低浓度TBBPA长期暴露对乳腺癌细胞增殖效应研究  

Effects of Chronic Exposure to Low-concentration Tetrabromobisphenol A on the Proliferation of Breast Cancer Cells

作　　者：庞晶 贾斌 朱慧瑞 姚杰[1] 弓韬[1] 刘志贞[1] 李卓玉 PANG Jing;JIA Bin;ZHU Huirui;YAO Jie;GONG Tao;LIU Zhizhen;LI Zhuoyu(Department of Biochemistry and Molecular Biology,Basic Medical College,Shanxi Medical University,Taiyuan 030001,China;Institute of Biotechnology,Key Laboratory of Chemical Biology and Molecular Engineering of,Ministry of Education,Shanxi University,Taiyuan 030006,China)

机构地区：[1]山西医科大学生物化学与分子生物学教研室,山西太原030001 [2]山西大学生物技术研究所,化学生物学与分子工程教育部重点实验室,山西太原030006

出　　处：《山西大学学报（自然科学版）》2024年第3期647-656,共10页Journal of Shanxi University(Natural Science Edition)

基　　金：山西省自然科学基金(202203021211232);山西赛奥生物科技有限公司与山西医科大学合作项目(2F022018020)。

摘　　要：四溴双酚A(Tetrabromobisphenol A,TBBPA)是生产量最高的反应型和添加型溴代阻燃剂,具有持久性有机污染物的特征,可通过多种途径对人体进行暴露。关于TBBPA暴露与人体健康风险等问题亟待研究,其中TBBPA作为一种潜在的环境雌激素干扰物,对于雌激素敏感性乳腺癌细胞的促癌机制尚不明确。因此,在本研究中将人源乳腺癌细胞MCF-7(ER+)细胞系和MDA-MB-231(ER-)细胞系分别连续5代暴露于环境浓度下人体血浆中残留的TBBPA(10^(-8)mol/L)以模拟长期暴露模式,观察细胞的增殖效应并探讨可能的作用机制。CCK8和集落形成实验的结果表明,10^(-8)mol/L TBBPA长期暴露可以显著促进MCF-7细胞增殖。活性氧(ROS)生成实验表明,MCF-7细胞在10^(-8)mol/L TBBPA长期暴露下ROS水平升高。蛋白印迹结果表明,10^(-8)mol/L TBBPA长期暴露于MCF-7细胞后,上调了Ras信号通路中Cyclin D1、c-Myc、p-ERK1/2和Ras的蛋白表达水平。Ras蛋白抑制剂(FTS)干预后阻断了Ras/p-ERK1/2/Cyclin D1/c-Myc并降低了细胞的增殖效应。此外,CCK8实验结果表明ERα的抑制剂Fulvestrant预处理MCF-7细胞可减弱TBBPA诱导的细胞增殖活性,证明TBBPA促进MCF-7细胞增殖与ER的存在有直接关联。同时选取ER-的MDA-MB-231细胞株进行了研究,在相同浓度的TBBPA长期暴露下,其增殖活性、ROS水平及Ras信号通路相关蛋白均无显著影响。研究结果表明10^(-8)mol/L TBBPA长期暴露可以通过激活Ras信号通路促进雌激素受体阳性的MCF-7细胞的增殖。Tetrabromobisphenol A(TBBPA)is the reactive and additive brominated flame retardants(BFRs)with the highest production volume,which has the characteristics of persistent organic pollutants and can be exposed to human bodies in various ways.Thus,the relationship between TBBPA exposure and human health risks needs to be studied urgently.In particular,as a potential environmental estrogen disruptor,the carcinogenic mechanism of TBBPA on estrogen-sensitive breast cancer cells remains unclear.Therefore,in this study,human MCF-7(ER-positive)cell lines and MDA-MB-231(ER-negative)cell lines were continuously treated with the actual plasma concentration of TBBPA(10-8mol/L)for five generations to simulate long-term exposure mode.The proliferation effect and the possible mechanism were examined.The data from CCK8 and colony formation experiments demonstrated that 10^(-8) mol/L TBBPA long-term exposure promoted the proliferation of MCF-7 cells.The content of ROS increased in MCF-7 cells after long-term exposure to 10-8 mol/L TBBPA.Western blotting results showed that 10^(-8) mol/L TBBPA upregulated the Ras expression and induced p-ERK1/2,c-Myc,and Cyclin DI expression in MCF-7 cells on long-term exposure.Moreover,the intervention of Ras inhibitor FTS blocked the Ras/p-ERK1/2/Cyclin D1/c-Myc and inhibited cell proliferation.In addition,the data from CCK8 showed that pretreatment of MCF-7 cells by Fulvestrant,an ERa inhibitor,could attenuate cell proliferation activity induced by TBBPA,suggesting that the promotion of MCF-7 cell proliferation by TBBPA was directly related to the ER.At the same time,the MDA-MB-231(ER-negative)cell line was selected for research.Under long-term exposure to the same concentration of TBBPA,its proliferation activity,ROS level,and Ras signaling pathway related proteins were not significantly affected.These results show that long-term exposure to 10^(-8) mol/L TBBPA can accelerate MCF-7(ER-positive)cell proliferation via the activation of the Ras signaling pathway.

关 键 词：TBBPA 乳腺癌 Ras信号通路 增殖效应 

分 类 号：R994.6[医药卫生—毒理学]