铜、锌对泥鳅细胞DNA损伤及金属硫蛋白表达的影响

Toxicity of Copper Sulfate and Zinc Chloride on DNA Damage and Metallothionein Gene Expression in Loach Cell Lines

采用0、100、200、400、800μmol/L的硫酸铜和0、200、400、800、1600μmol/L的氯化锌分别处理泥鳅鳍细胞系,培养24 h后,用单细胞凝胶电泳检测其DNA损伤情况,同时对金属硫蛋白基因的表达进行研究。试验结果表明,随着两种重金属浓度的增加,泥鳅鳍细胞的DNA损伤程度呈现明显的剂量依赖性。当硫酸铜浓度≥200μmol/L、氯化锌浓度≥400μmol/L时,细胞的拖尾率、彗尾DNA比例、彗星尾长、彗星尾距和Olive尾矩均显著增大(P<0.05)。泥鳅细胞金属硫蛋白基因的表达量随着硫酸铜浓度的升高呈现出短暂稳定后显著下降的趋势,而随着氯化锌浓度的增大呈现出显著上升再显著下降的趋势,表明氯化锌浓度≤400μmol/L时可显著诱导泥鳅鳍细胞系金属硫蛋白的转录表达。综合来看,与锌相比,泥鳅鳍细胞对铜的解毒能力和耐受力更差。因此,铜污染对泥鳅细胞的遗传毒性更加明显。本研究将为探讨重金属对生物的毒性效应及生物学监测提供理论依据。

The effects of copper sulfate and zinc chloride on DNA damage and metallothionein(MT)gene expression were studied in loach fin cell lines in this study.Cells were suffered by 0,100,200,400 and 800μmol/L copper sulfate,and 0,200,400,800 and 1600μmol/L zinc chloride,respectively,for 24 h,and the DNA damage(detected by single cell gel electrophoresis)and MT expression were studied.Results showed that the DNA damage were gradually obvious with the increasing concentrations of copper sulfate and zinc chloride,showing concentration dependent characteristics.When copper sulfate and zinc chloride were greater than or equal to 200μmol/L and 400μmol/L,respectively,the Tailing percent,comet tailing DNA ratio(Tail DNA),comet tail length(L Tail),comet tail distance(TM)and Olive tail moment(OTM)were significantly increased(P<0.05).MT mRNA expression levels were studied by Real-time PCR usingβ-actin as internal reference gene.The MT mRNA expression values were maintained at the level of the control group,and then decreased significantly with the increasing concentration of copper sulfate.Expressed levels of MT in loach cells increased at low concentration of zinc chloride(200 and 400μmol/L)and then decreased significantly with the increasing zinc chloride concentration.These findings indicated that MT mRNA were induced at zinc chloride concentration of less than or equal to 400μmol/L.In general,loach fin cells were more sensitive to copper sulfate than to zinc chloride,and the genetic toxicity was more obviously in the cells exposed to copper.The results of this study will allow us to understand the mechanism of genetic toxicity to loach cells under Cu and Zn exposure and provide some information about physiology regulation and environmental monitoring.