Several types of specific insulin-like growth factor binding proteins have been reported. These binding proteins are produced by peripheral tissue-derived cells and they modulate the functions of insulin-like growth factors. In this study, we investigated both the secretion of insulin-like growth factor binding protein 3 (IGFBP-3) from a human osteosarcoma cell line MG63, and the effects of 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) on the production of this binding protein. The beta subunit of IGFBP-3 was detected in perinuclear cytoplasm of MG63 cells by immunocytochemical study. Immunoblotting and SDS-PAGE analysis revealed that both 150KD MW entire molecules and 40-60KD MW beta subunit molecules of IGFBP-3 were present in cell-conditioned media. 1,25-(OH)2D3 stimulated the production of the IGFBP-3 molecule by MG63 cells. The concentration of IGFBP-3 in conditioned media began to rise at 12 hours after the addition of 10(-8) M of 1,25-(OH)2D3 and reached peak level at 48 hours. Dose-dependent effects of 1,25-(OH)2D3 were demonstrated. The its maximum effect was observed at 10(-10) M. The concentration of IGFBP-3 in cytosol also increased at a 10(-10) M concentration of 1,25-(OH)2D3. We conclude from these results that human osteosarcoma cells MG63 produce the IGFBP-3 molecule and that 1,25-(OH)2D3 stimulates the production of this protein. These data suggests that the synergistic effects of 1,25-(OH)2D3 on the action of IGF-I on osteoblastic cells, which we reported previously, may be modulated by locally produced IGFBP-3.