REGULATION OF C-FOS EXPRESSION AND TGF-BETA PRODUCTION BY GONADAL AND ADRENAL ANDROGENS IN NORMAL HUMAN OSTEOBLASTIC CELLS

Although the role of estrogens in bone formation is becoming clarified, the function of androgens in this process remains to be defined. Consequently, we have explored the mechanism of action for both gonadal and adrenal androgens in normal human osteoblastic (hOB) cells, which are responsible for the synthesis and mineralization of bone. Changes in the steady-state mRNA levels for two nuclear proto-oncogenes (c-fos and c-jun) and one cytokine (TGF-beta 1) were quantified in response to short (30 min) and long (24-48 h) treatments of these cells with physiologic concentrations of steroids. In addition, the levels of TGF-beta protein in the hOB cell conditioned-media were measured using a bioassay. The results indicated that neither 10 nM dihydrotestosterone, 10-20 nM testosterone, nor 10-100 nM androstenedione had a significant effect on the steady-state levels of c-fos, c-jun, or TGF-beta 1 mRNAs. Interestingly, 10-1000 nM dehydroepiandrosterone (DHEA) and 1-10 mu M DHEA-sulfate rapidly reduced the steady-state level of c-fos mRNA by 60-80% in a dose-dependent manner within 30 min. In contrast, neither of these adrenal steroids had a significant effect on the message levels for c-jun or TGF-beta 1. Surprisingly, although TGF-beta 1 mRNA levels remained unchanged, the total amount of TGF-beta activity in the hOB cell conditioned-media increased 2-5-fold in response to 24-48 h treatments of the cells with gonadal or adrenal androgens. This increase in TGF-beta activity by DHEA-sulfate was both time- and dose-dependent, and was not blocked by cotreatment with the specific aromatase inhibitor 4-hydroxyandrostenedione (1 mu M). Immunoprecipitations of the hOB cell conditioned-media with isoform-specific TGF-beta neutralizing-antibodies indicated that TGF-beta 2 was predominantly produced by the cells in response to DHEA-sulfate treatment. These results demonstrate that differences exist between the actions of estrogens and androgens on normal human osteoblasts with regard to the regulation of c-fos expression and TGF-beta production. Moreover, these data suggest that DHEA and DHEA-sulfate may play a distinct role in the regulation of human osteoblast function via the rapid repression of c-fos message levels and the slower increase in TGF-beta 2 protein levels.