摘要
目的探讨caveolin-1对喉鳞状细胞癌细胞系HEp2凋亡的影响。方法通过脂质体法把caveolin-1基因导入喉鳞癌细胞系HEp2细胞,筛选出稳定高表达caveolin-1的克隆,用荧光实时定量反转录-聚合酶链反应(real-timeRT-PCR)和免疫印迹法鉴定;流式细胞仪检测细胞周期与凋亡;Real-time RT-PCR检测survivin的mRNA水平;免疫印迹法检测细胞BCL-2和survivin蛋白的表达;用分光光度法方法检测caspase-3相对活性。结果成功筛选到稳定高表达caveolin-1的克隆,命名为HEp2-CAV1;与对照组相比,更多的HEp2-CAV1细胞处于G0/G1期,凋亡率增加,caspase-3相对活性水平上升2.72倍(P<0.05),survivinmRNA的表达水平减少74%(P<0.01),survivin与BCL-2蛋白表达水平明显减少。结论Caveolin-1能够促进喉鳞状细胞癌细胞株HEp2的凋亡,BCL-2和survivin的表达下调可能是其促凋亡的重要机制之一。
Objective To investigate the effects of caveolin-1 on apoptosis of laryngeal squamous cell carcinoma cell line HEp2.Methods Human caveolin-1 gene was transfected into HEp2 cell line,the positive clones with high expression of caveolin-1 were identified by fluorescence quantitative real time reverse transcriptase-polymerse chain reaction（real-time RT-PCR）and Western blot.Cell cycle and apoptosis were analyzed by flow cytometry.Caspase-3 relative activities were calculated according to the changs of optical density after reaction with caspase-3 substrate.Real-time RT-PCR was used to determine the expression level of survivin mRNA and the expression of BCL-2 and survivin protein were detected by Western blot.Results The clones stably overexpressing caveolin-1 were obtained as HEp2-CAV1.Overexpressing caveolin-1 resulted in the cell cycle arrest at G0/G1 phase and increased the apoptosis cell fraction.Compared with the parental HEp2 cells,the realtive activites of caspase-3 increased by 2.72 fold（P0.05）,the expression level of survivin mRNA reduced by 74%（P0.01） and the protein BCL-2 significantly decreased in HEp2-CAV1 cells,respectively.Conclusion Overexpression of caveolin-1induces apoptosis in laryngeal squamous cell carcinoma HEp2 cell line,downregulation of BCL-2 and survivin expression may be involved in its mechanism.
出处
《基础医学与临床》
CSCD
北大核心
2010年第9期971-975,共5页
Basic and Clinical Medicine
基金
湖州市科技攻关项目(2008GS07)
