吴茱萸碱抑制Huh7细胞生长并增强细胞对TRAIL的敏感性

Evodiamine inhibits growth of Huh7 cells and enhances their sensitivity to TRAIL

目的:探讨吴茱萸碱对人肝癌Huh7细胞生长和凋亡的影响,阐明吴茱萸碱促进肿瘤坏死因子凋亡诱导配体(TRAIL)抗肿瘤活性的分子机制。方法:MTT法检测吴茱萸碱对Huh7细胞活力的影响;流式细胞术观察吴茱萸碱对细胞周期的阻滞;TUNEL染色法检测细胞凋亡的变化;Western blot实验测定细胞内细胞周期和凋亡相关蛋白的表达水平。结果:Huh7细胞经吴茱萸碱处理后,细胞活力明显下降(P<0.05);同时,细胞发生G_2/M期阻滞,p27、cyclin B1、细胞分裂周期蛋白2(Cdc2)和p-Cdc2的蛋白水平上调(P<0.05);吴茱萸碱能够诱导Huh7细胞发生凋亡,促进多聚ADP核糖聚合酶(PARP)和caspase-3的切割。当吴茱萸与TRAIL联用后,Huh7细胞活力明显下降,PARP和caspase-3的切割增加;另外,吴茱萸上调Huh7细胞中死亡受体5(DR5)的蛋白水平。结论:吴茱萸碱通过抑制细胞活力和阻滞细胞周期于G_2/M期而抑制细胞生长,并诱导Huh7细胞发生凋亡;上调DR5的表达水平可能与吴茱萸碱增强Huh7细胞对TRAIL的敏感性相关。

AIM： To investigate the effects of evodiamine on the growth and apoptosis of human hepatocellular carcinoma Huh7 cells,and to illustrate the molecular mechanism that evodiamine enhances antitumor activity of tumors necrosis factor-related apoptosis-inducing ligand（ TRAIL） in Huh7 cells. METHODS： The cell viability was measured by MTT assay. The cell cycle distribution was analyzed by flow cytometry. The apoptosis rate was determined by TUNEL staining. The protein levels of cell cycle-and apoptosis-related proteins were detected by Western blot analysis. RESULTS：Treatment of Huh7 cells with evodiamine reduced the cell viability（ P 〈 0. 05）. Evodiamine induced cell cycle arrest in G2/M phase by upregulation of p27,cyclin B1,cell division cycle protein 2（ Cdc2） and p-Cdc2. Evodiamine triggered apoptosis accompanied by cleavage of caspase-3 and poly（ ADP-ribose） polymerase（ PARP）. Combination of evodiamine with TRAIL significantly reduced the cell viability and increased cleavage of caspase-3 and PARP as compared with the use of each agent alone. Moreover,evodiamine increased the expression of death receptor 5（ DR5） in the Huh7 cells. CONCLUSION： Evodiamine inhibits the cell growth by reducing the cell viability and inducing cell cycle arrest. Evodiamine also triggers cell apoptosis and enhances the sensitivity of Huh7 cells to TRAIL by upregulating the expression of DR5.