摘要
目的:研究Akt1基因沉默对人喉癌细胞系(Hep-2)增殖和凋亡的影响。方法:设计并合成针对Akt1的特异性小干扰RNA(siRNA)序列,利用转染试剂转入体外培养人Hep-2细胞,48h后收集细胞,RT-PCR和Westernblot验证siRNA的沉默效应,MTT法检测细胞增殖活性、台盼蓝细胞染色法计数细胞存活率、AnnexinV/PI检测细胞凋亡和坏死。结果:设计的Akt1siRNA能够有效抑制体外培养Hep-2细胞内Akt1表达实现其基因沉默效应;Akt1siRNA干扰组Akt1mRNA转录和蛋白表达水平均低于空白和阴性对照组;Akt1siRNA干扰组细胞增殖活性下降,细胞存活率下降,Akt1siRNA干扰后细胞凋亡率和坏死率增加。结论:RNA干扰Akt1基因沉默具有抑制喉癌细胞增殖,促进细胞凋亡的作用,Akt1可作为喉癌基因治疗的后选新靶点。
Objective： To investigate the effect of siRNA targeting Aktl gene on the proliferation and apoptosis of cuhured human laryngeal carcinoma cell line. Methods： siRNA targeting Akt1 was designed and transfected into Hep-2 cell. At 48 hours after transfection, RT-PCR and western blot technique were used to test the inhibitory effect of Aktl siRNA. The activity of cell proliferation was examined by MTT assay and survival rate was calculated after trypan blue staining. Cell apoptosis and necrosis were tested with AnnexinV/PI kit. Results： The Akt1 siRNA designed could interfere the Aktl expression of Hep-2. The mRNA and protein expression level of Aktl gene of Akt1 siRNA group were lower than that of blank and negatiw control group; The proliferative activity was inhibited, survival rate was decreased, cell apoptosis and necrosis rate increased after AktlsiRNA transfection. Conclusion： RNA interference can inhibit cell proliferation and induce apoptosis of laryngeal carcinoma cell line. Aktl may be a novel target in the gene therapy of laryngeal carcinoma.
出处
《中国当代医药》
2011年第36期7-9,13,共4页
China Modern Medicine
基金
辽宁省博士启动基金资助项目(20081042)
