敲低及过表达G6PD对肝癌细胞增殖和迁移的影响

Effect of knockdown or overexpression of G6PD on the proliferation and migration of hepatoma cells

目的研究在人肝癌细胞株PLC/PRF/5中,敲低及过表达G6PD对细胞株增殖、生长及迁移能力的影响。方法包装慢病毒颗粒,选用人肝癌细胞株PLC/PRF/5建立G6PD敲低及过表达稳转细胞株,通过PCR及Western blot检验过表达及敲低效果,利用建立好的细胞株进行功能实验:通过实时细胞分析系统(RTCA)记录细胞增殖及迁移曲线,EDU实验可直观显示处于DNA复制阶段的细胞比例,以克隆形成实验反映细胞的生长能力。结果在G6PD敲低株中,细胞的倍增时间较对照组延长,生长速率明显下降,EDU实验中处于增殖期细胞的比例较对照组下降43.2%,克隆形成率明显下调(P<0.05),敲低组的迁移速率亦明显降低,曲线分离显著;而在过表达株及其对照组两株细胞中,增殖、生长及迁移实验结果无明显差异。结论在肝癌细胞中降低G6PD的表达将抑制肝癌的增殖生长,为进一步研究肝癌的发生机制及治疗方案打下了基础。

Objective To investigate the effect of knockdown or overexpression of G6PD on proliferation, growth and migration of human hepatocellular carcinoma ceil PLC/PRF/5. Methods Lentivirus-mediated knock- down or overexpression of G6PD was achieved in human hepatoceliular carcinoma cell line PLC/PRF/5. RT-PCR and Western blotting assay were used to detect the overexpression or knockdown of G6PD. Cell proliferation and mi- gration curves were recorded by real-time cell analysis system （RTCA）, the cell proportion in the DNA replication phase can be directly displayed with EDU experiment, cell growth ability was detected by colony forming assay. Results The doubling time of cells in G6PD knockdown group was longer than that of the control group, and the cell growth rate decreased significantly, the proportion of cells in proliferative phase （43.2 % ） was lower than that in the control group, but the rates colony formation and migration were significantly decreased （P 〈 0.05, respective- ly）, and the migration curves separated apparently. While no significant differences in proliferation, growth and mi- gration of PLC/PRF/5 cells were found between the over-expressed strain and the control group. Conclusion The reduction of G6PD expression in HCC ceils inhibits the proliferation and growth of HCC, which may lay a foundation for the further study of the pathogenesis and treatment of HCC.