RNA干扰介导的生存素基因沉默对卵巢癌细胞系SKOV3和SKOV3/ADM凋亡的影响

[Effect of RNAi-mediated survivin gene silencing on apoptosis of ovarian cancer cell lines SKOV3 and SKOV3/ADM].

作者信息

Deng Kai-Xian, Zhong Ling, Jiang Mei-Xian, Chen Wei-Xian, Chen Ying, He Hua

机构信息

Department of Obstetrics and Gynecology, The Second Affiliated Hospital, Chongqing University of Medical Science, Chongqing, P. R. China.

出版信息

Ai Zheng. 2005 Aug;24(8):945-50.

PMID:16086871

Abstract

BACKGROUND & OBJECTIVE: RNA interference (RNAi) has been widely used in tumor gene therapy, antivirus, and gene drug selection. Survivin gene is highly expressed in ovarian cancer cell line SKOV3 and drug-resistant cell line SKOV3/ADM, and is an ideal target of gene therapy for ovarian cancer. This study was to explore effect of RNAi-mediated Survivin gene silencing on apoptosis of SKOV3 and SKOV3/ADM cells.

METHODS

Recombinant plasmid pshRNA-Survivin was transfected into SKOV3 and SKOV3/ADM cells. The expression of Survivin was detected by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The apoptosis of SKOV3 and SKOV3/ADM cells after transfection were evaluated by AO/EB dyeing, TUNEL, and flow cytometry.

RESULTS

After transfection of pshRNA-Survivin, mRNA and protein levels of Survivin gene in SKOV3 and SKOV3/ADM cells were obviously reduced; the apoptosis of SKOV3 and SKOV3/ADM cells were significantly higher in transfection group than in control group (P0.01); 48 h after transfection, the apoptosis rate was 14.05% in SKOV3 cells, and 21.02% in SKOV3/ADM cells, no obvious apoptosis was detected in control cells.

CONCLUSION

pshRNA-Survivin could reduce the expression of Survivin gene, and induce apoptosis of SKOV3 and SKOV3/ADM cells.

摘要

背景与目的

RNA干扰（RNAi）已广泛应用于肿瘤基因治疗、抗病毒及基因药物筛选。生存素基因在卵巢癌细胞系SKOV3及耐药细胞系SKOV3/ADM中高表达，是卵巢癌基因治疗的理想靶点。本研究旨在探讨RNAi介导的生存素基因沉默对SKOV3和SKOV3/ADM细胞凋亡的影响。

方法

将重组质粒pshRNA-Survivin转染至SKOV3和SKOV3/ADM细胞。采用半定量逆转录-聚合酶链反应（RT-PCR）和蛋白质印迹法检测生存素的表达。通过AO/EB染色、TUNEL法及流式细胞术评估转染后SKOV3和SKOV3/ADM细胞的凋亡情况。

结果

转染pshRNA-Survivin后，SKOV3和SKOV3/ADM细胞中生存素基因的mRNA和蛋白水平明显降低；转染组SKOV3和SKOV3/ADM细胞的凋亡率显著高于对照组（P<0.01）；转染48 h后，SKOV3细胞凋亡率为14.05%，SKOV3/ADM细胞凋亡率为21.02%，对照组细胞未检测到明显凋亡。