Role of Ascorbic Acid in the Osteoclast Formation: Induction of Osteoclast Differentiation Factor with Formation of the Extracellular Collagen Matrix*

Osteoclasts are bone-resorbing multinucleated cells. Tartrate-resistant acid phosphatase-positive (TRAP-positive) mononuclear and multinucleated cells, which are osteoclast-like cells (OCLs), were formed as a result of the coculture of mouse bone marrow cells and clonal stromal ST2 cells in the presence of 1alpha,25-dihydroxy-vitamin D3. Removal of ascorbic acid from the culture medium prevented the formation of TRAP-positive OCLs. Addition of ascorbic acid to the medium formed TRAP-positive OCLs, and the effect of ascorbic acid was dose-dependent. When we examined the level of messenger RNA (mRNA) for osteoclast differentiation factor (RANKL/ODF) in ST2 cells, we found that ascorbic acid caused an approximately 5-fold increase in the level of this mRNA. The half-life of the mRNA was unaffected by ascorbic acid. To characterize the mechanism of action of ascorbic acid, we investigated the relationship between formation of TRAP-positive OCLs and formation of the collagen matrix. Inhibitors of the formation of collagen triple helices blocked both the formation of TRAP-positive OCLs and the expression of the mRNA for RANKL/ODF in response to ascorbic acid. Our findings suggest that ascorbic acid might be essential for osteoclastogenesis and might induce the formation of TRAP-positive OCLs via induction of the synthesis of RANKL/ODF that is somehow mediated by the extracellular matrix.