Key Laboratory of Animal Physiology & Biochemistry, Nanjing Agricultural University, Nanjing 210095, PR China.
Steroids. 2010 Mar;75(3):245-51. doi: 10.1016/j.steroids.2009.12.009. Epub 2010 Jan 4.
Two in vitro systems were employed to delineate the estrogenic activity of daidzein (Da), alone or in combination with high or low concentrations of estrogen in two cell types possessing different estrogen-receptor (ER) isoforms, ERalpha and/or ERbeta: (1) vitellogenin II (VTG), the egg yolk precursor protein and the endpoint biomarker for estrogenicity, in chicken primary hepatocytes, and (2) CHO-K1 cells transiently co-transfected with ERalpha or ERbeta and estrogen-response elements (ERE) linked to a luciferase reporter gene. Da (100 microM) alone induced VTG mRNA expression in chicken hepatocytes, albeit with much less potency compared to estradiol (E(2)). Da exhibited different effects in the presence of 1 microM and 10 microM E(2). At a concentration of 100 microM, Da enhanced 1 microM E(2)-induced VTG transcription by 2.4-fold, but significantly inhibited 10 microM E(2)-induced VTG mRNA expression in a dose-dependent fashion from 1 to 100 microM. Tamoxifen completely blocked the estrogenic effect of daidzein, alone or in combination with 1 microM of E(2), but did not influence its anti-estrogenic effect on 10 microM E(2)-induced VTG mRNA expression. Furthermore, neither E(2) nor daidzein, alone or in combination, affected ERalpha mRNA expression, yet all the treatments significantly up-regulated ERbeta mRNA expression in chicken hepatocytes. E(2) effectively triggered estrogen-response elements (ERE)-driven reporter gene transactivation in CHO-K1 cells expressing ERalpha or ERbeta and showed much greater potency with ERalpha than with ERbeta. In contrast, daidzein was 1000 times more powerful in stimulating ERbeta- over ERalpha-mediated transactivation. Daidzein, in concentrations ranging from 5 nM to 50 microM, did not affect ERbeta-mediated transactivation induced by 1 nM E(2), but it significantly inhibited ERbeta-mediated transactivation induced by 10 nM E(2) at 500 nM. Despite the tremendous difference in sensitivity between the two in vitro systems, daidzein exhibited greater potency as an estrogen-antagonist for ERbeta-mediated activity.
采用两种体外系统，分别在两种具有不同雌激素受体（ER）同工型 ERα和/或 ERβ的细胞类型中，研究大豆苷元（Da）单独或与高、低浓度雌激素联合作用的雌激素活性：（1）卵黄蛋白原 II（VTG），即卵黄前体蛋白，也是雌激素活性的终点生物标志物，在鸡原代肝细胞中；（2）瞬时转染 ERα或 ERβ和与荧光素酶报告基因相连的雌激素反应元件（ERE）的 CHO-K1 细胞。Da（100 μM）单独作用于鸡肝细胞时可诱导 VTG mRNA 表达，但其效力远低于雌二醇（E2）。Da 在 1 μM 和 10 μM E2 存在时表现出不同的作用。在 100 μM 浓度下，Da 使 1 μM E2 诱导的 VTG 转录增强 2.4 倍，但以剂量依赖性方式从 1 至 100 μM 显著抑制 10 μM E2 诱导的 VTG mRNA 表达。他莫昔芬完全阻断了大豆苷元单独或与 1 μM E2 联合作用的雌激素作用，但不影响其对 10 μM E2 诱导的 VTG mRNA 表达的抗雌激素作用。此外，E2 或大豆苷元单独或联合作用均不影响 ERα mRNA 表达，但所有处理均显著上调鸡肝细胞中 ERβ mRNA 表达。E2 有效地触发了表达 ERα或 ERβ的 CHO-K1 细胞中雌激素反应元件（ERE）驱动的报告基因转录激活，且与 ERα 相比，其效力更高。相反，大豆苷元在刺激 ERβ-过 ERα介导的转录激活方面比 E2 强 1000 倍。Da 在 5 nM 至 50 μM 浓度范围内不影响 1 nM E2 诱导的 ERβ 介导的转录激活，但在 500 nM 时显著抑制 10 nM E2 诱导的 ERβ 介导的转录激活。尽管两种体外系统的敏感性存在巨大差异，但大豆苷元在作为 ERβ 介导的活性的雌激素拮抗剂方面表现出更高的效力。
