miR-4295靶向结合PTEN促进EMT和鼻咽癌细胞的侵袭  

miR-4295 promotes epithelial to mesenchymal transition and human nasopharyngeal carcinoma invasion by directly targeting

作　　者：肖烈钢[1] 黄季萌[2] 何本夫[3] 朱成全[4] PTEN Xiao Liegang;Huang Jimeng;He Benfu;Zhu Chengquan(Combined Chinese and Western Medicine Department,421 Hospital of PLA,Guangzhou 510318,China;Medical Department,421 Hospital of PLA,Guangzhou 510318,China;Oncology Department,421 Hospital of PLA,Guangzhou 510318,China;Outpatient Department,421 Hospital of PLA,Guangzhou 510318,China)

机构地区：[1]中国人民解放军第四二一医院中西医结合科,广州510318 [2]中国人民解放军第四二一医院医务处,广州510318 [3]中国人民解放军第四二一医院肿瘤科,广州510318 [4]中国人民解放军第四二一医院门诊部,广州510318

出　　处：《国际医药卫生导报》2018年第19期2934-2940,共7页International Medicine and Health Guidance News

基　　金：广州市科技计划项目（201707010025）

摘　　要：目的 探讨miR-4295在鼻咽癌细胞侵袭、转移过程中的作用及相关的分子机制.方法 首先通过PCR检测鼻咽癌细胞系中miR-4295的表达,通过用重组逆转录病毒介导建立稳定高表达miR-4295的鼻咽癌细胞株5-8F/miR-4295.分别应用Transwell侵袭实验、划痕损伤修复实验及三维培养实验检测过表达miR-4295后细胞株5-8F的迁移、侵袭能力;WB和QPCR方法检测E-Cadherin、Fibronectin、Vimentin在5-8F/miR-4295及5-8F/vector细胞中蛋白及miRNA的表达水平;荧光素酶活性检测miR-4295与PTEN靶向结合情况.结果 miR-4295在鼻咽癌细胞系中高表达,通过Transwell侵袭实验、划痕损伤修复实验、三维培养实验均显示过表达miR-4295后5-8F细胞的体外迁移及侵袭能力明显增强;WB及QPCR结果显示：过表达miR-4295后,5-8F细胞上皮细胞分子标志物E-Cadherin表达下调,间质细胞分子标志物Fibronectin、Vimentin表达上调;过表达miR-4295后PTEN蛋白表达抑制荧光素酶活性降低.结论 miR-4295靶向调节PTEN蛋白促进上皮细胞间质化的过程增强鼻咽癌细胞的转移及侵袭.Objective To investigate the possible functions and mechanism of miR-4295 in metastasis of human nasopharyngeal carcinoma （NPC）.Methods We tested the expression of miR-4295 in the NPC cell lines by reverse transcriptase polymerase chain reaction （QPCR）;packing retrovirus of pMSCV-puro and pMSCV-miR-4295 to infect 5-8F cells to establish stable cell lines.We tested the invasion and migration ability of cells by Transwell invasion assay,wound healing assay,and 3-D culture.Western blot （WB） and QPCR were used to test the E-Cadherin,Fibronectin,and Vimentin expression between 5-8F/miR-4295 and 5-8F/vector at both mRNA and protein levels;luciferase activity report was used to verify the target of miRNA.Results We found the miR-4295 had obviously higher expression in 5-8F cell.Transwell invasion assay,wound healing assay,and 3-D culture in matrigel found that the migration and invasion ability of 5-8F were strongly promoted by stable overexpression of miR-4295.Western blot and QPCR analysis showed that the expression level of epithelial markers E-cadherin in 5-8F/miR-4295 was downregulated,while the expression of mesenchymal markers Fibronectin and Vimentin were upregulated.PTEN was identified by luciferase reporter as a direct target of miR-4295.Its expression was downregulated by miR-4295.Furthermore,the results showed that the expression of miR-4295 was inversely correlated with the expression of PTEN protein.Conclusion miR-4295 might promote epithelial to mesenchymal transition （EMT） and human nasopharyngeal carcinoma metastasis by PTEN in vitro.

关 键 词：miR-4295 PTEN 上皮细胞间质化 鼻咽癌 细胞转移和侵袭 

分 类 号：R739.63[医药卫生—肿瘤]