High dose 1,25(OH)2D3 inhibits osteoblast mineralization in vitro

Vitamin D is essential for optimal calcium absorption needed for maintaining normal bone mineral density (BMD). Consequently, vitamin D-deficiency leads to poorly mineralized bone with diminished strength and load bearing capacity. Surprisingly, several animal and clinical studies have identified suppressive effects of high dose vitamin D supplementation on bone formation. These data suggest that while vitamin D is necessary for basal bone homeostasis, excessive concentrations may be detrimental to the skeleton. To further examine the direct effects of high dose vitamin D on the function of osteoblasts we differentiated primary osteoblast precursors and MC3T3 preosteoblastic cells, in the presence of supraphysiological doses of the active metabolite, 1,25-dihydroxyvitamin D-3 [1,25(OH)(2)D-3]. In vitro osteoblast mineralization was potently suppressed by high dose 1,25(OH)(2)D-3. To investigate the mechanism we used a bioassay to examine nuclear factor-KB (NF-kappa B) activation in MC3T3 cells. Although NF-kappa B agonists are generally potent inhibitors of osteoblast differentiation, surprisingly, 1,25(OH)(2)D-3 dose-dependently suppressed, rather than stimulated, NF-kappa B activation. Interestingly, 1,25(OH)(2)D-3 also suppressed Smad activation induced by the osteoblast commitment and differentiation factors transforming growth factor-beta (TGF-beta) and bone morphogenetic protein 2 (BMP2), which may account for the inhibitory activities of 1,25(OH)(2)D-3 on mineralization. Our data suggest that vitamin D has complex pleiotropic effects on osteoblast signal transduction. As the net balance of high dose 1,25(OH)(2)D-3 appears to be an inhibitory action on osteoblasts, our data suggest that the therapeutic value of vitamin D to maximize bone mass through indirect actions on calcium absorption may need to be carefully balanced with potential inhibitory direct effects on mineralizing cells. Our data suggest that indiscriminate over-dosing may be detrimental to bone formation and optimal concentrations need to be established for humans in vivo.