Treatment with non-hypercalcemic analogs of 1,25-dihydroxyvitamin D 3 increases responsiveness to 17β-estradiol, dihydrotestosterone or raloxifene in primary human osteoblasts

Abstract

Pretreatment with 1 nM 1,25-dihydroxyvitamin D 3 (1,25), or non-hypercalcemic Vitamin D analogs, upregulated the response of creatine kinase (CK) to 17β-estradiol (30 nM E 2), raloxifene (3000 nM RAL) or dihydrotestosterone (300 nM DHT) in primary human bone cells. Previously, we reported that these osteoblast-like cells responded to gonadal steroids in a sex specific manner. Bone cells derived from pre-menopausal women showed greater stimulation of CK specific activity by E 2 than bone cells from post-menopausal women; in male-derived cells no age related difference was found. In this study, we treated cells derived from female or male bones, at different ages, with the side chain modified analogs of Vitamin D: CB 1093 (CB), EB 1089 (EB), MC 1288 (MC) and the demonstrably non-calcemic hybrid analog JK 1624 F2-2 (JKF), by daily addition of 1 nM, for 3 days. On day 4, cells were incubated with sex steroids for 4 h and cell extracts were prepared. Pretreatment with JKF or CB significantly upregulated the response to 30 nM E 2 in all female-derived cells and to 300 nM DHT in mature male-derived cells. In cells from older males, only JKF caused augmentation of DHT action. Bone cells from pre- or post-menopausal females responded to 3000 nM RAL by increased CK activity to the same extent as to 30 nM E 2; however, RAL and E 2, when applied together, resulted in mutual annihilation of their agonist activities. Vitamin D analogs prevented the antagonistic effect of RAL in the presence of E 2, possibly due to increased numbers of ERs. Both estrogen receptors, α (ERα) and β (ERβ), were expressed in male- as well as in female-derived cells. However, only in female-derived cells were ERα and ERβ upregulated by pretreatment with Vitamin D analogs. This study raises the possibility of testing combined Vitamin D analog and estrogen replacement treatment for post-menopausal women to prevent osteoporosis.