Because postmenopausal osteoporosis, one of the major health problems in women, results from endogenous estrogen deficiency, the roles of estrogens and estrogen-like molecules, including isoflavones, in regulating bone cell activities are central to understanding the etiology, prevention, and treatment of this disease. Isoflavones have been shown to enhance bone retention in young ovariectomized (OVX) animal studies and in two trials of peri- or post-menopausal women. The cellular mechanisms by which isoflavones improve bone retention remain, however, uncertain. Three hypotheses were tested by the in vitro studies presented here: (1) isoflavones modulate the synthesis or mRNA expression of osteoclastogenesis-regulatory (OCR) proteins synthesized by osteoblastic cells; (2) regulation of OCR cytokines by isoflavones is estrogen receptor (ER)-dependent; (3) nuclear factor-kappa B (NF-kappaB), central to the control of interleukin-6 (IL-6) synthesis, is regulated by isoflavones.;During osteoblastic differentiation, isoflavones, at dietarily achievable concentrations, have been found to decrease IL-6 production and to increase osteoprotegerin (OPG) expression in MC3T3-E1, hFOB1.19, and hFOB/ER9 osteoblastic cells. The reduced IL-6 synthesis and stimulated expression of OPG by isoflavones were related to the different levels of ER expression in two hFOB osteoblastic cell lines. Responses to isoflavones were much stronger in the hFOB/ER9 cell line with higher numbers of ERs. After adding ICI-182,780, an ER blocker, the effects of isoflavones on OPG and IL-6 were reduced or neutralized. In addition, treatments of early differentiating hFOB/ER9 osteoblasts with isoflavones clearly inhibited NF-kappaB DNA-binding activities, and IL-6 synthesis. Also, isoflavone treatments were found to block the TNF-alpha-induced IL-6 production and NF-kappaB DNA-binding activities in hFOB/E9 cells. The addition of the anti-estrogen, ICI-182,780, reduced the inhibitory effects of isoflavones on NF-kappaB DNA-binding activities.;In summary, isoflavones, at dietarily achievable levels, are able to modulate the production of OCR cytokines in osteoblasts. These effects of isoflavones on OCR cytokines suggest that isoflavones may indirectly inhibit osteoblastic resorption through their actions on osteoblastic cells. Inhibition of IL-6 production by isoflavones results from an inhibition of NF-kappaB activation. At nutritionally-relevant concentrations, these effects of isoflavones on osteoblastic cells are mediated via an ER-dependent pathway.
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