The role of recombinant human M-CSF, IL-3, GM-CSF and calcitriol in clonal development of osteoclast precursors in primate bone marrow.

Evidence suggests that among the myeloid colony-stimulating factors (CSFs), macrophage-CSF (M-CSF) is important in normal development of osteoclasts. To understand the role of M-CSF and other myeloid CSFs on clonal growth of osteoclast progenitors, isolated monkey bone marrow cells expressing the CD34 antigen were grown in soft agar or methylcellulose cultures in the presence of recombinant human M-CSF, granulocyte-macrophage CSF (GM-CSF), interleukin-3 (IL-3), calcitriol and various combinations of these factors. Day-10 to day-20 colonies were stained for tartrate-resistant acid phosphatase (TRAP) in order to identify putative osteoclast progenitors. Colonies were scored as follows: if greater than 80% of the cells in a colony contained TRAP it was scored as TRAP-positive; between 20% and 80%, TRAP-mix; and less than 20%, TRAP-negative. Further identification of osteoclast progenitors was made by simultaneous autoradiographic detection of calcitonin receptors on TRAP-stained colonies grown in methylcellulose. Among the 3 CSFs tested alone, GM-CSF stimulated the greatest number of TRAP-positive colonies; however, M-CSF and IL-3 in combination stimulated an even greater number of TRAP-positive colonies at all time points. Such additive effect was not observed between GM-CSF and IL-3. When calcitriol was added, the following increases in day-20 TRAP-positive colonies occurred: with M-CSF, 189%; with IL-3, 109%; with the combination of M-CSF and IL-3, 52%; and with GM-CSF there was no change. In the presence of calcitriol, 75% of IL-3 plus M-CSF-induced TRAP-containing colonies demonstrated calcitonin receptors over their TRAP-positive cells, whereas with M-CSF, GM-CSF or IL-3 each alone, 0 to 25% of TRAP-containing colonies had calcitonin receptor over TRAP-positive cells. These results suggest that IL-3, M-CSF and calcitriol stimulate a subpopulation of hemopoietic progenitors, likely osteoclast progenitors, that can give rise to TRAP-positive cells simultaneously expressing calcitonin receptors.