Direct action of naturally occurring estrogen metabolites on human osteoblastic cells

This article describes experiments that were performed to examine the direc t action of estrogen metabolites on cultured human osteoblast cells. The hu man fetal osteoblastic cell line, hFOB/ER9, which expresses high levels of the estrogen receptor (ER) alpha, was used to examine the direct effects of 16 alpha-hydroxyestrone (16 alpha-OHE1) and 2-hydroxyestrone (2-OHE1) on o steoblast differentiation. The 16 alpha-OHE1 caused a decrease in osteocalc in (OC) secretion to a maximum of 40% of control values (vehicle-treated ce lls) at 10(-7) M. Alkaline phosphatase (AP) activity was significantly indu ced at 10-7 M 16 alpha-OHE1 with greater than 500% of control at 10(-6) M 1 6 alpha-OHE1. Finally, AP steady-state messenger RNA (mRNA) levels were inc reased within 24 h of 16 alpha-OHE1 treatment. In contrast to 16 alpha-OHE1 , 2-OHE1 had no effects on the secretion of OC, AP activity, or AP gene exp ression. The 2-OHE1 also did not display any antiestrogen activity because treatment in combination with 17 beta-estradiol (E-2) and 16 alpha-OHE1 had no significant effect on the reduction in OC secretion or induction of AP activity. Similar to E-2, 16 alpha-OHE1 stimulated the expression of an ear ly response gene, a TGF-beta inducible early gene, designated TIEG, as earl y as 60 minutes after treatment, whereas treatment with 2-OHE1 displayed no effect. Support that the 16 alpha-OHE1 regulation of these osteoblasts (OB ) markers was mediated through the ER is shown by the fact that the estroge n antagonist ICI 182,780 abrogated these effects. These data suggest that 1 6 alpha-OHE1 is a potent estrogen agonist on human osteoblastic hOB/ER9 cel ls. In contrast, 2-OHE1 displayed no estrogenic or antiestrogenic activity in this human osteoblast cell model.