2-hydroxyestradiol and indole-3-carbinol

To gain new insight into the roles of cruciferous vegetable-derived bioactive phytochemicals in bone cells, we investigated the effects of indole-3-carbinol (I3C) on cell proliferation and differentiation in estradiol (E2)-exposed calvarial osteoblasts that were obtained from neonatal rats.. Osteoblast activity was assessed by analyzing cellular DNA, cell-associated osteocalcin (OC) levels and alkaline phosphatase (AP) activity. We also examined [(3)H]-estrone (E1) metabolism and estrogen-agonistic and estrogen-antagonistic activities of 2-hydroxy (OH) E1 and 2-OHE2 and their capacity to displace [(3)H]-E2 at ER binding sites using competition studies.. I3C did not affect on cellular DNA, OC levels or AP activity. However, I3C completely inhibited E2-induced increases in cell proliferation and differentiation in neonatal rat osteoblasts. Metabolic studies demonstrated that I3C promoted the conversion of [(3)H]-E1 to 2-OHE1 and 2-OHE2 and those higher rates of conversion (twofold-threefold) were archived when a higher dose of I3C was applied. Proliferation and differentiation studies showed that 2-OHE2 but not 2-OHE1 inhibited E2-induced increases in cell proliferation and differentiation via an ER-mediated mechanism. Likewise, Esr1 was expressed at high level than Esr2. 2-OHE1 showed no activity or affinity for ER.. This study is the first to show that a bioactive compound derived from cruciferous vegetables, I3C, abolishes the E2-mediated stimulation of cell activities including, proliferation and differentiation, in rat osteoblasts and increases the 2-hydroxylation of E1, resulting in the formation of inactive and anti-estrogenic metabolites. These results suggest that in neonatal rat osteoblasts, the anti-estrogenic effect of I3C is mediated by 2-OHE2 through ER-α.

Topics: Animals; Animals, Newborn; Anticarcinogenic Agents; Cell Differentiation; Cell Proliferation; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Estradiol; Estrogen Antagonists; Estrogen Receptor Antagonists; Female; Indoles; Osteoblasts; Rats; Rats, Wistar; Real-Time Polymerase Chain Reaction; Receptors, Estrogen; Reverse Transcriptase Polymerase Chain Reaction

Compounds like indole-3-carbinol (I3C) have been shown to increase catechol estrogen formation and reduce mammary tumor incidence in mice. These compounds may exert a protective effect for breast cancer development by decreasing the overall estrogen pool available for the formation of 16 alpha-hydroxyestrone (16 alpha-OHE1), a metabolite that retains significant estrogenic activity, may be mutagenic and could represent a potential carcinogenic intermediate of estradiol degradation. I3C and ascorbigen originate from the breakdown of glucobrassicin. We have compared the inductive effects of I3C with ascorbigen and beta-naphthaflavone (Bnf) in microsomes from rats pretreated with these compounds using isotope dilution GC-MS and a radiometric method. Incubated microsomes from rats pretreated with I3C and ascorbigen yielded high levels of 2-hydroxyestradiol (2-OHE2) that were comparable to levels induced by Bnf and were significantly above control group levels (p < 0.005). Absolute values determined by the radiometric method were approximately 40% lower than 2-OHE2 concentrations determined by GC-MS, although the relative changes in each group were the same. These differences may be attributed to the radiolabel becoming trapped in microsomal intermediates in the sequence leading to tritium entering the aqueous compartment. Both ascorbigen- and Bnf-treated animals exhibited significant increases in 2-hydroxyestrone (2-OHE1) (p < 0.05). The ability of ascorbigen to induce estradiol C-2 hydroxylation has not been previously reported. Based on these data, we speculate that ascorbigen will act as an anticarcinogenic agent and will inhibit or reduce the incidence of mammary tumor formation.

Topics: Animals; Ascorbic Acid; Benzoflavones; beta-Naphthoflavone; Estradiol; Estrogens, Catechol; Female; Gas Chromatography-Mass Spectrometry; Hydroxyestrones; Hydroxylation; Indoles; Microsomes, Liver; Radiometry; Rats; Rats, Sprague-Dawley