siRNA沉默Livin基因表达对肝癌细胞HepG2生物学行为的影响

Influence of siRNA silencing expression of Livin gene on the biological behavior of HepG2 cells

目的:研究应用siRNA沉默Livin基因表达后肝癌细胞HepG2的生物学功能变化。方法:用脂质体lipofectamineTM2000将携带Livin基因的siRNA载体转染至HepG2细胞内,RT-PCR、Westernblot检测转染前后Livin基因mRNA和蛋白质的表达改变;流式细胞技术(FCM)和TUNEL检测转染前后肝癌细胞凋亡变化。结果:RT-PCR及Western blot检测发现转染siRNA后Livin基因的mRNA和蛋白质表达均明显下降(P<0.05);FCM检测发现实验组、阴性对照组和空白组肝癌细胞HepG2在G1期所占比例分别是(58.99±1.173)%、(41.85±2.82)%、(41.25±1.24)%,实验组与其他两组比较差异均有统计学意义(P<0.05);TUNEL技术检测表明实验组肝癌细胞HepG2凋亡数目明显增加,细胞凋亡率为(67.56±2.56)%,凋亡率明显高于阴性对照组(27.21±12.58)%和空白组(14.09±5.16)%(P<0.05)。结论:在肝癌细胞HepG2中,Livin基因沉默具有诱导肝癌细胞凋亡、抑制肝癌细胞生长的作用。

Objective： To explore the biological significance of silencing Livin gene expression with siRNA in hepatocellular carcinoma HepG2 cells. Methods： HepG2 cells were transfected with synthetic small interfering RNA （siRNA） targeting Livin. Expression of Livin mRNA and protein wererespectively measured by reverse transcription polymerase chain reaction （RT-PCR） and Western blot. Flow cytometry （FCM） and TUNEL assay were used to examine cell cycle and apoptosis in transfected ceils. Results： The result of RT-PCR and Western blot showed that the mRNA and protein levels of Livin declined markedly in transfected cells （P〈0.05）. FCM demonstrated that the cell cycle was arrested in the G1 phase, the apoptotic rate in those groups were （58.99± 1.173）%, （41.85± 2.82）%, （41.25 ± 1.24）%, as the transfected group, there were significant differences compared with other two groups （P〈0.05）. The result of TUNEL assay indicated that the number of apoptotic cells in transfected group were obviously increase, the apoptotic rate was （67.56 ± 2.56） %, there were significant differences compared with negative control group （27.21 ±12.58）% and mock group （14.09 ± 5.16）%（P〈0.05）. Conclusion： siRNA-mediated downregulation of Livin expression can induce apoptosis and diminish the growth, proliferation in hepatocellular carcinoma HepG2 cells.