摘要目的 探讨以Survivin为靶基因的胰腺癌基因治疗的可能性,为胰腺癌基因治疗提供依据.方法 采用化学合成的小十扰RNA(siRNA)和pGCSi载体中的小发夹RNA(shRNA)抑制胰腺癌细胞系PaTu8988的Sunrivin基因表达,通过观察胰腺癌细胞株Survivin基因表达的下调以及细胞形态、细胞凋亡、细胞活力、凋亡信号途径等的改变评价Survivin作为靶基因的治疗效果.结果 不同序列的siRNA和shRNA抑制胰腺癌细胞Survivin的表达后,Survivin mRNA和蛋白表达水平明显下降(P<0.05);碘化吡啶(PI)染色法观察发现RNA干扰(RNAi)后细胞出现核皱缩、细胞凋亡率>20%;流式细胞仪检测发现RNAi后在G0/G1期前出现了亚二倍体峰(P<0.05);Western blot法检测发现RNAi后Casptrqe-3被激活(P<0.05).结论 通过抑制胰腺癌细胞PaTu8988的Survivin表达可以诱导肿瘤细胞启动凋亡程序,加速肿瘤细胞凋亡,由此可望提高胰腺癌的治疗效果.

abstractsObjective To investigate the influence of gene therapy using survivin as a gene target on biological behavior of pancreatic carcinoma cell line. Methods Chemically synthesized siRNA and shRNA in pGCSi vector were used to silence survivin expression of pancreatic carcinoma cell line PaTu8988. The therapeutical effects of survivin as a gene target were evaluated through determination of the down-regulation of survivin gene expression, cellular shape, cell apoptosis, cell viability and apoptosis signal pathway changes. Results After transfection of different arrays of siRNA and shRNA vectors to silence the survivin expression, survivin mRNA and protein levels were significantly decreased (P < 0.05) ; PI staining revealed the presence of karyopyknosis, the cell apoptosis index was more than 20%; hypodiploid DNA content before G0/G1 detected by flow cytometry ; cell viability measured by MTT assay was significantly decreased (P <0.05) ; the activity of caspase-3 remarkably increased (P < 0. 05). Conclusions The pancreatic carcinoma cell line PaTu8988 be induced to promote spontaneous apoptosis procedure through silencing survivin expression by RNAi, which could accelerate carcinoma cell apoptosis and improve therapeutic effect on pancreatic carcinoma.