表没食子儿茶素没食子酸酯诱导人肝癌细胞HepG2凋亡与TGF-β1-Smad通路激活有关

目的 探讨表没食子儿茶素没食子酸酯(EGCG)对人肝癌细胞HepG2细胞周期和凋亡的影响及其机制.方法 四甲基偶氮唑蓝(MTT)法检测EGCG对HepG2细胞增殖的影响,流式细胞术检测EGCG对HepG2细胞周期的影响以及细胞凋亡情况,逆转录聚合酶链反应(RT-PCR)和萤光素酶报告基因系统检测验证HepG2细胞中TGF-β1-Smad通路的完整性,实时聚合酶链反应(realtime PCR)检测EGCG对HepG2细胞中Smad2、Smad3、Smad4和Smad7 mRNA表达的影响.结果 高浓度EGCG干预时,HepG2细胞的增殖能力随着EGCG浓度和作用时间的增加而下降,呈明显的时效和量效关系,作用72 h,HepG2细胞的IC50为111.76 μmol/L.随EGCG剂量增大,HepG2细胞中G0/G1期细胞的比例逐渐增高,80、120和160 μmol/L EGCG处理组分别为44.9%、54.8%和91.3%;相反,S期细胞比例逐渐降低,80、120和160 μmol/L EGCG处理组依次为38.5%、29.2%和3.0%.随着EGCG剂量增大,HepG2细胞的早期凋亡增加,80、120和160 μmol/L EGCG处理组的早期凋亡率分别为1.82%、4.22%和6.83%;晚期凋亡也随之增加,80、120和160 μmol/L EGCG处理组的晚期凋亡率分别为7.92%、24.19%和27.92%.HepG2细胞中TGF-β1-Smad通路是完整的.EGCG对HepG2细胞中Smad2、Smad3和Smad4 mRNA的表达无明显影响,但下调Smad7 mRNA的表达.结论 EGCG能诱导人肝癌细胞HepG2的凋亡,其机制可能涉及TGF-β1-Smad通路的激活.

Epigallocatechin gallate induces apoptosis in human hepatocellular carcinoma HepG2 cells via TGF/Smad signaling pathway

Objective To investigate the cytotoxic effect of epigallocatechin gallate(EGCG)on human hepatocellular carcinoma cell line HepG2 cells and corresponding changes of TGF-β1-Smad pathway.Methods The cytotoxic effect of EGCG on HepG2 cells was determined by MTT assay.Cell cycle and apoptosis rate were detected by flow cytometry.RT-PCR and luciferase assay were used to verify whether TGF-β1-Smad signaling pathway is intact in HepG2.The mRNA expression of Smad 2,Smad3,Smad4 and Smad7 was detected by real-time PCR.Results EGCG induced apoptosis in the HepG2 ceils in a time-and concentration-dependent manner.The proportion of G1 phase cells was increased gradually as the concentration increased.However,the percentage of cells in S phase was decreased gradually.Annexin V/PI assay demonstrated that early apoptosis increased as the concentration increased,and late apoptosis also increased,when treated with high-concentration EGCG.The intact TGF-β1-Smad pathway was verified by luciferase assay and RT-PCR.There was no significant effect of EGCG on mRNA level of Smad 2,Smad 3,and Smad 4 in HepG2 cells,but downregulated mRNA level of Smad 7.Conclusion EGCG can reduce apoptosis in human hepatocellular carcinoma cell line HepG2 cells.The activation of TGF-β1-Smad signaling pathway may be involved in its cytotoxicity mechanisms.