Heat-treated osteoblastic cell (ROS17/2.8)-conditioned medium induces the formation of osteoclast-like cells

To study the effects of osteoblast products on osteoclast formation, we added the conditioned medium (CM) of rat osteoblastic cell line ROS 17/2.8 to rat bone marrow cultures, in which tartrate-resistant acid phosphatase (TRAP)-positive osteoclast-like multinucleate cells (MNCs) formed in the presence of 10⁻⁸ M 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃). The formation of 1,25(OH)₂D₃-dependent TRAP-positive MNC at day 7 of culture was strongly inhibited by the >10 kDa fraction of ROS 17/2.8 cell-CM (ROSCM), but heat treated ROSCM (htROSCM) expressed marked stimulation in the formation of the MNCs. The expression of several osteoclastic phenotypes of the MNCs induced by htROSCM and 1,25(OH)₂D₃ was more enhanced compared with that of the MNCs induced by 1,25(OH)₂D₃ alone. The MNCs induced by htROSCM and 1,25(OH)₂D₃ were highly motile, were sensitive to calcitonin (CT), and had high bone resorbing activity. These data suggest that htROSCM promotes the osteoclast differentiation in the presence of 1,25(OH)₂D₃ in a rat bone marrow culture system. The stimulatory activity of TRAP-positive MNC formation in htROSCM is derived from heat-stable protein(s) that is (are) thought to be different from colony-stimulating factors (CSFs) such as macrophage-CSF (M-CSF) or granulocyte-macrophage-CSF (GM-CSF).