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Basic investigation on influence of environmental chemicals on Sex-steroid hormone secretion

环境化学物质对性类固醇激素分泌影响的基础研究

基本信息

批准号：
16590095
负责人：
NAKAJIN Shizuo
金额：
$ 1.41万
依托单位：
Hoshi University School of Pharmacy and Pharmaceutical Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16590095/
关键词：
Environmental chemicals Sex-hormone Steroid Endocrine

项目摘要

The in vitro assay system to examine the influence of the environmental chemicals on the androgen production was constructed by using the primary culture of pig Leydig cell. We found that organotin compounds, such as tributyltin chloride (TBT), dibutyltin dichloride and triphenyltin chloride, strongly suppressed the testosterone production level at a concentration without cytotoxicity by using this assay system. It was clarified as action mechanism that TBT exposure suppressed the rise of intracellular cAMP level and decreased the CYP17 mRNA level up-regulated by human chorionic gonadotropin or forskolin (FSK). Furthermore, TBT inhibited 17β-HSD activity in vivo as well as in vitro, which enzyme was essential for testosterone production. The estrogen biosyntheses enzyme, aromatase is a rate-limiting enzyme of the estrogen biosyntheses, and an indispensable enzyme to the estrogen production. We clarified that aromatase was expressed in human adrenocortical carcinoma cell line, H295R and promoter II and 1.3 were selected for its expression, and the transcription was activated by EGF and PGE_2 stimulation. Then, the in vitro assay system to estimate aromatase activity or the transcriptional activity was constructed by using this H295R cell, and the influence of the environmental chemicals on the estrogen production was examined. Mono-(2-ethylhexyl) phthalate (MEHP), a main metabolite of di-(2-ethylhexyl) phthalate (DEHP) which is widely used as plasticizers for polyvinylchloride plastics suppressed aromatase activity in a dose-dependent manner and its transcription level, which are up-regulated with FSK. It was suggested as action mechanism that the suppression of aromatase activity and the transcription level by exposure of NCI-H295R cells to MEHP was regulated through the rapid and transient expression of orphan nuclear receptor Nur77 gene.

利用猪Leydig细胞原代培养物构建体外检测系统，检测环境化学物质对雄激素产生的影响。我们发现，通过使用该测定系统，有机锡化合物，例如三丁基氯化锡（TBT）、二丁基二氯化锡和三苯基氯化锡，在没有细胞毒性的浓度下强烈抑制睾酮的产生水平。其作用机制被阐明为TBT暴露抑制细胞内cAMP水平的升高并降低人绒毛膜促性腺激素或毛喉素(FSK)上调的CYP17 mRNA水平。此外，TBT 在体内和体外都能抑制 17β-HSD 活性，而该酶对于睾酮的产生至关重要。雌激素生物合成酶，芳香酶是雌激素生物合成的限速酶，是雌激素生成过程中不可缺少的酶。我们阐明了芳香化酶在人肾上腺皮质癌细胞系中表达，选择H295R和启动子II和1.3进行其表达，并通过EGF和PGE_2刺激激活转录。然后，利用该H295R细胞构建了评估芳香酶活性或转录活性的体外测定系统，并检查了环境化学物质对雌激素产生的影响。邻苯二甲酸单(2-乙基己基)酯(MEHP)是邻苯二甲酸二(2-乙基己基)酯(DEHP)的主要代谢物，广泛用作聚氯乙烯塑料的增塑剂，它以剂量依赖性方式抑制芳香酶活性及其转录水平，而FSK则上调芳香酶活性和转录水平。据认为，NCI-H295R细胞暴露于MEHP对芳香酶活性和转录水平的抑制是通过孤儿核受体Nur77基因的快速和瞬时表达来调节的作用机制。