雌激素受体ER和甲状腺激素受体TR介导的典型环境内分泌干扰物效应研究

发布时间：2018-09-03 19:37

【摘要】：内分泌干扰物是近年来环境科学领域研究的热点之一。目前,我国还未建立有效的内分泌干扰物清单,也缺乏能够快速、有效地对化合物内分泌干扰效应进行高通量筛选的平台。本研究构建了以中国仓鼠卵巢癌细胞(CHO-K1)为宿主细胞,通过雌激素受体α(ERα)和甲状腺激素受体β(TRβ)介导的双荧光报告基因系统,并结合T-screen实验和计算毒理学技术,对3类化合物——限用杀虫剂锐劲特及其代谢产物、20种典型持久性有机污染物多氯联苯(PCBs)和9种新型工业替代品有机磷阻燃剂(OPFRs)的内分泌干扰效应进行了定性及定量评价。主要的研究工作和成果分为以下3部分内容:(1)锐劲特及其代谢产物的内分泌干扰效应评价:在双荧光报告基因体系中,锐劲特砜化物具有抗雌和抗甲活性,比其母本化合物(只有抗雌活性)表现出了更强的内分泌干扰效应。此外,通过分子对接后发现:在与TRβ的结合方面,锐劲特代谢产物(砜化物)比母本化合物具有更强的亲和力,即具有更强的抗甲状腺激素活性,这一点也有力地支持了前期细胞实验的结果。(2)20种PCBs的甲状腺激素效应评价:在检测的20种PCBs中,PCB44,PCB52,PCB101,PCB110和PCB180具有类甲状腺激素活性;而PCB99,PCB118,PCB128,PCB138,PCB163,PCB180,PCB187,PCB194和PCB199则具有抗甲状腺激素活性,其中PCB180既具有类甲活性也具有抗甲活性。此外,分子对接技术也对PCBs活性大小差异的潜在原因进行了探讨和解释。从构效层面分析,氯原子的取代个数或取代位置可能会影响PCBs与甲状腺受体TRβ间的相互作用。(3)9种OPFRs的甲状腺激素效应评价:通过TRβ介导的报告基因实验检测发现,9种OPFRs均无激动活性,而其中4种则表现出显著的抗甲状腺激素效应,活性大小顺序如下:TCPTBPTCPPTDCPP。这一结果在T-screen实验中得到了部分验证。此外,分子对接结果显示OPFRs甲状腺激素活性的强弱部分程度上取决于它和TRβ不同的结合方式,结构中含有苯环、卤素及短链的OPFRs可能更易表现出拮抗效应。
[Abstract]:Endocrine disruptors are one of the hotspots in the field of environmental science in recent years. At present, China has not established an effective list of endocrine disruptors, nor is there a platform for rapid and effective screening of endocrine disrupting effects of compounds. In this study, a double fluorescence reporter gene system, mediated by estrogen receptor 伪 (ER 伪) and thyroid hormone receptor 尾 (TR 尾), was constructed with Chinese hamster ovarian cancer cells (CHO-K1) as host cells. The system was combined with T-screen experiment and computational toxicology. In this paper, the endocrine disrupting effects of three kinds of chemical compounds, Reagin and its metabolite, 20 typical persistent organic pollutants (PCBs) (PCBs) and 9 new industrial substitutes, organophosphorus flame retardants (OPFRs), were evaluated qualitatively and quantitatively. The main research work and achievements are as follows: (1) Evaluation of endocrine disrupting effect of Trigonil and its metabolites: in the system of double fluorescence reporter gene, the sulfoxide has anti-female and anti-A activities. These compounds exhibit stronger endocrine disrupting effects than their female parent compounds (with only anti-female activity). In addition, after molecular docking, it was found that in binding to TR 尾, the sulfoxide metabolite of Fipronil had stronger affinity than the female compound, that is, it had stronger antithyroid hormone activity. This also strongly supports the results of previous cell experiments. (2) the evaluation of thyroid hormone effect of 20 PCBs: among the 20 PCBs tested, PCB44-PCB52PCB101 PCB110 and PCB180 have thyroid-like hormone activity, while PCB99,PCB118,PCB128,PCB138,PCB163,PCB180,PCB187,PCB194 and PCB199 have anti-thyroid hormone activity. Among them, PCB180 has both armour-like activity and anti-nail activity. In addition, molecular docking technique also discussed and explained the potential reasons for the difference of PCBs activity. From a structure-activity perspective, The number or position of substitution of chlorine atoms may affect the interaction between PCBs and thyroid receptor TR 尾. (3) the evaluation of thyroid hormone effect of nine OPFRs: through the TR 尾 mediated reporter gene assay, 9 OPFRs were found to be inactive. Four of them showed significant antithyroid hormone effects, and the order of activity was as follows: TCPTBPTPTDCPP. This result is partially verified in T-screen experiment. In addition, the molecular docking results showed that the activity of OPFRs thyroid hormone depended in part on its different binding mode with TR 尾, and that OPFRs with benzene ring, halogen and short chain might be more likely to exhibit antagonistic effect.
【学位授予单位】：浙江工业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：X503

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