hTERT启动TRAIL基因的腺病毒载体对人舌鳞癌细胞影响

Effect of TRAIL Gene in Squamous Cell Carcinoma Cells Induced by Adenovirus

目的:探讨以凋亡基因TRAIL为靶基因,腺病毒(Ad)为载体,人端粒酶(TERT)启动子引导的重组基因治疗药物AdTERT-TRAIL诱导人鳞状细胞癌细胞(TCa8113)凋亡的作用。方法:含有绿色荧光蛋白基因(EGFP)的腺病毒载体(AdTERT-EGFP)转染TCa8113细胞,确定重组腺病毒载体转染效率;AdTERT-EGFP为对照组,AdTERT-TRAIL为实验组感染TCa8113细胞,采用RT-PCR方法检测TRAIL的表达。应用倒置显微镜观察细胞形态,应用MTT检测细胞的存活率,流式细胞仪检测细胞凋亡率。结果:AdTERT-EGFP在1000p/cell时转染效率为100%。RT-PCR可检测到594bp的TRAIL基因。细胞转染AdTERT-TRAIL和AdTERT-EGFP后,随时间细胞存活率逐渐下降,但AdTERT-TRAIL组较AdTERT-EGFP组下降更快,统计学分析有显著意义(P<0.001)。AdTERT-TRAIL和AdTERT-EGFP均能诱导TCa8113细胞凋亡,而AdTERT-TRAIL组引起细胞凋亡的程度明显比AdTERT-EGFP组大,统计学有显著差异(P<0.0001)。结论:AdTERT-TRAIL在体外能明显抑制TCa8113细胞的活性,并能促进其凋亡。

Objective：To study the apoptotic effect on the squamous cell carcinoma cell line TCa8113 induced by recombined adenovirus vector containing TRAIL gene and TERT promoter.Methods：The TCa8113 cell line was firstly infected with AdTERT-EGFP containing enhanced green fluorescence protein gene（EGFP）,in order to obtain the definite titre of this adenovirus vector.Then,the TCa8113 cell line was infected with AdTERT-TRAIL,and the expression of TRAIL gene was detected by means of RT-PCR,the activity of TCa8113 cell line was evaluated by MTT and the apoptosis were detected by flow cytometer.Results：The proper titre was 1000p/cell,and TCa8113 cell line could be infected 100% in this titre.TRAIL gene was detected by RT-PCR after the cells were infected with AdTERT-TRAIL.The activity of TCa8113 decreased in both groups,but the AdTERT-TRAIL group decreased more sharply than AdTERT-EGFP group,the statistic analysis is significant（P〈0.001）.Both AdTERT-TRAIL and AdTERT-EGFP could lead to apoptosis of TCa8113 cells,but the AdTERT-TRAIL function more efficiently than AdTERT-EGFP.Especially there was remarkable statistic difference between two groups（P〈0.001）.Conclusion：AdTERT-TRAIL could effectively decrease the activity of TCa8113 cells and induce apoptosis.