基础医学与临床 ›› 2012, Vol. 32 ›› Issue (2): 175-180.

• 研究论文 • 上一篇    下一篇

阻断PLCε基因与膀胱癌细胞侵袭性相关机制研究

欧俐苹1,罗春丽2   

  1. 1. 重庆医科大学
    2. 重庆医科大学医学检验系
  • 收稿日期:2010-12-31 修回日期:2011-08-22 出版日期:2012-02-05 发布日期:2012-01-12
  • 通讯作者: 罗春丽 E-mail:luochunli79@hotmail.com
  • 基金资助:
    重庆市教委自然科学基金

Mechanism of PLCepsilon gene blockage on invasive power of human bladder cancer cells

Li-Ping OU1,Chun-Li LUO   

  • Received:2010-12-31 Revised:2011-08-22 Online:2012-02-05 Published:2012-01-12
  • Contact: Chun-Li LUO E-mail:luochunli79@hotmail.com
  • Supported by:
    Research Item Foundation of Educational Committee in Chongqing

摘要: 目的 构建PLCε基因的shRNA表达载体转染膀胱癌细胞T24株,观察其对T24细胞转移侵袭等恶性行为的影响。方法 设计并合成针对PLCε基因mRNA的寡核苷酸序列,插入绿色荧光蛋白质粒真核表达载体pGenesil中,构建重组载体pGenesil-PLCε。重组载体转染T24细胞后,RT-PCR、Western blot检测对T24细胞PLCε表达的作用;以侵袭小室、明胶酶谱分析及免疫组化检测其对T24细胞侵袭能力变化。结果 成功构建了针对PLCε基因的shRNA表达载体。RT-PCR检测显示,2个阳性重组质粒转染膀胱癌细胞T24,对其PLCεmRNA表达抑制率分别为76.0%,73.9%,Western blot检测显示:对PLCε蛋白表达抑制率分别为65.4%,64.8%。对T24细胞的侵袭转移能力, 阳性质粒P1和P2转染组穿膜细胞数分别为(25.8±6.2) 和P2转染组(26.8±5.8),明显少于空白对照组(34.8±6.9)(P<0.01)和阴性质粒HK-A转染组(33.8±5.7)(P<0.01);明胶酶谱及免疫细胞化学显示转染P1、P2均使细胞分泌MMP-2、MMP-9明显低于空白对照和HK-A转染组(p<0.01)。结论 通过阻断PLCε基因,能有效抑制膀胱癌T24细胞株中PLCε基因和其蛋白表达,并且对T24细胞的侵袭转移能力具有一定的抑制作用。

关键词: 膀胱癌, 侵袭, PLCε

Abstract: Objective The aim of this study was to construct a shRNA expression plasmid against gene PLCε and to observe the inhibition of PLCε gene expression in bladder cancer cell line T24.Methods pGenesil-PLCε plasmids were constructed and insert T24 cells,then RT-PCR, Western blot analysis was taken to know about inhibition of PLCε gene expression after the transfection of plasmids. invasive power of T24 were measured before and after transfection by the membrane invasion culture system(Transwell chamber), gelatin enzymography and immunochemistry of cells. Results It was confirmed by digesting and sequencing that the two recombinant plasmids had been constructed successfully. The inhibition rate of PLCε mRNA was 76.0% and 73.9%, and this of protein expression was 65.4% and 64.8% . The invasion number of T24 transfected with P1(25.8±6.2) and P2(26.8±5.8) are both lower than the the group of HK-A (33.8±5.7 )and group without transfection(34.8±6.9) (P<0.01); MMP-2,MMP-9 had lower expression in gelatin enzymography assay and imunocytochemistry of cells than the groups HK-A and control(p<0.01). Conclusion:RNAi of PLCε might decrease invasive power of bladder cancer cells so as to inhibit the development of tumor.

Key words: Bladder Cancer, invasive power, PLCε

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