Membrane estrogen receptor-alpha-mediated nongenomic actions of phytoestrogens in GH3/B6/F10 pituitary tumor cells.
Membrane estrogen receptor-alpha-mediated nongenomic actions of phytoestrogens in GH3/B6/F10 pituitary tumor cells.
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DOI：
10.1186/1750-2187-4-2
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发表时间：
2009-04-28
影响因子：
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通讯作者：
Watson CS
中科院分区：
文献类型：
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作者：
Jeng YJ;Kochukov MY;Watson CS
雌激素（E2）通过几种雌激素受体（ER）介导来自质膜的各种细胞内信号级联。垂体是一个雌激素敏感的组织，我们曾报道E2可激活膜ERα（mERα）富集的GH 3/B6/F10大鼠垂体瘤细胞系中的丝裂原活化蛋白激酶（MAPK），如ERK 1/2和JNK 1/2/3。植物雌激素是在植物和食物中发现的化合物，如大豆，苜蓿芽和红葡萄。它们在结构上与E2相似，并通过与ER结合而具有相似的作用机制。植物雌激素以低得多的亲和力与核ER结合，因此在介导基因组应答方面不太有效。然而，关于它们通过mER介导非基因组效应的能力知之甚少。为了研究不同的非基因组途径的激活，并确定mERα的参与，我们通过放射免疫测定法测量了催乳素（PRL）的释放，通过定量平板免疫测定法测量了MAPK激活（ERK 1/2和JNK 1/2/3），并通过Fura-2荧光成像测量了E2或四种不同植物雌激素（香豆雌酚，大豆黄酮，染料木素和反式白藜芦醇）处理的细胞内[Ca 2 +]。在GH 3/B6/F10细胞中，香豆甾醇和大豆苷元类似于E2增加PRL释放，而染料木黄酮和反式白藜芦醇没有影响。除染料木黄酮外，所有这些化合物在1-10皮摩尔浓度下激活ERK 1/2信号传导; JNK 1/2/3在100纳摩尔浓度下被所有化合物激活。所有化合物也引起快速的钙摄取，虽然在独特的剂量依赖性的钙响应模式的几个方面的这种响应。表达低水平mERα的GH 3细胞亚克隆（GH 3/B6/D9）对任何植物雌激素处理均无反应，表明这些非基因组效应是通过mERα介导的。植物雌激素通过mERα介导这些非基因组反应（激活MAPK、PRL释放和增加细胞内[Ca 2 +]）的效力远高于先前报道的基因组反应。由E2和所有测试的植物雌激素引起的独特的非单调剂量反应和变体信号模式表明，可能涉及复杂和多个信号通路或结合伴侣。通过激活这些不同的非基因组信号通路，植物雌激素可能对垂体细胞功能产生重要的生理后果。
Estradiol (E2) mediates various intracellular signaling cascades from the plasma membrane via several estrogen receptors (ERs). The pituitary is an estrogen-responsive tissue, and we have previously reported that E2 can activate mitogen-activated protein kinases (MAPKs) such as ERK1/2 and JNK1/2/3 in the membrane ERα (mERα)-enriched GH3/B6/F10 rat pituitary tumor cell line. Phytoestrogens are compounds found in plants and foods such as soybeans, alfalfa sprouts, and red grapes. They are structurally similar to E2 and share a similar mechanism of action through their binding to ERs. Phytoestrogens bind to nuclear ERs with a much lower affinity and therefore are less potent in mediating genomic responses. However, little is known about their ability to act via mERs to mediate nongenomic effects. To investigate the activation of different nongenomic pathways, and determine the involvement of mERα, we measured prolactin (PRL) release by radio-immunoassay, MAPK activations (ERK1/2 and JNK1/2/3) via a quantitative plate immunoassay, and intracellular [Ca2+] by Fura-2 fluorescence imaging in cells treated with E2 or four different phytoestrogens (coumestrol, daidzein, genistein, and trans-resveratrol). Coumesterol and daidzein increased PRL release similar to E2 in GH3/B6/F10 cells, while genistein and trans-resveratrol had no effect. All of these compounds except genistein activated ERK1/2 signaling at 1–10 picomolar concentrations; JNK 1/2/3 was activated by all compounds at a 100 nanomolar concentration. All compounds also caused rapid Ca2+ uptake, though in unique dose-dependent Ca2+ response patterns for several aspects of this response. A subclone of GH3 cells expressing low levels of mERα (GH3/B6/D9) did not respond to any phytoestrogen treatments for any of these responses, suggesting that these nongenomic effects were mediated via mERα. Phytoestrogens were much more potent in mediating these nongenomic responses (activation of MAPKs, PRL release, and increased intracellular [Ca2+]) via mERα than was previously reported for genomic responses. The unique non-monotonic dose responses and variant signaling patterns caused by E2 and all tested phytoestrogens suggest that complex and multiple signaling pathways or binding partners could be involved. By activating these different nongenomic signaling pathways, phytoestrogens could have significant physiological consequences for pituitary cell functions.