硫胺素和核黄素对H_2O_2诱导的DNA氧化损伤的影响

Effects of thiamine and riboflavin on H_2O_2-induced DNA oxidative damage

目的探讨硫胺素和核黄素对H2O2诱导人脐静脉内皮细胞ECV304的DNA氧化损伤的影响。方法于细胞培养液中分别加入硫胺素(终浓度为10、100、500、1000mg/L)或核黄素(终浓度为20、100、300、500nmol/L)预处理24h,给予H2O2(终浓度为25μmol/L)反应30min,采用单细胞凝胶电泳技术(SCGE)分析DNA氧化损伤情况。以不加H2O2、硫胺素和核黄素者为阴性对照组;以加H2O2而不加硫胺素、核黄素者为阳性对照组。结果H2O2诱导ECV304细胞DNA断裂损伤,SCGE分析显示,阳性对照组拖尾细胞率、彗星尾长、尾部DNA含量和Olive尾矩等指标均较阴性对照组升高;经10、100、500mg/L硫胺素或20、100、300nmol/L核黄素预处理后,各DNA损伤指标均较阳性对照组显著下降(P<0.05或P<0.01);经1000mg/L硫胺素或500nmol/L核黄素预处理后,各DNA损伤指标与阳性对照组比较差异无统计学意义(P>0.05)。结论适宜浓度的硫胺素和核黄素能降低H2O2诱导的DNA氧化损伤,高浓度的硫胺素和核黄素不能保护H2O2诱导的DNA氧化损伤。

Objective To explore the effects of thiamine and riboflavin on H2O2-induced DNA oxidative damage in human umbilical vein endothelial cell line ECV304.Methods ECV304 cells were incubated with 10,100,500,1000 mg/L of thiamine or 20,100,300,500 nmol/L of riboflavin for 24 h,and then oxidative damage of cells were induced by 25 mol/L H2O2 for 30 min.DNA damage was detected with single cell gel electrophoresis（SCGE）assay.ECV304 cells incubated without H2O2,thiamine and riboflavin were served as negative controls,and those incubated with H2O2 and without thiamine and riboflavin were served as positive controls.Results H2O2 induced DNA damage,and the indices of percent of DNA damage cells,percent of tail DNA,tail length and Olive tail moment were increased.The indices of cells pretreated with 10,100,500 mg/L of thiamine or 20,100,300 nmol/L riboflavin were significantly decreased（P〈0.05 or P〈0.01）.There was no significant difference in the indices between cells treated with 1000 mg/L of thiamine or 500nmol/L of riboflavin and positive controls（P〉0.05）.Conclusion Proper supplementation of thiamine and riboflavin may decrease H2O2-induced DNA oxidative damage,while excess thiamine and riboflavin supplementation may be harmful to DNA and enhance the susceptibility to H2O2 potentially.