摘要
目的:研究整合素β1(integrinβ1,ITGB1)在人乳腺癌MCF-7细胞过表达对其迁移侵袭能力及三苯氧胺(tamoxifen, TAM)耐药的影响。方法:构建携带ITGB1的质粒(pBABE-puro-myc-ITGB1),通过包装反转录病毒转染人乳腺癌MCF-7细胞系,构建稳定高表达ITGB1的细胞株MCF-7-ITGB1;实验设置MCF-7组(对照组)、MCF-7-vector组(阴性对照组)和MCF-7-ITGB1(实验组);采用qRT-PCR和Western blot技术分别检测各组ITGB1 mRNA和蛋白表达量;MTT法检测不同TAM浓度下细胞增殖活性;Transwell实验检测不同ITGB1表达情况下细胞迁移及侵袭能力;Western blot检测上皮间质转换(epithelial-mesenchymal transition, EMT)标志物E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、间质表型标志物波形蛋白(vimentin)、纤维黏连蛋白(fibronectin)等相关蛋白的表达情况。结果:在人乳腺癌细胞MCF-7中,成功构建稳定高表达ITGB1细胞系;MCF-7及MCF-7-ITGB1的TAM半数抑制浓度(half-maximal inhibitory concentrations, IC_(50))分别为1.24 nmol/L和20.28 nmol/L;MCF-7-ITGB1的耐药指数为16.35。MCF-7-ITGB1较MCF-7及MCF-7-vector细胞迁移及侵袭能力显著增强,差异有统计学意义(P<0.05)。MCF-7-ITGB1的EMT标志物E-cadherin显著下调(P<0.05),fibronectin、N-cadherin、vimentin蛋白显著上调(P<0.05)。结论:稳定过表达ITGB1促进乳腺癌MCF-7细胞迁移侵袭能力增强及三苯氧胺的耐药,其作用机制可能与ITGB1活化上皮间质转换通路诱导细胞EMT相关。
Objective:To explore the effects of over-expressed integrin β1(ITGβ1)on invasion,migration and tamoxifen(TAM)resistance of breast cancer cell line MCF-7.Methods:Recombinant plasmid pBABE-puro-myc-ITGB1 was formed,and retrovirus was packaged.MCF-7 cells were then transfected with the retrovirus carrying ITGB1 gene or the controls.Blank group(MCF-7),negative control group(MCF-7-vector),the ITGB1 over-expressed group(MCF-7-ITGB1)were set up.After treatment with puromycin,the expression of ITGB1 in MCF-7 cells was determined by quantitative real-time PCR and Western blot assays.The growth rates of cells at different TAM concentrations were detected by MTT method.The abilities of cancer cells to migrate and invade were analyzed by Tr answell assays.The expressions of epithelial-mesenchymal transition(EMT)markers(E-cadherin,vimentin,fibronectin and N-cadherin)in MCF-7 cells were detected by Western blot.Results:Stably over-expressed ITGB1 cell line(MCF-7-IT-GB1)was successfully constructed in human breast cancer cell line MCF-7.The half-maximal inhibitory concentrations(IC_(50))of TAM for MCF-7 and MCF-7-ITGB1 were 1.24 nmol/L and 20.28nmol/L,respectively.The resistance index of MCF-7-ITGB1 was 16.35.Compared to MCF-7 and MCF-7-vector cells,MCF-7-ITGB1 cells have better ability to migrate and invade(P<0.05);the E-cadherin expression in the former was lower than that in the latter(P<0.05);and vimen-tin,fibronectin and N-cadherin expressions in the former were higher than that in latter(P<0.05).Conclusion:ITGB1 promotes TAM resistance and enhances the ability of cells to migrate and invade.Its possible mechanism may be related to EMT in breast cancer cell line MCF-7.
作者
杨钱
陈振海
胡淞
付伟杰
Yang Qian;Chen Zhenhai;Hu Song;Fu Weijie(Department of General Surgery,Chongqing University Central Hospital(i.e.Chongqing Emergency Medical Center,or the Fourth People’s Hospital of Chongqing),Chongqing 400000,China)
出处
《肿瘤预防与治疗》
2022年第6期500-505,共6页
Journal of Cancer Control And Treatment
基金
重庆市自然科学基金面上项目(编号:cstc 2021jcyj-msxmX1031)
