EGCG对H_2O_2诱导HLE细胞氧化损伤的保护作用

Protective Effects of EGCG on H_2O_2-induced Oxidative Damage in HLE Cells

目的:探讨表没食子酸酯(EGCG)对体外培养的人晶状体上皮(human lens epithelial,HLE)细胞氧化损伤的保护作用及可能机制。方法:HLE细胞传代培养,分为阴性对照组:以正常培养液培养;氧化损伤组:100μmol·L^(-1)的H_2O_2作用12 h;EGCG低浓度组:10μmol·L^(-1)EGCG孵育24 h后,加入H_2O_2作用12 h;EGCG高浓度组:100μmol·L^(-1)EGCG孵育24 h后,加入H_2O_2作用12 h。MTT比色法检测细胞活力,流式细胞仪检测细胞凋亡率,Hochest33258染色观察凋亡细胞形态,比色法检测凋亡相关因子caspses-3及caspase-9的表达。结果:EGCG能明显抑制H_2O_2诱导的HLE细胞活力的下降,用不同浓度EGCG处理后,HLE细胞活性分别提高到51.00%±2.37%和63.67%±2.29%,与氧化损伤组(40.33%±2.86%)比较差异具有统计学意义(P<0.05);经不同浓度EGCG处理后,HLE细胞凋亡率分别下降至33.33±3.12%和22.80±1.67%,与氧化损伤组(43.03±2.43%)比较差异具有统计学意义(P<0.05);此外,EGCG还能明显减少H_2O_2所致HLE细胞内caspses-3及caspase-9的表达。结论:EGCG通过抑制caspses-3及caspase-9的表达有效抑制了H_2O_2对HLE细胞的损伤,从而为其用于治疗HLE细胞损伤提供可靠的实验依据。

Objective： To discuss the protective effects and possible mechanisms of EGCG in vitro cultured human les epithelial （HLE） cells induced by oxidative damage. Methods： HLE cells were subcultured, devided into negative control group： cultured with normal culture medium; oxidative injury group：100 μmol·L^-1 H2O2 treated for 12 h; EGCG low dose group： 10 μmol·L^-1 EGCG incubated for 24 h then treated with 100μmol·L^-1 H2O2 for 12 h; and EGCG high dose group： 100 μmol·L^-1 EGCG incubated for 24 h then treated with 100 μmol·L^-1 H2O2 for 12 h. Cell viability were tested by MTT colorimetric detection, apoptosis rate was detected by flow cytometry, apoptotic cell morphology was observed by hochest33258 staining, expression of apoptosis-related factors caspses-3 and caspase-9 were tested by colorimetric detection. Results： EGCG inhibited H2O2-induced cell viability decrease in HLE cells, after treated with different concentrations of EGCG, HLE cells activity increased to 51.00%± 2.37% and 63.67% ±2.29 %, which has statistical significance difference compared with oxidative damage group （40.33%± 2.86%）（P〈0.05）; after treated with different concentrations of EGCG, the apoptosis rate of HLE cells decreased to 33.33 ±3.12% and 22.80± 1.67%, respectively, which has statistical significance difference compared with oxidative damage group （43.03 ±2.43% ） （P〈0.05）; in addition, EGCG also significantly reduced the H2O2-induced expression of caspses-3 and caspase-9 in HLE cells. Conclusion： EGCG effective inhibited H2O2-induced HLE cells damage by inhibiting the expression of caspses-3 and caspase-9, which provide reliable experimental basis for the treatment of injuries in HLE cells.