外源性导入抑癌基因FHIT对肝癌细胞Hep3B生长的影响

Effects of fragile histidine triad gene transduction on growth of Hep3B human hepatocellular carcinoma cells

目的:研究外源性人脆性组氨酸三联体(FHIT)基因的高表达对人肝癌细胞Hep3B生长的影响.方法:构建一个包含FHIT基因的真核表达载体pcDNA3.1(+)/FHIT,体外转染人肝癌细胞Hep3B,逆转录-聚合酶链反应(RT-PCR),蛋白印迹(Western blot)分别检测目的基因不同水平的表达,通过细胞生长试验、流式细胞仪分析技术检测细胞转染前后的细胞增殖、细胞周期和细胞凋亡的变化.结果:转染pcDNA3.1(+)/FHIT后Hep3B细胞的FHIT-mRNA,FHIT蛋白表达阳性,转染后对Hep3B细胞的生长有抑制作用,早期细胞凋亡增加,与对照组比较差异有显著性(72.23±0.84 vs 54.36±0.78,53.17±0.52,P<0.05).结论:FHIT基因通过诱导肿瘤细胞发生凋亡并导致其发生G_1期阻滞,在肿瘤基因治疗方面发挥作用.

AIM： To evaluate the effects of the human fragile histidine triad gene on cell proliferation and apoptosis of the Hep3B human hepatocellular carcinoma line. METHODS： A recombinant pcDNA3.1 （＋）/FHIT including functional region of human fragile histidine triad gene （FHIT） was constructed for transfer into human hepatocellular carcinoma cells in vitro, mRNA and protein expressions of the gene in the transfected cells were detected by reverse transcriptase-polymerase chain reaction and Western blot, respectively. MTT [3-（4, 5-dimethylthiazol-2-yl）-2, 5-diphenyltetrazolium bromide] assay was used to assess the effects of FHIT on proliferation. Cell cycle changes and apoptosis were measured by flow cytometry. RESULTS： Hep3B cells expressed high levels of FHIT-mRNA and FHIT protein after infection with pcDNA3.1 （＋）/FHIT. The growth of Hep3B cells treated with pcDNA3.1 （＋）/FHIT was significantly inhibited, pcDNA3.1 （＋）/FHIT-infected Hep3B cells showed a significant increase in G0-G1 phase and FHIT-induced apoptosis compared with controls （72.23 ± 0.84 vs 54.36 ± 0.78 and 53.17 ± 0.52, P 〈 0.05, respectively）. CONCLUSION： Transfer of the FHIT gene inhibited the growth of human hepatocellular carcinoma cells and induced cell apoptosis.