Characterization of immortalized osteoclast precursors developed from micetransgenic for both bcl-X-L and simian virus 40 large T antigen

We recently developed an immortalized osteoclast (OCL) precursor cell line that forms large numbers of OCLs. This cell line was derived from mice doub ly transgenic for bcl-X-L, and large T antigen that was targeted to cells i n the OCL lineage (bcl-X-L/Tag cells). We have now characterized these cell s in terms of their surface and enzymatic phenotype, responsiveness to oste otropic factors, and differentiation potential. The bcl-X-L/Tag cells expre ssed interleukin-l receptors 1 and 2, gelatinase B (MMP9), as well as Mac-1 , CD16/CD32 (Fc gamma receptors), CD45.2 (common leukocyte marker), CD86 (c ostimulatory molecule expressed on B cells, follicular dendritic cells, and thymic epithelium), major histocompatibility complex I, and nonspecific es terase when cocultured with MC3T3E1 cells. However, they did not express th e antigens for F4/80 (mature macrophage/dendritic cell marker) by immunosta ining. Treatment of bcl;X-L/Tag cells, cocultured with MC3T3E1 cells, with the combination of 1,25-dihydroxyvitamin D-3, and dexamethasone induced hig h levels of OCL formation. The bcl-X-L/Tag cells formed large numbers of OC Ls when cultured with RANK ligand and macrophage colony-stimulating factor in the absence of feeder cells. In the absence of RANK ligand and a feeder cell layer, 100% of the cells differentiated into F4/80-positive cells. How ever, neither PTH nor PTH-related protein enhanced OCL formation by bcl-X-L /Tag cells even when they were cocultured with primary osteoblasts, suggest ing that they differ from primary mouse bone mat-row cells in their respons iveness to PTWPTK-related protein. Thus, bcl-X-L/Tag cells have many of the properties of primary mouse OCL precursors and should be very useful for s tudies of OCL differentiation and divergence of OCL precursors from the mac rophage lineage.