3-氯-1,2-丙二醇对人胚肾293细胞DNA损伤作用
3-monochloropropane-1,2-diol induces oxidative DNA damage in HEK 293 cells
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摘要: 目的 研究3-氯-1,2-丙二醇(3-MCPD)对人胚肾293(HEK293)细胞DNA损伤作用。方法 以HEK293细胞为靶细胞,3-MCPD设5个剂量组:0,1.25,2.5,5,10 mmol/L,应用单细胞凝胶电泳检测3-MCPD诱导的HEK293细胞DNA损伤;将3-MCPD设4个剂量组:0,2.5,5,10 mmol/L,以2,7-二氢二氯荧光素(DCFH)法测定HEK293细胞内活性氧(ROS)水平;将3-MCPD设4个剂量组:0,1.25,2.5,5 mmol/L,通过免疫组化法测定8-羟基脱氧鸟苷(8-OHdG)在HEK293细胞内的表达水平。结果 与对照组比较,1.25~10 mmol/L的染毒组细胞DNA链断裂,细胞形成彗星样脱尾,尾DNA%明显上升,且呈剂量依赖关系(P<0.05)。2.5~10 mmol/L的染毒组细胞内ROS水平表达升高,在10 mmol/L时明显升高(P<0.05)。1.25~5 mmol/L的染毒组细胞内8-OHdG表达水平上升,在2.5和5 mmol/L时8-OHdG表达明显增强(P<0.05)。结论 3-MCPD导致了HEK293细胞DNA损伤,可能是通过细胞内ROS升高及8-OHdG形成增加所造成的氧化性DNA损伤作用。Abstract: Objective The DNA damage induced by 3-monochloropropane-1,2-diol(3-MCPD)and the underlying mechanisms were studied in HEK293 cells.Methods As target cells,HEK293 cells were treated at the concentrations of 0,1.25,2.5,5,and 10 mM with 3-MCPD at 37℃ for 1 hr.The single cell gelelectrophoresis assay(SCGE)was applied for quantitative analysis of DNA damage which was caused by 3-MCPD.We also measured the level of intracellular ROS by use of the 2,7-dichlorofluorescein diacetate(DCFH-DA)assay when HEK293 cells were treated with different concentrations of 3-MCPD(0,2.5,5,and 10 mM)at 37℃ for 1 hr.The level of oxidative DNA damage was evaluated using immunope roxi-dases taining for 8-hydroxydeoxyguanosine(82OHdG)when HEK293 cells were treated with different concentrations of 3-MCPD(0,1.25,2.5,5 mM)at 37℃ for 3 hr.Results The results showed that 3-MCPD at all tested concentrations (1.25-10 mM)caused a significant increase of DNA strand breaks in HEK293 cells.The numerical values for tail DNA% significantly increased compared to the control(P<0.05).The level of intracelluar ROS was significantly increased when exposed to 10 mM of 3-MCPD(P<0.05).The formation of 82OHdG was significantly increased at the concentration of 2.5-5 mM of 3-MCPD(P<0.05).Conclusion 3-MCPD can induce DNA damage in HEK293 cells.The DNA damage is probably induced by the inc reased level of in tracellular ROS and formation of 8-OHdG,which cause oxidatively generated DNA damage and formation of DNAs trand breaks.
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