EGCG诱导人肝癌SMMC-7721细胞早期凋亡及基因和蛋白谱的变化研究  

Induction of early apoptosis in human hepatocellular carcinoma cell line SMMC-7721 by epigallocatechin gallate(EGCG)and alteration of genomic and proteomic expression profiles

作　　者：李刚[1] 容敏华[1] 钟艳平[1] 丁兰芳[1] 覃健[1] 廖明[1] 萧浩[1] 何敏[1] 

机构地区：[1]广西医科大学医学科学实验中心,广西南宁530021

出　　处：《中国药理学通报》2009年第11期1482-1486,共5页Chinese Pharmacological Bulletin

基　　金：广西应用基础研究专项资助项目(桂科应No0731063);广西科学研究与技术开发计划资助项目(桂科能No05112001-4A;0630006-5E;0842009)

摘　　要：目的探讨表没食子儿茶素没食子酸酯(EGCG)诱导人肝癌SMMC-7721细胞早期凋亡的变化规律,以及EGCG作用后SMMC-7721细胞基因及蛋白谱的变化。方法通过MTT法初步筛选EGCG诱导SMMC-7721细胞凋亡的作用浓度,流式细胞Annexin V-FITC/PI法检测EGCG对SMMC-7721细胞增殖的抑制作用和早期凋亡的诱导效应;运用Af-fymetrix U133A2.0人基因组表达谱芯片检测EGCG作用后SMMC-7721细胞基因表达谱的变化,以及SELDI检测EGCG作用后SMMC-7721细胞蛋白图谱的变化,Real-time PCR验证3个表达差异显著基因。结果EGCG诱导SMMC-7721细胞早期凋亡的最佳作用剂量是218.2μmol·L-1,早期凋亡的作用呈剂量依赖性;SMMC-7721细胞经EGCG作用后,表达差异两倍以上的基因共196个,包括上调基因132个和下调基因64个;EGCG作用后表达差异两倍以上的蛋白共43个,包括23个表达上调蛋白,20个表达下调蛋白。Real-time PCR验证DIO2、Id3基因表达与芯片结果一致。结论EGCG有明显的诱导肝癌SMMC-7721细胞早期凋亡的作用,其诱导凋亡作用涉及多基因和多蛋白变化。Aim To investigate the changes of apoptosis inducing hepatocellular carcinoma cell line SMMC-7721 by epigallocatechin gallate（EGCG）and observe the alteration of genomic and proteomic expression profiles in SMMC-7721 cells treated by EGCG.Methods MTT assay was used to preliminarily detect the effect concentration of EGCG on induction of apoptosis in SMMC-7721 cells.The growth inhibition and early apoptosis of SMMC-7721 cells was measured by flow cytometry using Annexin V-FITC/PI staining.Alteration of genomic and proteomic expression profiles of SMMC-7721 cells were detected by Affymetix U133A 2.0 array and SELDI respectively.Three differentially expressed genes were verified by Real-time PCR.Results The best dose of EGCG effecting on induction of early apoptosis in SMMC-7721 cells was 218.2 μmol·L-1,in a dose-dependent manner.When the SMMC-7721 cells were treated by EGCG,a total of 196 differential genes were found out,with a difference of more than two times in expression levels.Of the 196 genes,132 genes were up-regulated and 64 genes were down-regulated.Meanwhile,a total of 43 differential proteins were found out,with a difference of more than two times in expression levels.Of the 43 proteins,23 proteins were up-regulated and 20 proteins were down-regulated.The Real-time PCR confirmed expression levels of DIO2 and Id3,which were consistent with results using microarray technology.Conclusions EGCG significantly induces early apoptosis in human hepatocellular carcinoma SMMC-7721 cells.The apoptosis is associated with alteration of many genomic and proteomic expression profiles.

关 键 词：EGCG 细胞早期凋亡 基因谱 蛋白谱 REAL-TIME PCR 肝癌 

分 类 号：R284.1[医药卫生—中药学] R329.25[医药卫生—中医学]