石决明提取液对人晶状体上皮细胞氧化损伤的保护作用

摘要

目的 研究石决明提取液对氧化损伤的人晶状体上皮细胞(HLEC)的作用.方法 实验研究.采用双氧水(H2O2)干预体外培养HLEC氧化损伤模型,同时加入不同浓度(0.001％、0.01％、0.1％、0.3％)的石决明提取液,分成空白对照组、阳性对照组即H2O2组和不同浓度的石决明干预组,于1、3、5 d时用CCK-8检测各组HLEC的活力,倒置相差显微镜下观察细胞增殖和形态改变,3d后用化学比色法检测超氧化物歧化酶(SOD)、还原型谷胱甘肽(GSH)和丙二醛(MDA)的水平.采用单因素方差分析进行样本均数间的多重比较,组间两两比较采用LSD-t检验.结果 不同时间点时各实验组间HLEC细胞活力的吸光度(A)值活力存在变化,第1、3、5天各实验组间HLEC细胞活力的A值分别是:空白对照组0.88、1.28、1.32；阳性对照组0.73、1.02、1.06；0.001％石决明提取液组0.73、1.03、1.06;0.01％石决明提取液组0.76、1.10、1.13;0.1％石决明提取液组0.79、1.22、1.21；0.3％石决明提取液组0.79、1.21、1.21；组间比较差异具有统计学意义(1d时F=23 922.42,p =0.05;3 d时,F=120 605.86,P＜0.05；5d时,F=150 939.45,P＜0.05).H2O2使细胞的活力降低,石决明提取液提高其活力,且在一定的时间和浓度范围内具有依赖性,其中第3天时0.1％石决明提取液组HLEC细胞活力的A值为最高(1.22).H2O2组损伤后,HLEC中SOD、GSH的水平降低(SOD 158.05 U/mgprot、GSH 15.05 mg/1000 mgprot)、而MDA量增高到18.11 nmol/mgprot,石决明提取液组使氧化损伤所致HLEC中抗氧化水平提高(SOD 188.64 U/mgprot、GSH 21.05 mg/ 1000 mgprot),脂质过氧化物升高有下降(MDA 14.16 nmol/mgprot),组间比较差异均具有统计学意义(P＜0.05).改变具有统计学意义(SOD:F=983.04,P＜0.05; GSH:F =444.44,P＜0.05; MDA:F=830.52,P＜0.05);阳性对照组中可见细胞核染色质浓缩、聚集,石决明组中细胞染色质的聚集有不同程度的减轻.结论 石决明提取液对体外培养HLEC氧化损伤具有保护作用,可提高细胞内抗氧化水平,减少有害产物的生成.%Objective To study the influence of haliotidis extractive on the oxidative damage in the human lens epithelial cells cultured in vitro.Methods Experimental study.Cultured human lens epithelial cells in vitro were intervened with hydrogen peroxide caused oxidative damage model,at the same time added different concentrations of concha haliotidis extractive.With control experiment research cells were divided into the blank control group,positive control group hydrogen peroxide group and hydrogen peroxide and different concentrations of concha haliotidis group,and on the first,third,fifth day the activity of Cultured human lens epithelial cells were detected with Cell Counting Kit-8 (CCK-8),cellular proliferation and morphological changes were observed with interred phase contrast microscope,and then on the third day chemical colorimetric were used to detect the homogenates superoxide dismutase(SOD),glutathione(GSH) and malondialdehyde(MDA)level.Results (1)At different time points there were variations between the activity of HLEC in each experimental group,Among each experimental group HLEC OD value of the cell vitality at 1 d,3 d,5 d,respectively were blank control group:0.88,1.28,1.32 ;Positive control group:0.73,1.02,1.06; 0.001％ concha haliotis extract group:0.73,1.03,1.06; 0.01％ concha haliotis extract group:0.76,1.10,1.13 ; 0.1％ concha haliotis extract group:0.79,1.22,1.21 ; 0.3％ concha haliotis extract group:0.79,1.21,1.21 ; the difference between groups was statistically significant (P ＜ 0.05) (1 d,F=23922.42,P＜0.05;3 d,F=120605.86,P＜0.05;5 d,F=150939.45,P＜0.05).H2O2 made the vitality of the cells reduce,concha haliotidis enhance its vitality,and in a certain range of time and concentrations there was dependence,with which the third day and 0.1％ was the best.(2)After adding H2O2,the SOD and GSH level of HLEC reduced,(SOD 158.05 U/mgprot,GSH 15.05 mg/gprot) but MDA increased to 18.11 nmol/mgprot,concha haliotidis groups made the increase of antioxidant level (SOD 188.64 U/mgprot,GSH 21.05 mg/1000 mgprot)and the decrease of lipid peroxidation in oxidative damaged HLECs(MDA 14.16 nmol/mgprot),change had a statistical significance (P ＜ 0.05) (SOD:F =983.04,P ＜0.05; GSH:F =444.44,P ＜0.05; MDA:F =830.52,P ＜0.05).(3) The chromatin of the positive control group concentrated and aggregated obviously,the aggregation of chromatin in coneha haliotidis group lightened.Conclusion The concha haliotidis can protect the cultured human lens epithelial cells in vitro which are oxidative injured,increased intracellular antioxidant levels,reduce the generation of hazardous products.