THE BIOLOGICAL ROLES OF THE 3RD COMPONENT OF COMPLEMENT IN OSTEOCLASTFORMATION

We previously reported that 1alpha,25-dihydroxyvitamin D3 [1alpha,25-( OH)2D3] tissue-specifically stimulated the production of the third com ponent of complement (C3) in bone in vitro and in vivo. In the present study, we examined the possible roles of C3 in bone using bone marrow cultures with antibodies against C3 and C3 receptors (Mac 1, 8C12, an d 7G6) and the purified mouse C3 protein. The C3 protein produced pref erentially by stromal cells in response to 1alpha,25-(OH)2D3 was distr ibuted in macrophage-like mononuclear cells and small cells with few n uclei. Adding anti-C3 antibody together with 1alpha,25-(OH)2D3 to bone marrow cultures greatly inhibited not only the appearance of tartrate -resistant acid phosphatase (TRAP)-positive mononuclear and multinucle ated cells, but also the growth of macrophage-like mononuclear cells a nd stromal cells. The inhibitory effect of anti-C3 antibody on osteocl ast-like cell formation was most prominent when it was added between d ays 2-4 of the 6-day culture period, which corresponded to the late pr oliferative phase and the early differentiation phase of osteoclast de velopment. Adding anti-C3 receptor antibodies also inhibited osteoclas t-like cell formation induced by 1alpha,25-(OH)2D3. When C3 receptors were detected by the binding of C3-coated sheep red blood cells or imm unostaining, the localization of C3 receptor-positive cells coincided exactly with that of C3 protein-positive cells. C3 receptors were expr essed mainly in macrophage-like mononuclear cells, TRAP-positive monon uclear cells, and TRAP-positive small cells with few nuclei. TRAP-posi tive large cells with many nuclei were totally negative for C3 recepto rs. When macrophage-colony-stimulating factor (M-CSF), the purified C3 protein, and 1alpha,25-(OH)2D3 were added to bone marrow methylcellul ose cultures, separately or in combination, M-CSF-dependent colony for mation was strikingly inhibited by 1alpha,25-(OH)2D3, but the inhibiti on was prevented by simultaneously adding C3. These results provide ad ditional evidence that osteoclast progenitors are indeed cells of the monocyte-macrophage lineage. It is likely that the C3 produced by stro mal cells in response to 1alpha,25-(OH)2D3 is somehow involved in oste oclast development by potentiating M-CSF-dependent proliferation of bo ne marrow cells and induction of osteoclast differentiation.