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目的:探讨通过反义RNA技术下调Raptor基因的表达,研究宫颈癌细胞株C33a中Raptor下调后宫颈癌细胞增殖及对化疗药物的敏感性。方法:C33a细胞分3组,实验组和阴性对照组分别转染Raptor siRNA和阴性对照siRNA,空白对照组不做任何处理。通过荧光实时定量PCR和蛋白印记法分别检测C33a转染siRNA后Raptor在mRNA和蛋白水平的表达,CCK-8法检测各组细胞的增殖和对顺铂敏感性的变化,流式细胞仪检测各组及加入顺铂后的凋亡情况。结果:转染siRNA 72 h后,Raptor分别在mRNA水平及蛋白水平较阴性对照下调为82%及78%(P<0.05),转染72 h后CCK-8法测细胞增殖,A值由空白对照组1.828±0.059、阴性对照组1.723±0.044明显降低为Raptor siRNA组的1.349±0.040(P<0.05),各组加入8μM顺铂后,Raptor siRNA组凋亡率为(40.14±1.57)%较空白对照组(21.76±1.23)%和阴性对照组(20.81±1.45)%明显增加(P<0.05)。结论:Raptor siRNA能显著下调C33a细胞中Raptor在mRNA和蛋白水平的表达,抑制C33a细胞的增殖,增加对化疗药物顺铂的敏感性和促凋亡作用。
OBJECTIVE: To investigate the down-regulation of Raptor gene expression by antisense RNA and to study the effect of Raptor on cervical cancer cell proliferation and chemosensitivity after Raptor down-regulation in cervical cancer cell line C33a. Methods: C33a cells were divided into 3 groups. The experimental group and the negative control group were transfected with Raptor siRNA and negative control siRNA respectively. The blank control group was not treated. The expression of Raptor mRNA and protein in C33a transfected siRNA was detected by real-time quantitative PCR and Western blotting. The proliferation and the sensitivity to cisplatin in each group were detected by CCK-8, Group and the apoptosis after adding cisplatin. RESULTS: After 72 h siRNA transfection, Raptor mRNA levels and protein levels were down-regulated to 82% and 78%, respectively (P <0.05). After 72 hours of transfection, cell proliferation was measured by CCK-8 assay. 1.828 ± 0.059 in the control group and 1.723 ± 0.044 in the negative control group were significantly decreased to 1.349 ± 0.040 (P <0.05) in the Raptor siRNA group. The apoptotic rate in the Raptor siRNA group was (40.14 ± 1.57)% The percentage of the control group (21.76 ± 1.23%) and the negative control group (20.81 ± 1.45)% were significantly increased (P <0.05). CONCLUSION: Raptor siRNA can down-regulate the expression of Raptor mRNA and protein in C33a cells, inhibit the proliferation of C33a cells and increase the sensitivity to cisplatin and promote the apoptosis of C33a cells.