Department of Medical Oncology, Yijishan Hospital Wannan Medical College, Anhui Wuhu 241000, China.
Cell Biochem Biophys. 2012 Mar;62(2):337-41. doi: 10.1007/s12013-011-9315-0.
The objective of the study was to evaluate the expression of survivin, cell proliferation, and apoptosis in survivin-specific siRNA-transfected human gastric cancer cell line MGC-803. For this purpose, the target gene fragments were cloned into pSilencer3.1-Hl neo vector. Recombinant eukaryotic expression vector, pSilencer3.1-SVV was successfully constructed and then the recombinant vector was transfected into gastric cancer MGC-803 cells. The mRNA expression of survivin was determined by reverse-transcriptase polymerase chain reaction (RT-PCR). Survivin protein expression was detected by Western blot. Cell cycle distribution and apoptosis were determined by flow cytometry. Our data regarding RT-PCR and Western blot showed that pSilencer3.1-SVV vector could knockdown the expression of survivin mRNA and protein. In contrast with the control group, the apoptotic index of MGC-803 cells increased remarkably. Survivin-specific siRNA caused cells accumulation in the G2/M phase and the number of cells in the G0/G1 phase decreased after transfection. It was, therefore, concluded that the siRNA targeting survivin gene could inhibit the proliferation of gastric cancer cells and induce apoptosis. The use of survivin siRNA may provide a novel approach for gene therapy of gastric cancer.
本研究旨在评估 Survivin 表达、细胞增殖和凋亡在 Survivin 特异性 siRNA 转染的人胃癌细胞系 MGC-803 中的作用。为此，将靶基因片段克隆到 pSilencer3.1-Hl neo 载体中。成功构建了重组真核表达载体 pSilencer3.1-SVV，然后将重组载体转染入胃癌 MGC-803 细胞。采用逆转录聚合酶链反应 (RT-PCR) 检测 Survivin 的 mRNA 表达。采用 Western blot 检测 Survivin 蛋白表达。采用流式细胞术检测细胞周期分布和凋亡。我们关于 RT-PCR 和 Western blot 的数据表明，pSilencer3.1-SVV 载体可下调 Survivin mRNA 和蛋白的表达。与对照组相比，MGC-803 细胞的凋亡指数显著增加。转染后，Survivin 特异性 siRNA 导致细胞在 G2/M 期积聚，G0/G1 期细胞数量减少。因此，结论是靶向 Survivin 基因的 siRNA 可抑制胃癌细胞的增殖并诱导凋亡。Survivin siRNA 的应用可能为胃癌的基因治疗提供一种新方法。
