Evidence that estrogens modulate activity and increase the number of 1,25-dihydroxyvitamin D receptors in osteoblast-like cells (ROS 17/2.8)

A detailed understanding of the mechanism of action of estrogen on bones is still lacking. The present study was designed to examine possible modulation by 17β-estradiol (E₂) on the effects of 1,25-dihydroxyvitamin D₃ [1,25-(OH)₂D₃] and on vitamin D receptors (VDR) in the ROS 17/2.8 osteoblast-like cell line. Cells were grown in phenol-red free medium supplemented with charcoal-stripped fetal calf serum (FCS). Total cellular VDR were measured in cell homogenates after extraction with a KCl hypertonic buffer. VDR-binding capacity doubled in the presence of 10 nM E₂ (16.2 ± 2.3 vs. 7.0 ± 1.3 fmol/mg protein, respectively; P < 0.01), while the K_d for 1,25-(OH)₂D₃ did not change (∼0.1 nM). Tamoxifen alone had no effect on VDR, while it completely abolished the E₂-induced increase in VDR, indicating that the effect was specific for E₂ and estrogen receptor mediated. 1,25-(OH)₂D₃ inhibited cell proliferation, determined by [³H] thymidine incorporation to DNA, in a dose-dependent fashion between 0.01-100 nM. The inhibitory effect of 1,25-(OH)₂D₃ on cell proliferation was significantly augmented in the presence of E₂ (10 nM). 1,25-(OH)₂D₃ increased osteocalcin secretion to the medium by the cells in a dose-dependent fashion between 0.01-100 nM. In the presence of E₂ (10 nM), maximal osteocalcin secretion in response to 1,25-(OH)₂D₃ was 3.5-fold higher than that in response to 1,25-(OH)₂D₃ alone. These results indicate that E₂ modulates 1,25-(OH)₂D₃ activity in osteoblast-like cells, and that this effect can be attributed to an increase in VDR.