured in the presence of osteoblasts develop morphologies indistinguishable from osteoclasts. We will utilize monoclonal antibody technology to generate osteoclast specific phenotypic markers. These markers in turn will be utilized in conjunction with the giant cell culture system and matrix implantation studies to trace osteoclast lineage. This will be accomplished by utilizing monoclonal antibodies to localize and quantitate (peroxidase localization, R.I.A., ELISA assays) the emergence of the osteoclast. The culture systems will be exploited to determine what factors effect the emergence of specific osteoclast markers. Specific osteoclast surface components will be analyzed for functional roles by monoclonal antibody blocking techniques and characterized bichemically. The goal of this proposal is to begin to gain insight into cell surface changes associated with osteoclast development and function that may regulate normal and pathological bone loss.
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