ALKBH5抑制肝癌细胞系HepG2和肝细胞系L-02的增殖能力

ALKBH5 inhibits proliferation of HepG2 and L-02 cells

目的探讨Alk B同源蛋白5(Alk B homologue 5,ALKBH5)对正常肝细胞系L-02和肝癌细胞系Hep G2的增殖、周期、凋亡的影响。方法通过慢病毒稳定转染ALKBH5基因的重组载体(p EGFP-C1b-ALKBH5)至肝癌细胞系Hep G2和肝细胞系L-02中,Western印迹鉴定绿色荧光蛋白(GFP-ALKBH5)的稳定表达;实验分GFP-ALKBH5慢病毒组和GFP慢病毒组;细胞增殖实验(cell counting kit-8,CCK-8)检测两组细胞的增殖能力;流式细胞术检测两组细胞周期和细胞凋亡的情况;平板克隆形成实验检测两组细胞的生长能力。结果与GFP对照组相比,过表达GFP-ALKBH5可显著抑制Hep G2和L-02细胞的增殖,引起Hep G2和L-02细胞的周期阻滞,但不影响细胞的凋亡。结论ALKBH5可显著抑制肝癌细胞Hep G2和肝细胞L-02的增殖能力,扮演了抑癌基因的角色,可能在肝细胞癌的发生和发展中起重要作用。

Objective To investigate the effect of AlkB homologue 5 （ALKBH5） on proliferation, cell cycle, and apoptosis of HepG2 and L-02 cells. Methods Recombinant plasmid vector containing the CDS region of ALKBH5 （pEGFP-Clb-ALKBHS） was stably transfected into HepG2 and L-02 cells. Western blotting was used to detect the expression of green fluonescence protein （GFP）-ALKBHS. There were two groups in our experiment： GFP-ALKBH5 lentivirus group and GFP lentivirns group. Characteristics, such as proliferation, cell cycle and apoptosis of HepG2 and L-02, were detected through Cell Counting Kit-8 （CCK-8） assay, flow cytometry and clone formation, respectively. Results The result of Western blotting revealed that ALKBH5 was efficiently up-regulated at protein levels. Despite apoptosis, phenotypic analysis revealed that the proliferation and cell phases were significantly inhibited in ALKBH5 overexpressed stable cell strains compared with the control cells （both P 〈 0.05 ）. Conclusion ALKBH5 can restrain fetal liver cell （L-02） and hepatocellular carcinoma cell （ HepG2 ） from proliferating. Taken together, our results strongly suggest that ALKBH5 can play a key role in the generation and progression in HCC as a tumor suppressor.