1α, 25-dihydroxyvitamin D 3 (1α, 25(OH) 2 D 3) acts on the osteoblasts to enhance the expressions of receptor activator of nuclear factor κB ligand (RANKL) and macrophage-colony stimulating factor (M-CSF) and induce the formation of osteoclasts. However, the mechanism in osteoblasts by which 1α, 25(OH) 2 D 3 promotes osteoclasto-genesis has not yet been completely understood. This study aimed to select the first generation of murine osteo-blasts to explore the underlying mechanism of 1α, 25(OH)

more »

... 2 D 3-induced osteoclastic formation from bone marrow mononuclear cells (BMMNCs). We discovered the activation of osteoblastic NF-κB pathway under 10-8 mol/L 1α, 25(OH) 2 D 3 treatment, as evidenced by the transfer of NF-κB p65 from cytoplasm to nuclei. Then, the NF-κB p65-siRNA was designed, constructed, and transfected into osteoblastic cells. Immunofluorescence assay confirmed the successfully silenced NF-κB p65 gene in osteoblasts. In the co-culture system of osteoblasts and BMMNCs with 1α, 25(OH) 2 D 3 added, the multinucleated osteoclast-like cells containing 2-3 nuclei were observed in BMMNCs co-cultured with non-transfection osteoblasts, conversely, silencing osteoblastic NF-κB p65 resulted in failed differentiation of BMMNCs along with substantial vacuolar degeneration in cytoplasm. In addition, the expressions of RANKL and M-CSF were notably decreased in NF-κB p65-silenced osteoblasts. Taken together, our data indicated that osteoblastic NF-κB pathway was involved in 1α, 25(OH) 2 D 3-induced osteoclast-like cells formation from BMMNCs through regulating the expression of RANKL and M-CSF. Therefore, our findings further identified the mechanism of 1α, 25(OH) 2 D 3-induced osteoclastogenesis on the basis of prior studies.