乳腺癌上皮-间质转化后引发BCRP介导的多药耐药的研究

Multidrug resistance in breast cancer cells during epithelial-mesenchymal transition is modulated by breast cancer resistant protein

背景与目的:研究发现肿瘤的侵袭浸润和转移增强是上皮-间质转化(epithelial mesenchymal transition,EMT)所致,而EMT的发生也与肿瘤细胞多药耐药现象的发生密切相关。本研究通过分析乳腺癌组织和细胞中EMT与乳腺癌耐药蛋白表达的相关性,探讨EMT对乳腺癌中乳腺癌耐药蛋白(breast cancer resistant protein,BCRP)介导多药耐药的影响。方法:免疫组织化学方法检测乳腺癌组织中Snail、BCRP的表达;构建Snail真核表达载体pCDNA3.1-Snail,转染至MCF-7细胞,采用免疫荧光、Westernblot、Real-timePCR检测转录抑制因子snail、上皮标志物E-钙粘素、间质标志物vimentin以及多药耐药蛋白BCRP的表达;MTT法检测细胞对米托蒽醌的耐药指数。结果:免疫组化结果显示乳腺癌组织中Snail、BCRP的表达呈显著相关;免疫荧光、Westernblot、Real-timePCR显示与亲本MCF-7细胞相比,转染Snail后的MCF-7细胞中E-cadherin表达水平明显降低,而snail、vimentin和BCRP的表达水平明显增加;MTT结果显示细胞对米托蒽醌的耐药指数增加至9.93。结论:转染snail真核表达载体pCDNA3.1-snail可使乳腺癌MCF-7细胞发生EMT,并导致细胞中BCRP表达增加,引发BCRP介导的多药耐药。

Background and Objective： Epithelial-mesenchymal transition （EMT） not only initiates invasion and metastasis of tumors, but also induces multidrug resistance in tumor cells. Our experiment analyzed the dependability between breast cancer resistant protein （BCRP） and EMT in breast cancer to explore the effect of EMT on BCRP-mediated multidrug resistance. Methods： The expressions of BCRP and transcription inhibitor Snail （Snail） in breast cancer were detected by immunohistochemistry. The eukaryotic expression vector pCDNA3.1-Snail was constructed and then transfected into human breast cancer cell line MCF-7. Snail, epithelial marker gene E-cadherin, interstitial marker gene Vimentin, multidrug resistance protein BCRP, and relative drug resistance were measured by immunofluorescence, Western blot, real-time polymerase chain reaction （PCR）, and MTT assay. Results：Immunohistochemistry showed that Snail was highly correlated with BCRP in breast cancer. Immunofiuorescence, Western blot, and real-time PCR revealed that compared with parent cell MCF-7, after transfected with Snail, the expression of E-cadherin in MCF-7 decreased, but Snail, Vimentin, and BCRP increased. MTT displayed that the relative drug resistance increased to 9.93. Conclusion. After transfected with eukaryotic expression vector pCDNA3.1- Snail, breast cancer cells MCF-7 showed EMT with BCRP-mediated multidrug resistance.