ESTROGEN RECEPTOR ALPHA AND G-PROTEIN COUPLED RECEPTOR 30 MEDIATE THE NEUROPROTECTIVE EFFECTS OF 17β-ESTRADIOL IN NOVEL MURINE HIPPOCAMPAL CELL MODELS
ESTROGEN RECEPTOR ALPHA AND G-PROTEIN COUPLED RECEPTOR 30 MEDIATE THE NEUROPROTECTIVE EFFECTS OF 17β-ESTRADIOL IN NOVEL MURINE HIPPOCAMPAL CELL MODELS
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DOI：
10.1016/j.neuroscience.2010.06.076
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发表时间：
2010-09-29
期刊：
影响因子：
3.3
通讯作者：
Belsham, D. D.
中科院分区：
文献类型：
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作者：
Gingerich, S.;Kim, G. L.;Belsham, D. D.
海马体是一个多方面的，复杂的大脑结构，被认为是学习中心。原代海马细胞培养的使用揭示了参与整体生理功能的重要细胞机制。然而，原代培养的使用本质上是困难的，并且显然缺乏来自小鼠海马的永生化细胞系用于分子水平的机制研究。我们有永生化细胞系从胚胎（E18）和成人衍生的海马原代细胞培养使用逆转录病毒感染的SV 40 T抗原。4个克隆胚胎系mHippoE-2、mHippoE-5、mHippoE-14、mHippoE-18和1个混合成体系mHippoA-mix表现出神经元形态，具有神经突延伸和神经元标记物表达，具有独特的基因表达谱。我们使用这些细胞模型来研究17 β-雌二醇（E2）对谷氨酸诱导的神经毒性的神经保护作用。这些细胞系表达一系列相关的基因和受体，这些基因和受体被认为在神经保护中发挥作用，包括雌激素受体ER α、ER β和GPR 30。我们发现用E2（10或100 nM）预处理24 h显著降低了由谷氨酸mHippoE-14和mHippoE-18细胞诱导的细胞死亡，但不是mHippoA混合物。使用特异性雌激素受体（ER）激动剂4，4 '，4”-（4-丙基[1H]-吡唑-1，3，5-三基）三酚（PPT）和二芳基丙腈2，3-双（4-羟基苯基）丙腈（DPN）预处理24 h，我们将E2介导的神经保护作用与ER α联系起来，但仅在mHippoE-18细胞中。由于E2激活PI 3 K/Akt和STAT 3信号通路，我们还测试了膜结合E2受体GPR 30是否参与其神经保护作用。用GPR 30激动剂G-1（10和100 nM）预处理1小时，但不预处理24小时，可显著减弱mHippoE-14和mHippoE-18细胞的细胞死亡。使用特异性ER拮抗剂ICI 182780和GPR 30拮抗剂G-15将这些作用与ER和GPR 30受体联系起来。这是GPR 30可能在雌激素对海马神经元的保护作用中发挥作用的第一个证据。(C)2010年IBRO。由爱思唯尔有限公司出版。保留所有权利。
The hippocampus is a multifaceted, complex brain structure considered to be the learning center. The use of primary hippocampal cell cultures has uncovered important cellular mechanisms involved in overall physiological function. Yet, the use of primary culture is inherently difficult, and the lack of immortalized cell lines from the murine hippocampus for mechanistic studies at the molecular level is evident. We have immortalized cell lines from embryonic (E18) and adult-derived hippocampal primary cell culture using retroviral infection of SV40 T-antigen. Four clonal embryonic lines, mHippoE-2, mHippoE-5, mHippoE-14, mHippoE-18, and one mixed adult line, mHippoA-mix, exhibited neuronal morphologies with neurite extensions and expression of neuronal markers, with unique gene expression profiles. We used these cell models to study the neuroprotective effects of 17 beta-estradiol (E2) on glutamate-induced neurotoxicity. The cell lines express a relevant array of genes and receptors suggested to play a role in neuroprotection, including estrogen receptors ER alpha, ER beta, and GPR30. We find that pretreatment with E2 (10 or 100 nM) for 24 h significantly reduced cell death induced by glutamate mHippoE-14 and mHippoE-18 cells, but not the mHippoA-mix. Using 24 h pretreatment with the specific estrogen receptor (ER) agonists, 4,4',4"-(4-propyl[1H]-pyrazole-1,3,5-triyl)trisphenol (PPT) and diarylpropionitrile, 2,3-bis(4-Hydroxyphenyl)-propionitrile (DPN), we linked the E2-mediated neuroprotection to ER alpha, but only in the mHippoE-18 cells. Since E2 activated both PI3K/Akt and STAT3 signaling pathways, we also tested whether the membrane-bound E2 receptor GPR30 was involved in its neuroprotective action. Pretreatment with the GPR30 agonist G-1 (10 and 100 nM) for 1 h, but not 24 h, significantly attenuated cell death in both mHippoE-14 and mHippoE-18 cells. The use of specific ER antagonist ICI 182780 and GPR30 antagonist G-15 linked these effects to both ER and GPR30 receptors. This is the first evidence that GPR30 may play a role in the protective effects of estrogen in hippocampal neurons. (C) 2010 IBRO. Published by Elsevier Ltd. All rights reserved.