Recently, membrane-type matrix metalloproteinase-1 (MT1-MMP) was identified, and found that it can activate proMMP-2 on the cell membrane, degrade bone matrix, and participate in bone formation. Since bone is a target tissue of 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3], in the present study we observed the effects of 1alpha,25(OH)2D3 on MTI-MMP expression, and proMMP-2 activation in normal human ostcoblast-like cells (hOB). Western immunoblots showed 1alpha,25(OH)2D3 time and dose dependently stimulated MTI-MMP production. By ELISA, we found that the activation of proMMP-2 in cultures of hOB was intensified by 1alpha,25(OH)2D3. Our studies suggest that 1alpha,25(OH)2D3 induces MMP-2 activation in part by up-regulating MTI-MMP expression, and since MTI-MMP plays a role in bone metabolism, the induction of MT1-MMP levels by 1alpha,25(OH)2D3 in hOB cells may contribute to a new mechanism by which 1alpha,25(OH)2D3 promotes bone formation and stimulates bone resorption.