天津医药

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人剪切修复基因XPD对人肝癌SMMC-7721细胞中Ets-1和Cdk6基因的调控作用

王娟1,杜芳腾2,杜瑶3,张吉翔4   

  1. 1. 江西省南昌市民德路1号南昌大学第二附属医院消化内科
    2. 江西省南昌大学第二附属医院消化科
    3. 江西省南昌大学医学院
    4. 南昌大学第二附医院消化内科
  • 收稿日期:2012-02-28 修回日期:2012-06-18 出版日期:2012-09-15 发布日期:2012-09-15
  • 通讯作者: 王娟

Role of XPD in Hepatoma Cells regulating Ets-1 and Cdk6

  • Received:2012-02-28 Revised:2012-06-18 Published:2012-09-15 Online:2012-09-15
  • Contact: Juan WANG

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摘要： 目的：将野生型人剪切修复基因XPD转染入人肝癌细胞SMMC-7721后，观察转染后细胞内Ets-1和Cdk6基因的表达变化及对SMMC-7721肝癌细胞增殖的影响。方法：将人工合成的pEGFP-N2-XPD重组质粒通过Lipofectamine 2000TM转染SMMC-7721细胞。本实验分为重组质粒转染细胞SMMC-7721-pEGFP-N2-XPD组、空载质粒转染细胞SMMC-7721-pEGFP-N2组、脂质体组、无转染空白对照组。分别用逆转录-聚合酶链反应（RT-PCR）和蛋白印迹法（Western blot）检测细胞中XPD、Ets-1、Cdk6基因的mRNA和蛋白质的表达量，并用流式细胞仪检测细胞周期变化，四甲基偶氮唑盐微量酶反应比色法（MTT）检测各组细胞的增殖活力。结果：RT-PCR与Western blot检测结果示XPD组中的XPD的mRNA和蛋白质表达较其他3组明显增高（P<0.01），而Ets-1、Cdk6的mRNA和蛋白质表达较其他3组明显减少（P<0.01）。流式细胞仪检测发现转染pEGFP-N2-XPD重组质粒后细胞停滞在G1期，难于进入S期。MTT检测示转染了野生型XPD的SMMC-7721细胞增殖能力减弱。结论：XPD基因可能通过抑制Ets-1、Cdk6基因的表达影响肝癌细胞的生长。

关键词: 肝肿瘤, XPD, Ets-1, Cdk6, 转染, 细胞周期

Abstract: Objective：To transfect wild-type XPD gene into human hepatoma cell line SMMC-7721 and to observe the expression of Ets-1 and Cdk6 gene and the influence in the growth and proliferation of SMMC-7721 cells. Methods：The pEGFP-N2-XPD was transfected into SMMC-7721 cells by Lipofectamine 2000TM. There were four groups in my study including SMMC-7721-pEGFP-N2-XPD group(XPD group)，SMMC-7721-pEGFP-N2 group(N2 group)，Lipofectamine group(Lip group) and blank control group. The expressions levels of XPD，Ets-1 and Cdk6 were detected by RT-PCR and Western blot. Flow cytometry(FCM) was used to analyze the cell cycle of SMMC-7721 cells. The cell proliferation was measured by MTT assay. Results: Compared with blank control group，N2 group and Lip group，the expression of XPD mRNA and protein was enhanced significantly in XPD group(P<0.01)，the expression of Ets-1 and Cdk6 mRNA and protein were decreased obviously(P<0.01). FCM showed that XPD prevented the hepatoma cells from G1 stage to S stage. The proliferation ability of SMMC-7721 cells was observably reduced after transfected by wild-type XPD gene(P<0.01). Conclusion: XPD gene may regulate the hepatoma cell proliferation by inhibiting the Ets-1 and Cdk6 gene.

Key words: liver neoplasms, XPD, Ets-1, Cdk6, transfect, cell cycle