cAMP promotion of osteoclast-like cell development from mouse bone marrow cells requires a permissive action of...

Recruitment of osteoclasts from monocytic precursors is modulated by local signals. We previously showed that monoblastic differentiation in U937 cells is stimulated by 1,25-(OH)2D3 and cAMP in series. We investigate here the combined effects of these agents to stimulate differentiation of osteoclast-like cells from mouse marrow. Cells from mouse marrow were harvested and cultured in alpha-MEM with 10% fetal bovine serum. The appearance of tartrate-resistant acid phosphatase-containing multinuclear cells was measured after 8 days in culture by cytochemical staining. Continuous exposure of cultures to 10 nM 1,25-(OH)2D3 positively stimulated development of these cells after 8 days (101 +/- 3 cells per well, n = 74). No osteoclast-like cells were found when 1,25-(OH)2D3 was added for the first 4 days followed by 4 days more with no treatment. PGE2 (1 microM) as a single agent added during the last 4 days of culture was not able to recruit osteoclast-like cells. However, cultures exposed to 1,25-(OH)2D3 during the first 4 days and 1 microM PGE2 during the second 4 days developed osteoclast-like cells at 8 days [66 +/- 8% of the formation seen with 1,25-(OH)2D3 alone, p less than 0.05]. Dibutyryl cAMP (1 microM to 3 mM) was also not effective used as a single agent, but was able to stimulate formation of TRAP-positive multinuclear cells when 1,25-(OH)2D3 preceded its addition to culture medium. cAMP analogs therefore mimicked the effect of 1 microM PGE2, but these experiments do not allow us to assign the PGE2 action entirely to activation of cAMP second messenger.(ABSTRACT TRUNCATED AT 250 WORDS)