Botanical Modulation of AhR-ERα by Crosstalk Inhibitors Promotes Estrogen (E2) Detoxification

串扰抑制剂对AHR-ERα的植物调控促进雌激素(E_2)解毒 雌激素受体(ER)阳性的乳腺癌对绝经后妇女的健康构成重大风险， 部分由雌激素(E_2)代谢物--雌二醇-3，4-苯二酚调节，它能产生净化加合物。 并导致突变。P450 1b1是雌二醇转化为4-羟基化的主要酶。 产物，可导致遗传毒性的4-苯二酚，而P450 1A1，通常是表观遗传抑制 通过E2激活的雌激素受体α(ERα)，将雌二醇转化为一种非遗传毒性的2-羟化产物。 激活的芳香烃受体诱导ERα的降解及其转录 分别翻译成P450 1A1和1b1。激活剂和配体 AHR(串扰抑制药)已被证明优先上调CYP1A1，并最终上调 非遗传毒性的2-羟基雌二醇代谢产物(雌激素解毒途径)，通过下调 细胞色素P450 1A1的表观遗传抑制。越来越多的女性转向植物性膳食补充剂(BDS)， 替代传统激素替代疗法(HRT)的研究结果发布以来，雌激素+ 黄体酮会增加患乳腺癌的风险。有趣的是，淫羊藿苷是淫羊藿属中的一种生物活性化合物，a. 用于妇女健康目的的植物，已被证明能激活AhR。这是一个假设 建议一些女性保健植物含有抗雌激素、AhR激活化合物，这些化合物 降解ERα(Aim 1)和逆转表观遗传的CYP1A1抑制优先激活雌激素 体外细胞培养模型(AIM 2)和体内去卵巢大鼠模型(AIM 3)的解毒途径。 抑制Ishikawa细胞雌激素依赖的碱性磷酸酶活性，以及未能抑制A 植物类化合物荧光偏振酶检测雌二醇-ERα复合体的研究 提供不通过ERα发挥作用的抗雌激素化合物。这些化合物将在- MCF7(ER+)细胞对ERα抗体的免疫抑制作用，以寻找能降解ERα的化合物。其他内容 在HepG2细胞中XRE-荧光素酶的活性将表明AhR激活化合物，而甲基化减少 通过DNMT和XRE的芯片检测，以及CyP1A1/1b1的qRT-PCR将表明 雌二醇介导的表观遗传抑制导致雌激素解毒途径上调 通过体外相声抑制剂。这一前提将被带到一个去卵巢的大鼠模型上，该模型涉及雌二醇和 在处死和分析子宫重量前的植物化合物，进行E2的LC-MS分析 代谢产物，并用免疫组织化学方法定量ERα表达水平。这项研究可能会揭示 一些具有生物活性的女性健康BDS化合物表现出抗雌激素结果的机制，以及 体内绝经后模型中E2解毒途径在促进健康方面的重要性。

Botanical Modulation of AhR-ERα by Crosstalk Inhibitors Promotes Estrogen (E2) Detoxification Estrogen receptor (ER) positive breast cancer poses significant health risks for postmenopausal women, and is in part mediated by the estrogen (E2) metabolite, estradiol-3,4-quinone, which causes depurinating adducts and leads to mutations. P450 1B1 is the primary enzyme for the conversion of estradiol to the 4-hydroxylated product, which can result in a genotoxic 4-quinone, while P450 1A1, which is normally epigenetically inhibited by E2-activated estrogen receptor alpha (ERα), converts estradiol to a nongenotoxic 2-hydroxylated product. Activated aryl hydrocarbon receptor (AhR) induces degradation of ERα as well as the transcription of both CYP1A1 and CYP1B1, which are translated into P450 1A1 and 1B1 respectively. Activators and ligands of AhR (Crosstalk Inhibitors) have been shown to preferentially upregulate CYP1A1 and ultimately the nongenotoxic 2-hydroxylated estradiol metabolite (estrogen detoxification pathway), through downregulation of the epigenetic inhibition of CYP1A1. Women have increasingly turned to botanical dietary supplements (BDS), instead of traditional hormone replacement therapy (HRT), since the release of the findings that estrogen + progesterone increases breast cancer risk. Interestingly, icaritin, a bioactive compound from Epimedium sp., a botanical used for women's health purposes, has been shown to activate AhR. The hypothesis of this proposal is that some women's health botanicals contain antiestrogenic, AhR activating compounds which degrade ERα (Aim 1) and reverse epigenetic CYP1A1 inhibition to preferentially activate the estrogen detoxification pathway in an in vitro cell culture model (Aim 2) and an in vivo ovariectomized rat model (Aim 3). Inhibition of estrogen-dependent alkaline phosphatase activity in Ishikawa cells, and a failure to inhibit a fluorescent estradiol-ERα complex by botanical compounds in a fluorescence polarization enzyme assay will provide antiestrogenic compounds which do not act through ERα. These compounds will be subjected to in- cell western of MCF-7 (ER+) cells against an ERα antibody to find compounds which degrade ERα. Additional XRE-luciferase activity in HepG2 cells will indicate AhR activating compounds, while a decrease in methylation through a ChIP assay of DNMT and XRE, and qRT-PCR of CYP1A1/1B1 will show that the reversal of estradiol mediated epigenetic CYP1A1 inhibition leads to upregulation of the estrogen detoxification pathway by Crosstalk Inhibitors in vitro. This premise will be taken to an ovariectomized rat model involving E2 and botanical compounds before sacrificing and analyzing uterine weight, performing LC-MS analysis of the E2 metabolites, and quantifying ERα expression levels using immunohistochemistry. This research may reveal mechanisms by which some bioactive women's health BDS compounds exhibit antiestrogenic outcomes, and the importance of the E2 detoxification pathway in promoting wellness in an in vivo postmenopausal model.