微RNA-200c通过EZH2/E-cad信号通路抑制肺癌A549/MTX耐药细胞的迁移和侵袭能力  

MicroRNA-200c depresses the migration and invasion of methotrexateresistant lung cancer A549/MTX cells through EZH2/E-cad signaling pathway

作　　者：张晓蕾[1] 邓芳[1] 单武林 杨臣欢 魏敏[1] 李程[1] 吴坤[1] 韩丹丹[1] 张婧[1] 李明[1] 

机构地区：[1]安徽省肿瘤医院检验科,安徽合肥230001

出　　处：《肿瘤》2015年第10期1076-1082,共7页Tumor

基　　金：安徽省自然科学基金资助项目(编号:1308085QH144)~~

摘　　要：目的 :探讨微RNA-200c(micro RNA-200c,mi R-200c)对耐受甲氨蝶呤(methotrexate,MTX)的人非小细胞肺癌A549细胞迁移和侵袭能力的影响,及其可能的分子作用机制。方法 :通过浓度递增结合低剂量持续诱导,获得耐受MTX的人肺癌A549/MTX细胞系后,观察诱导前后细胞的形态学改变。将mi R-200c模拟物(mimic)转染A549/MTX细胞株后,分别采用细胞划痕愈合实验和Transwell细胞迁移和侵袭实验检测细胞的迁移和侵袭能力;再用实时荧光定量PCR法检测mi R-200c过表达的A549/MTX细胞中人Zeste基因增强子同源物2(human enhancer of Zeste homolog 2,EZH2)和E-钙黏蛋白(E-cadherin,E-cad)的m RNA表达水平。结果 :肺癌A549/MTX耐药细胞构建成功。miR-200c minic转染后A549/MTX耐药细胞表达mi R-200c水平比转染阴性片段组高6.41倍(P<0.05),表明转染成功。mi R-200c mimic转染后A549/MTX细胞的迁移和侵袭能力显著降低(P值均<0.05);而且A549/MTX细胞中EZH2 m RNA表达水平明显降低,而E-cad m RNA水平明显升高(P值均<0.05)。结论 :mi R-200c高表达可以抑制A549/MTX耐药细胞的迁移和侵袭能力,其机制可能与其下调EZH2表达和上调E-cad水平有关。Objective： To investigate the effects of microRNA-200c （miR-200c） on migration and invasion of methotrexate （MTX）-resistant human non-small cell lung cancer A549 cells and its possible molecular mechanism. Methods： MTX-resistant lung cancer A549/MTX cells were obtained by treatment with dose- escalated and continuous low-dose MTX. The change of morphology of A549/MTX cells was observed. After miR-200c mimic was transfected into A549/MTX cells, the cell invasion and migration abilities were examined by Transwell cell invasion and migration assay and scratch wound healing assay, respectively. The mRNA expression levels of human enhancer of Zeste homolog 2 （EZH2） and E-cadherin （E-cad） in miR-200c-overexprssed A549/MTX cells were detected by real-time fluorescent quantitative-PCR. Results： The lung cancer A549/MTX cells were successfully established. The expression level of miR-200c in A549/MTX cells after transfection with miR-200c mimic was 6.41 times higher than that of the negative fragment-transfected A549/MTX cells （P 〈 0.05）, which indicated that the transfection was successful. The migration and invasion abilities of A549/ MTX cells were significantly reduced after miR-200c mimic transfection （both P 〈 0.05）. In addition, the expression level of EZH2 mRNA in A549/MTX cells was significantly reduced after miR-200c mimic transfection, but the expression of E-cadherin mRNA was significantly increased （both P 〈 0.05）. Conclusion： Overexpression of miR-200c can inhibit the migration and invasion of A549/ MTX cells. The mechanism may be related to down-regulation of EZH2 expression and up- regulation of E-cad level.

关 键 词：癌 非小细胞肺 甲氨蝶呤 微RNAs 基因表达 细胞迁移 测定 

分 类 号：R734.202[医药卫生—肿瘤]