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目的研究微囊藻毒素-LR(MC-LR)对人肝细胞内超氧阴离子自由基(O2.-)、羟自由基(.OH)水平的影响,探讨不同荧光探针的检测效果。方法调整人肝癌HepG2细胞密度为5×104/mL,分别设终浓度为10,30,100μmol/L的MC-LR染毒组和去铁敏(DFO,1 mmol/L)对照组以及MC-LR(100μmol/L)+DFO(1 mmol/L)染毒组,另设溶剂对照组(0.1%DMSO),培养4 h后应用荧光探针氢化乙啶(HE)和2-[6-(4’-羟基)苯氧基-3H-占吨醇-3-on-9-yl]-苯甲酸(HPF)分别检测细胞内O2.-和.OH含量。结果 30~100μmol/L的MC-LR染毒4 h后细胞内O2.-和.OH含量明显高于溶剂对照组(P<0.05),染毒剂量与.OH含量之间呈现剂量-反应关系,去铁敏和MC-LR联合染毒组的O2.-和.OH含量明显低于100μmol/L MC-LR染毒组(P<0.05)。结论 MC-LR可引起人肝癌HepG2细胞内O2.-和.OH含量升高,用荧光探针HE和HPF能够快速、灵敏地检测细胞内的超氧阴离子自由基和羟自由基水平。
Objective To investigate the effect of microcystin-LR (MC-LR) on the level of superoxide anion radical (OH) and hydroxyl radical (OH) in human hepatocytes and to explore the detection effect of different fluorescent probes. Methods The density of HepG2 cells was adjusted to 5 × 104 / mL. MC-LR and DFO (1 mmol / L) control groups and MC-LR (100μmol / L) and DFO (1 mmol / L), another solvent control group (0.1% DMSO) -hydroxy) phenoxy-3H-tritol-3-on-9-yl] -benzoic acid (HPF) were used to detect intracellular O2.- and .OH content respectively. Results The contents of O2.- and .OH in the cells treated with 30-100 μmol / L MC-LR for 4 h were significantly higher than those in the solvent control group (P <0.05). There was a dose-response relationship between the dose and the OH content , And the content of O2.- and OH in the combined deferoxamine and MC-LR groups was significantly lower than that in the MC-LR group of 100μmol / L (P <0.05). Conclusion MC-LR can induce the increase of O2.- and OH contents in human hepatocellular carcinoma HepG2 cells. Fluorescent probes HE and HPF can rapidly and sensitively detect the intracellular superoxide anion and hydroxyl radical levels.