RNA干扰技术沉默宫颈癌Hela细胞Survivin基因对细胞生物学特点的影响

Effect of RNAi-mediated silencing Survivin gene on apoptosis of cervical cancer Hela cell line

目的:通过RNA干扰技术沉默宫颈癌Hela细胞中的Survivin基因,观察干扰后核酸表达及细胞凋亡情况发生的变化,为进行宫颈癌的基因治疗提供理论依据。方法:设定空白组、阴性对照组及实验组,空白组Hela细胞未进行基因干扰,阴性对照组为非特异序列干扰,实验组为针对Survivin基因设计的干扰RNA(siRNA)干扰,每组均设三个平行标本。转染后48h收集细胞,RT-PCR技术检测Survivin mRNA表达,流式细胞术检测细胞凋亡情况。结果:转染siRNA 48h后,Hela细胞中Survivin mRNA拷贝数明显减少,实验组mRNA表达量为空白组的23.4%,存在显著性差别(P<0.01);为阴性对照组的23.3%,存在显著性差别(P<0.01)。流式细胞术检测结果表明,实验组Hela细胞凋亡率是空白组的3.08倍(P<0.01);是阴性对照组的2.69倍(P<0.01),均有显著性差异。结论:通过RNA干扰沉默宫颈癌Hela细胞Survivin基因能够促进细胞凋亡的发生,Survivin可能成为基因治疗宫颈癌的理想靶基因。

Objective： To explore effect of RNA interference（ RNAi）- mediated on expression of Survivin gene and apoptosis of cervical cancer Hela cells,and to provide a theoretical basis for cervical cancer targeted therapy.Methods： The experiment was divided into blank group,negative control group and experimental group. The blank group was normal cells without gene interference,the negative control group was non- specific interference,the experimental group Survivin gene RNA（ siRNA） interference. Collected the cells 48 h after transfection. The expression of Survivin was detected by reverse transcription- polymerase chain reaction（ RT- PCR）. The apoptosis of Hela cells after transfection were evaluated by flow cytometry. Results： 48 h after transfection of siRNA- Survivin,mRNA of Survivin gene in Hela cells were obviously reduced; the quantity of mRNA of Hela cells in experimental group was23. 4% for the blank group,had significant difference（ P 0. 01）; 23. 3% for the negative control group,had significant difference（ P 0. 01）. Apoptosis rate of Hela cells in the experimental group was 3. 08 times that of blank group（ P 0. 01）,was 2. 69 times that of negative control group（ P 0. 01）,there were significant differences. Conclusion： RNAi- mediated silencing Survivin gene could induce apoptosis of Hela cells,Survivin may be the ideal target gene for gene therapy of cervical cancer.