OSTEOCLAST DEVELOPMENT FROM HEMATOPOIETIC STEM-CELLS - APPARENT DIVERGENCE OF THE OSTEOCLAST LINEAGE PRIOR TO MACROPHAGE COMMITMENT

To further clarify the progression of osteoclast development, the rela tionship of clonogenic osteoclast progenitors (CFU-O) to macrophage or more primitive progenitors was examined. Serum-free culture supernata nt of a tumor clone (CESJ) was used as a source of an osteoclast colon y stimulating factor (O-CSF). CFU-O-derived colonies were identified b y their characteristic positive staining for tartrate resistant acid p hosphatase (TRAPase). The effect of macrophage colony stimulating fact or (M-CSF) and stem cell factor (SCF) on osteoclast progenitors was ex amined by pre-culturing mouse bone marrow (BM) cells in agar medium co ntaining M-CSF or SCF and overlaying CESJ medium 0-7 days later. The n umber of TRAPase(+) colonies decreased while TRAP(-) macrophage coloni es increased in M-CSF pre-cultures as overlays of CESJ medium were del ayed. On the other hand, TRAPase(+) and mixed colonies persisted in SC F pre-cultures with CESJ medium overlays. Conversely, all colonies wer e TRAPase(+) and no macrophage colonies developed in O-CSF pre-culture s overlaid with M-CSF. CFU-O, but not CFU-M, survived 7 days without e xogenous CSFs in agar medium. In fractionated BM, the majority (>99%) of CFU-O were in the c-kit positive population; however, a specific an tibody to SCF did not affect O-CSF-induced TRAPase(+) colony formation , suggesting the proliferation and differentiation of osteoclast proge nitors are independent of c-kit-SCF interactions. These studies provid e further experimental evidence to support the concept that O-CSF acts on progenitors in earlier stages of development, supporting their dif ferentiation into the osteoclast lineage prior to macrophage commitmen t.