【作者】 李萍；

【导师】 张迎梅；

【作者基本信息】 兰州大学 ， 动物学， 2010， 硕士

【摘要】 本文通过采用人肝正常细胞(HL-7702)的体外实验来探讨表没食子儿茶素没食子酸酯(Epigallocatechin Gallate, EGCG)对镉(Cd)诱导肝细胞损伤的保护作用及其机制。研究采用MTT比色法检测镉的细胞毒性及EGCG对细胞活力的影响；Hoechst33258染色实验和琼脂糖凝胶电泳检测细胞凋亡水平；紫外-可见全波段扫描检测(UV-Vis)及核磁共振(NMR)谱分析EGCG与镉离子之间的化学反应；DCFH-DA染色后通过荧光分光光度仪检测细胞内活性氧自由基(ROS)水平。研究结果如下：1.镉处理后HL-7702细胞活力显著性降低(P<0.05),而且细胞内ROS含量显著上升(P<0.01)；并且Hoechst33258染色实验和琼脂糖凝胶电泳检测结果显示镉诱导DNA产生了明显的凋亡片段并诱导了细胞凋亡2.使用不同浓度的EGCG (50、100、200μM)对镉(80μM)预处理48 h的肝细胞分别进行3h的干预后可以使受损细胞的细胞活力得到不同程度的修复,修复率分别为13.8%、33.9%和71.4%。与此同时,EGCG对镉诱导的细胞内的ROS含量也具有明显的清除作用(P<0.01)。3.紫外-可见全波段扫描及核磁共振结果显示EGCG和镉离子(Cd2+)可能发生化学反应。本研究结果表明,EGCG与Cd2+可能发生化学反应,但是对于受到镉损伤的HL-7702细胞,EGCG的保护机制主要是通过清除细胞内镉诱导产生的ROS。更多还原

【Abstract】 The present study aims to investigate the protective effect and potential mechanism of epigallocatechin gallate (EGCG) on Cd-induced cytotoxicity in normal human liver cell (HL-7702). The cytotoxicity of cadmium and the protective effect of EGCG on normal human liver cell were assessed by using MTT assay; apoptosis was shown by using Hoechst33258 staining and agarose gel electrophoresis; changes in intracellular ROS (Reactive Oxygen Species) production were detected by using fluorescent dyes (DCFH-DA); the formation of chelate between EGCG and the cadmium ion was investicated by using UV-visible detection of wide-band scan (UV-Vis) and nuclear magnetic resonance hydrogen spectrum. The results are as follows:1. HL-7702 cells viability decreased significantly (P<0.05) by Cd, and Hoechst33258 staining and agarose gel electrophoresis experiments showed that cadmium induced the induction of DNA fragments and apoptosis.2. Significant elevation of cell viability was observed after the cells were pre-treated (80μM) for 48 h incubating with EGCG at 50,100,200 uM for 3 h respectively. The repair rates were 13.8%,33.9%and 71.4%respectively. Furthermore, EGCG could decreased the intracellular ROS level significantly(P< 0.01).3.EGCG might combining with Cd2+forming Cd2+-EGCG complex.The present result suggested that EGCG could reduce Cd-induced cytotoxicity in HL-7702 cells directly by decreasing the intracellular ROS level.更多还原