Nonhypercalcemic analogs of vitamin D stimulate creatine kinase B activity in osteoblast-like ROS 17/2.8 cells and up-regulate their responsiveness to estrogens.
We have reported that pretreatment with 1 alpha, 25(OH)2D3(1, 25) up-regulates responsiveness and sensitivity to 17 beta estradiol (E2) in osteoblast-like cells, as measured by parallel stimulation of [3H]thymidine incorporation into DNA and the specific activity of creatine kinase BB (CK). Increased responsiveness was correlated with increased E2 receptor concentration. In this study, we have extended these observations to new nonhypercalcemic analogs of 1,25. We compared the analogs hexafluoro vitamin D3 (FL), and the side chain modified derivatives: EB 1089 (EB), CB 1093 (CB) and MC 1288 (MC) with 1,25 and 25 (OH)D3(25 D3). Treatment with 30 nM E2 for 4 h stimulated CK activity in ROS 17/2-8 cells by 40%; there was no further increase after 3 daily additions of E2. Treatment by 3 daily additions, at 1 nM, of all analogs except 25 D3 caused a 2-3-fold increase in CK specific activity. This schedule of treatment also upregulated the response to 4 h exposure to 30 nM E2 by 30-70% above the response to vitamin D analogs alone, and by up to 2 fold compared to E2 without pretreatment. At 1 pM, the analogs doubled CK activity, and, except for 1,25, upregulated the response to E2 to levels characteristic of each analog. Pretreatment with vitamin D analogs also increased the sensitivity to E2 by lowering the dose for a comparable response to E2 by one or two orders of magnitude. Stimulation of specific activity of CK by the analogs was paralleled by increases in the steady state level of mRNA for CKB, but not in its half life. Whereas pretreatment by vehicle followed by E2 for 2 h was unable to increase CKB mRNA, pretreatment with the analogs made possible detection of mRNA responsiveness to E2. These results add to the evidence for the interaction of estrogens and antiestrogens with vitamin D metabolites in regulation of bone growth in vitro. They also strengthen the potential for treatment of bone loss, as occurs in postmenopausal osteoporosis, by a combination of nonhypercalcemic vitamin D analogs and estrogens.
Sömjen
D
D
Endocrine Unit, Tel-Aviv Sourasky Medical Center, Israel.
Waisman
A
A
Weisman
J
J
Kaye
A M
AM
D000818:Animals; D003402:Creatine Kinase / Q000378:metabolism*; D004789:Enzyme Activation; D004967:Estrogens / Q000502:physiology*; D006934:Hypercalcemia; D007527:Isoenzymes; D010006:Osteoblasts / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D051381:Rats; D014407:Tumor Cells, Cultured; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000494:pharmacology
D004967:Estrogens; D007527:Isoenzymes; D014807:Vitamin D; D003402:Creatine Kinase