抑制NEK7促进肝癌细胞凋亡的实验

目的 采用体外实验验证抑制NEK7表达后肝癌细胞增殖、衰老及凋亡的变化，并探索其机制。方法 Western blot及RT-PCR检测不同肝癌细胞系及人肝永生化THLE-2细胞系中的NEK7表达，针对NEK7基因序列设计抑制其表达的shRNA，转染肝癌细胞后，观察细胞体外增殖活性、衰老、凋亡及细胞周期的变化，Western blot检测抑制NEK7表达后细胞周期相关因子的变化。结果 肝癌细胞中NEK7呈高表达，shRNA慢病毒转染肝癌细胞抑制NEK7表达后，肝癌细胞增殖能力受到抑制，细胞衰老及凋亡比例显著升高，S及G2/M期细胞数明显减少，细胞周期受到阻滞，C-myc、c-Fos、cyclin D1、cyclin E等细胞周期因子表达水平受到抑制，P16及P27表达水平升高，CDK2、CDK4及CDK6的表达无明显变化。结论 靶向抑制NEK7表达，降低了肝癌细胞的增殖能力，并促进细胞衰老、诱导其凋亡，同时肝癌细胞周期进程受阻。

Experiment on Inhibiting NEK7 to Promote Apoptosis of Hepatocellular Carcinoma Cells

Objective To use in vitro experiments to verify the changes of proliferation, senescence and apoptosis of hepatocellular carcinoma cells after inhibiting the expression of NEK7, and to explore the related molecular mechanism. Methods Western blot and RT-PCR were used to detect the expression of NEK7 in hepatocellular carcinoma cells and THLE-2 cells. A viral vector was designed to inhibit the expression of NEK7 based on the gene sequence. After hepatocellular carcinoma cells were transfected, we observed the changes of proliferation activity, cell senescence, cell apoptosis and cell cycle in vitro. Western blot was used to detect the expression of cell cycle-related factors. Results Compared with THLE-2 cells, NEK7 was highly expressed in hepatocellular carcinoma cells. After inhibiting the expression of NEK7 with shRNA, the proliferation of hepatocellular carcinoma cells was inhibited, the proportions of cell senescence and apoptosis were increased, meanwhile, the cell number in stage S and G2/M was significantly reduced, the cell cycle progression was blocked, the expression levels of C-myc, c-Fos, cyclin D1 and cyclin E were inhibited, P16 and P27 expression were increased, and CDK2, CDK4 and CDK6 expression were not significantly changed. Conclusion After inhibiting the expression of NEK7, the proliferation ability of hepatocellular carcinoma cells is reduced, cell senescence is promoted and apoptosis is induced; meanwhile, the cell cycle progress is blocked.