壬基酚通过抑制ESR2表达激活ERK通路促进结直肠癌细胞增殖的作用机制  

Mechanism of nonylphenol promoting colorectal cancer cell proliferation by activating ERK signaling pathway via inhibiting ESR2 expression

作　　者：曲颜 张念杰 尹硕 王彪 杨雪峰 Qu Yan;Zhang Nianjie;Yin Shuo;Wang Biao;Yang Xuefeng(Department of Gastrointestinal Surgery,the Second Affiliated Hospital of Zunyi Medical University,Zunyi Guizhou 563006,China;Department of Gastrointestinal Surgery,Affiliated Hospital of Zunyi Medical University,Zunyi Guizhou 563000,China)

机构地区：[1]遵义医科大学第二附属医院胃肠外科,贵州遵义563006 [2]遵义医科大学附属医院胃肠外科,贵州遵义563000

出　　处：《遵义医科大学学报》2024年第9期850-856,867,共8页Journal of Zunyi Medical University

基　　金：遵义医科大学2017年度学术新苗培养及创新探索专项项目[NO:黔科合-平台人才(2017)5733-081];遵义医科大学第二附属医院硕士科研启动基金项目(NO:SQ-2021-15,SQ-2021-14)。

摘　　要：目的探讨壬基酚环境内分泌干扰物对结直肠细胞增殖的影响及其与ERK通路激活的关系。方法通过癌症基因组图谱及HPA数据库分析ERβ在CRC组织中的表达。不同浓度NP干预的COLO205细胞后,实时荧光定量聚合酶链反应(qRT-PCR)检测细胞中ESR2的表达,蛋白印迹(Western blot)检测细胞中ERβ的表达。细胞分为空白对照组、ESR2干扰对照组、壬基酚(10^(-6)mol/L)组、ESR2干扰组、壬基酚+ESR2干扰组。利用特异性小干扰RNA片段(si-ESR2)抑制ESR2的表达,NP(10^(-6)mol/L)干预COLO205细胞24 h后,通过CCK-8、流式细胞术检测COLO205细胞增殖及凋亡水平变化,Western blot检测凋亡相关蛋白(Bad、Bcl-2、Cleaved caspase-3)表达及ERK通路激活情况。结果TCGA及HPA分析发现,CRC组织中ERβ的表达均显著低于正常组织(P<0.001)。构建干扰载体对细胞进行转染,si-ESR2-3#的干扰效率超过60%,si-NC对照组无显著差异。NP干预后,可显著抑制COLO205细胞中ESR2及ERβ的表达(F=27.791,P<0.001),与空白对照组比较,NP(10^(-6)mol/L)显著提高细胞增殖能力(F=107.3,P<0.001),抑制细胞凋亡(F=51.1,P=0.0068),降低Bad(F=46.56,P=0.0032)、Cleavedcaspase-3(F=18.04,P=0.0035)的表达,促进Bcl-2(F=32.58,P=0.0054)的表达,p-ERK的表达被显著增加(F=39.07,P=0.0011);与NP组比较,NP联合si-ESR2组细胞增殖能力增强(F=107.3,P<0.001)、凋亡率显著降低(F=51.1,P=0.0003),Bad(F=46.56,P=0.0009)与Cleavedcaspase-3(F=18.04,P=0.0019)的表达显著降低,Bcl-2(F=32.58,P=0.0019)与p-ERK的表达显著增加(F=39.07,P=0.0081)。结论壬基酚可以通过ESR2激活ERK通路促进CRC细胞增殖。Objective To investigate the effect of Xenoestrogen Nonylphenol(NP)on the proliferation of colorectal cancer cells and its relationship with the activation of ERK pathway.Methods The expression of ERβin colorectal cancer cells was analyzed by The Cancer Genome Atlas(TCGA)and Human Protein Atlas(HPA).The expression of ESR2 in COLO205 cells was detected by quantitative fluorescent polymerase chain reaction(qRT-PCR)after treatment with different concentrations of NP.Western blot was used to analyze the effect of NP on ERβexpression in COLO205 cells.Cells were divided into Control group,si-NC group,NP(10^(-6)mol/L)group,si-ESR2 group,and NP+si-ESR2 group.The expression of ESR2 was inhibited by using a small interfering RNA fragment(si-ESR2).After NP(10^(-6)mol/L)was applied to COLO205 cells for 24 h,the proliferation and apoptosis of COLO205 cells were detected by CCK-8 and flow cytometry.The expression of apoptosis-related proteins(Bad,Bcl-2,and Cleaved caspase-3)and activation of ERK pathway were detected by Western blot.Results TCGA and HPA analysis showed that the expression of ERβin CRC tissues was significantly lower than that in normal tissues(P<0.001).The interference vector was constructed and transfected into the cells.The interference efficiency of si-ESR2-3#was more than 60%,and there was no significant difference in the si-NC control group.NP intervention significantly inhibited the expression of ESR2 and ERβin COLO205 cells(F=27.791,<0.001),and NP(10^(-6)mol/L)significantly increased cell proliferation(F=107.3,P<0.001)and inhibited cell apoptosis(F=51.1,P=0.0068)compared with the Control group.The expression of Bad(F=46.56,P=0.0032)and Cleaved caspase-3(F=18.04,P=0.0035)were decreased,the expression of Bcl-2 was increased(F=32.58,P=0.0054),and the expression of p-ERK was significantly increased(F=39.07,P=0.0011).Compared with NP group,the viability and proliferation of cells in NP combined with si-ESR2 group were increased(F=107.3,P<0.001),the apoptosis rate was significantly decreased(F=51.1,P=0.00

关 键 词：结直肠癌 壬基酚 ESR2 ERK通路 细胞增殖 

分 类 号：R735.35[医药卫生—肿瘤]