敲低雌激素受体α基因抑制A549细胞的增殖、迁移及裸鼠移植瘤的形成  

Knockdown of estrogen receptor alpha inhibits the proliferation and migration of A549cells and the formation of transplanted tumors in nude mice

作　　者：陈沛锐 吕静[1] 曾春芳[2] 何小艳[3] 刁明强[1] CHEN Peirui;LV Jing;ZENG Chunfang;HE Xiaoyan;DIAO Mingqiang(Department of Cardiothoracic Surgery,Deyang People's Hospital,Deyang,Sichuan 618000,P.R.China;Department of Respiratory Medicine,Deyang People's Hospital,Deyang,Sichuan 618000,P.R.China;Department of Pathology,Deyang People's Hospital,Deyang,Sichuan 618000,P.R.China)

机构地区：[1]德阳市人民医院心胸外科,四川德阳618000 [2]德阳市人民医院呼吸内科,四川德阳618000 [3]德阳市人民医院病理科,四川德阳618000

出　　处：《中国呼吸与危重监护杂志》2021年第9期649-655,共7页Chinese Journal of Respiratory and Critical Care Medicine

基　　金：四川省科技厅科技计划项目(2016SZ0049)。

摘　　要：目的探究干扰RNA(sh RNA)敲低非小细胞肺癌(NSCLC)A549细胞雌激素受体α(ERα)基因后对A549细胞生物学活性及裸鼠肿瘤生长的影响。方法采用sh RNA敲低A549细胞中的ERα基因,逆转录-聚合酶链反应(RT-PCR)及蛋白免疫印迹法(Western blot)检测敲低后的基因表达与蛋白表达,克隆形成实验检测细胞的增殖能力,RT-PCR检测相关增殖蛋白Ki-67和PCNA的表达,流式细胞检测细胞凋亡率;Transwell实验检测细胞的侵袭能力,Western blot检测上皮型钙黏蛋白(E-cad)和神经型钙黏蛋白(N-cad)的表达。对照组和转染ERα-sh RNA1的A549细胞分别注射到裸鼠皮下,构建移植瘤,免疫组织化学检测肿瘤组织中Ki-67与N-cad的表达。结果与对照组相比,转染ERα-sh RNA1和ERα-sh RNA2后,ERα的m RNA和蛋白表达量均明显下调(均P<0.05),采用干扰效率高的sh RNA1进行后续实验。A549细胞转染ERα-sh RNA1后,集落形成率较对照组显著下调(P<0.05),细胞凋亡比率明显升高(P<0.05),Ki-67和PCNA的表达显著下调(P<0.05),侵袭细胞数目明显减少,E-cad表达升高,N-cad表达降低(均P<0.05)。裸鼠成瘤实验结果表明干扰ERα的表达可显著抑制肿瘤的生长(P<0.05),下调Ki-67与N-cad的阳性细胞比率(均P<0.05)。结论敲低ERα抑制了A549细胞的生长、运动能力及裸鼠移植瘤的发生和发展。Objective To explore the effect of interfering RNA(sh RNA)on biological activity of A549 cells and tumor growth in nude mice after knockdown of estrogen receptorα(ERα)gene.Methods The ERαgene in A549 cells was knocked down by sh RNA.RT-PCR and Western blot were used to detect the gene expression and protein expression after knockdown;colony formation experiment was used to detect the proliferation of cells,and RT-PCR was used to detect the expression of Ki-67 and PCNA;flow cytometry was used to detect apoptosis rate;transwell assay was used to detect cell invasion ability;Western blot was used to detect the expression of epithelial cadherin(E-cad)and neuropathic cadherin(N-cad)protein.The control group and A549 cells transfected with ERα-sh RNA1 were injected subcutaneously in nude mice to construct transplanted tumors.Immunohistochemistry was used to detect the expression of Ki-67 and N-cad in tumor tissues.Results Compared with the control group,after transfection of ERα-sh RNA1 and ERα-sh RNA2,the m RNA and protein expressions of ERαwere reduced significantly(P<0.05),and sh RNA1 with high interference efficiency was used for subsequent experiments.Compared with the control group,the A549 cells were transfected with ERα-sh RNA1,the colony formation rate was down-regulated significantly(P<0.05),the apoptosis rate was increased significantly(P<0.05),the expression of Ki-67 and PCNA were down-regulated significantly(P<0.05),the number of invasive cells was reduced significantly,the expression of E-cad was increased,and the expression of N-cad was decreased(P<0.05).The results of tumor formation in nude mice showed that interfering with ERαexpression can significantly inhibit tumor growth(P<0.05),and down-regulate the rate of Ki-67 and N-cad positive cells(P<0.05).Conclusion Knockdown of ERαinhibits the proliferation and migration ability of NSCLC cells and the occurrence and development of transplanted tumors in nude mice.

关 键 词：雌激素受体Α A549细胞 细胞生长 侵袭 肿瘤形成 

分 类 号：R734.2[医药卫生—肿瘤]