雷公藤甲素对人肝癌细胞株HepG2体内外作用的研究

The antitumor effect of triptolide on HepG2 cells in vitro and in vivo

目的:探讨雷公藤甲素在体内外对人肝癌细胞HepG2的作用及可能的分子机制。方法:应用不同浓度的雷公藤甲素作用于体外培养的HepG2细胞,MTT法检测细胞生长抑制率,流式细胞仪检测细胞凋亡,RT-PCR法和Western blot法分别检测热休克蛋白70(heat shock protein 70,HSP70)mRNA和蛋白表达变化;caspase-3活性检测试剂盒分析其凋亡机制,建立人肝癌细胞HepG2裸鼠皮下种植瘤模型,实验组和对照组分别用雷公藤甲素(0.4mg/kg,每天1次)和相同体积的生理盐水,用药3周后观察肿瘤生长情况。结果:雷公藤甲素可显著抑制体外培养HepG2细胞增殖和诱导细胞发生凋亡(P<0.05),并呈现明显的量-效与时-效关系。应用雷公藤甲素后HepG2细胞中的HSP70的mRNA和蛋白表达明显的降低,呈现浓度依耐性,同时细胞质内caspase-3的活性随着剂量的增加而增加。此外,雷公藤甲素实验组种植瘤体积明显小于对照组(P<0.05)。结论:雷公藤甲素能通过诱导凋亡在体内外发挥对人肝癌细胞HepG2抗肿瘤作用,其机制可能与抑制HSP70蛋白和增加caspase-3的活性有关。

Objective： To study the effect of triptolide on human hepatocelluar carcinoma HepG2 cells proliferation and apoptosis in vitro and in vivo,as well as its possible mechanisms.Methods：HepG2 cells were cultured in triptolide media with different concentrations.Cell viability was assessed by MTT assay.Cell apoptosis was observed by flow cytometry（FCM）.The heat shock protein 70（HSP70） was assayed by Western blotting and RT-PCR methods.Caspase-3 activity was examined by caspase-3 colorimetric assay.In a xenograft tumor model,the control group and experimental group were treated with 0.9% NaCl solution and triptolide（0.4mg/kg,q.d.）,respectively.Three weeks after treatment,the influence of triptolide on the growth of hepatocellular carcinoma was observed.Results：Triptolide treatment resulted in dose-and time-dependent proliferation and apoptosis of HepG2 cells（P 〈 0.05）.Triptolide decreased HSP70 mRNA and protein levels in a dose-dependent pattern,whereas caspase-3 activity was significantly increased in a dose-dependent manner.Moreover,Mice receiving triptolide therapy had significantly smaller tumors than controls（P 〈 0.05）.Conclusion：Triptolide causes human hepatocelluar carcinoma HepG2 cells death in vitro and in vivo by inducing apoptosis and its mechanism of action might be mediated via the inhibition of HSP70 and up-regulation of caspase-3 activity.