上海交通大学学报（医学版）

›› 2013, Vol. 33 ›› Issue (4): 415-.doi: 10.3969/j.issn.1674-8115.2013.04.007

• 论著（基础研究） • 上一篇    下一篇

RNA干扰抑制EGFR基因表达对子宫内膜癌细胞上皮间质转化的影响

徐 玮, 王 娟, 徐妍力, 马 莉, 艾志红, 滕银成   

  1. 上海交通大学附属第六人民医院妇产科, 上海 200233
  • 出版日期:2013-04-28 发布日期:2013-05-03
  • 通讯作者: 滕银成, 电子信箱: teng1008@sohu.com。
  • 作者简介:徐 玮（1986—）, 男, 硕士生； 电子信箱: xuwei_1003@hotmail.com。
  • 基金资助:

    上海市科委基金（07ZR14059）

Influence of inhibition of EGFR gene expression by RNA interference on epithelial-mesenchymal transition in endometrial carcinoma cells

XU Wei, WANG Juan, XU Yan-li, MA Li, AI Zhi-hong, TENG Yin-cheng   

  1. Department of Obstetrics and Gynecology, the Sixth Peoples´|Hospital, Shanghai Jiaotong University, Shanghai 200233, China
  • Online:2013-04-28 Published:2013-05-03
  • Supported by:

    Shanghai Science and Technology Committee Foundation, 07ZR14059

PDF (PC)

1419

摘要：

目的 构建针对表皮生长因子受体(EGFR)的shRNA质粒载体，转染人子宫内膜癌Ishikawa细胞株，检测RNA干扰对抑制EGFR基因表达及子宫内膜癌细胞上皮间质转化（EMT）的影响。方法 体外构建针对EGFR的shRNA质粒（pRNAT-EGFR shRNA），脂质体法转染Ishikawa细胞。实验设阴性对照组（未经任何转染）、pRNAT-EGFRneg组（空白质粒载体）、pRNATEGFR1组、pRNAT-EGFR2组和pRNAT-EGFR3组。采用实时荧光定量RT-PCR和Western blotting技术检测EGFR mRNA及蛋白的表达变化，并检测转染前后Ishikawa细胞中EMT相关指标E-cadherin、α-catenin、N-cadherin和Vimentin的表达变化。结果 pRNAT-EGFR2组和pRNAT-EGFR3组细胞EGFR mRNA和蛋白的表达量均明显低于阴性对照组，差异有统计学意义（P<0.05）；pRNAT-EGFR1组EGFR mRNA和蛋白的表达量与阴性对照组相比，差异无统计学意义（P>0.05）。RNA干扰抑制EGFR基因表达后，与阴性对照组比较，pRNAT-EGFR2组和pRNAT-EGFR3组Ishikawa细胞上皮标志物E-cadherin和α-catenin的mRNA及蛋白表达水平均上调（P<0.05），间叶标志物N-cadherin和Vimentin的mRNA及蛋白表达水平则显著下调（P<0.01）。结论 抑制EGFR基因表达可部分逆转子宫内膜癌细胞EMT相关基因的表达。

关键词: 表皮生长因子受体, RNA干扰, 子宫内膜癌, 上皮间质转化

Abstract:

Objective To construct targeting epidermal growth factor receptor (EGFR) short hairpin RNA (shRNA) plasmid vector and transfect it into human endometrial carcinoma Ishikawa cells, evaluate the influence of RNA interference (RNAi) to the EGFR gene expression and epithelial-mesenchymal transition (EMT) in endometrial carcinoma cell. Methods shRNA plasmid targeting EGFR (pRNATEGFR shRNA) was constructed in vitro, and was transfected into Ishikawa cells with lipofectamine technique. Negative control group (without transfection), pRNAT-EGFRneg group (blank plasmid vector), pRNAT-EGFR1 group, pRNAT-EGFR2 group and pRNAT-EGFR3 group were established. RT-PCR and Western blotting were employed to detect the expression of EGFR mRNA and protein, and the expression of EMT related markers such as E-cadherin, α-catenin, N-cadherin and Vimentin in Ishikawa cells was determined before and after transfection. Results The expression of EGFR mRNA and protein in Ishikawa cells in pRNAT-EGFR2 group and pRNAT-EGFR3 group was significantly lower than that in negative control group (P<0.05), while there was no significant difference in the expression of EGFR mRNA and protein in Ishikawa cells between pRNAT-EGFR1 group and negative control group (P>0.05). After RNA interference inhibited the expression of EGFR gene in Ishikawa cells, the expression of E-cadherin and α-catenin mRNA and protein in pRNAT-EGFR2 group and pRNAT-EGFR3 group was significantly higher than that in negative control group (P<0.05), while the expression of N-cadherin and Vimentin in pRNAT-EGFR2 group and pRNAT-EGFR3 group was significantly lower than that in negative control group (P<0.01). Conclusion The inhibition of EGFR gene expression could partially reversed the expression of EMT related genes in endometrial carcinoma cells.

Key words: epidermal growth factor receptor, RNA interference, endometrial carcinoma, epithelial-mesenchymal transition