Effects Of Copper And Nickel Combined Exposure On Oxidative Stress And Endogenous And Exogenous Apoptosis Pathways In Mice Liver

Objective:With the development of society and human industrialization,heavy metals have been widely used in many fields.Heavy metals such as copper are widely used in the manufacture of communication cables,high and low voltage wires and various motors.However,while heavy metals are widely used in industrial development,their pollution and health effects have become important health issues.A previous research team investigating workers exposed to heavy metals in magnesium mines and glass factories found that the content of copper and nickel in urine among workers in both factories increased significantly,indicating that workers in both factories were exposed to copper and nickel.Whether copper-nickel joint exposure interacts with liver oxidative stress and cell apoptosis has not been reported yet.To this end,the Cu-Ni combined exposure mouse model was constructed to observe the interaction of copper and nickel on the oxidative and mitochondrial apoptosis pathway and death receptor apoptosis pathway in mouse liver to investigate the effects of Cu-Ni combined exposure on oxidative stress and endogenous apoptotic pathways in liver injury.Methods:Sixty male Kunming mice were selected,their initial body weights were weighed,and they were randomly divided into 4 groups with 15 mice in each group.The control group drank distilled water,the copper exposure group was 150ppm Cu SO₄,the nickel exposure group was 300ppm Ni SO₄·6H₂O,and the copper-nickel combined exposure group was 150ppm Cu SO₄+300ppm Ni SO₄·6H₂O.Record the weight every week,weigh the liver and calculate the viscera-to-body ratio.Use atomic absorption to detect liver copper and nickel levels.Mouse serum ALT and AST were detected by kit Hepatic sections were observed by HE staining and TUNEL staining The kit detects the oxidative stress indicators of GSH,CAT and T-AOC in mouse liver.Western Blot detects the relative protein expression levels of Bcl-2,Bax,Fas,Fas L,TNFR1,TRADD,FADD,Caspase-8,Cleaved-Caspase-8 in mouse liver.Results:1.The general condition of mice is affected by the combination of copper and nickel:no significant changes in each group.2.After 12 weeks of exposure,the copper content in the liver of the copper group and the copper-nickel combination group was higher than that of the control group and the nickel group（P<0.01）.The nickel content in the liver of the mice in the nickel exposure group and the copper-nickel combined exposure group was higher than that in the control group and the copper group（P<0.01）.3.Toxic effects of combined copper-nickel exposure on mouse liver:HE staining heavy metal exposure group hepatocyte central vein structure is disordered,the gap is enlarged and loose,and inflammatory cell aggregation.The liver coefficient of each exposed group was significantly lower than control group（P<0.05 or P<0.01）,and the ALT of each group was significantly higher than control group（P<0.05 or P<0.01）.The ALT level in the combined exposure group was significantly higher than that in the individual exposure（P<0.05 or P<0.01）.Analysis shows that copper and nickel have no interaction with ALT.The AST of other heavy metal exposure groups increased significantly in the control group（P<0.01）.Factor analysis showed that copper and nickel exposure interacted with serum AST levels in mice.The average value can be drawn that the AST level of the copper-nickel combined exposure group is higher than that of the copper exposure group,but lower than that of the nickel exposure group,indicating that copper antagonizes the effect of nickel on AST,and nickel synergizes the effect of copper on AST.4.Oxidative stress:the GSH and CAT of the combined exposure group decreased significantly compared with the control group（P<0.01）,the T-AOC content of each exposure group decreased significantly compared with the control group（P<0.05 or P<0.01）,and the GSH content of the combined exposure group was significantly lower than that of the copper group（P<0.01）,and other indicators were somewhat different compared to the single exposure group Decrease,but the significance is not obvious.The results of factorial analysis showed that Cu-Ni combined exposure had no interaction with GSH,CAT and T-AOC in the liver of mice.5.Effects of Cu-Ni combined exposure on mouse liver mitochondria and death receptor pathway mediated apoptosis:After 12weeks of exposure,TUNEL staining showed that the number of hepatocyte apoptosis in each heavy metal group was significantly higher than control group,Cu-Ni combined exposure group increased the most.The mitochondrial pathway suggests that the relative expression of Bcl-2 protein in each exposed group was significantly lower than that in the control group（P<0.01）.Bax increased more（P<0.05 or P<0.01）,combined exposure compared to a single exposure group,The relative expression of Bcl-2 protein decreased more（P<0.01）,whereas Bax expression decreased more（P<0.05 or P<0.01）.Factor analysis shows that,Cu and Ni do not interact with Bcl-2 and Bax.Mouse liver death receptor pathway:the relative expression level of liver Fas、Fas L、FADD、TNFR1、TRADD and Cleaved-Caspase-8 protein in exposed group was significantly higher than that in control group（P<0.05 or P<0.01）.The combined exposure and single exposure were significantly higher than other proteins except Fas L（P<0.01）;The expression of liver Caspase-8 decreased significantly in all exposed groups（P<0.01）,combined exposure of copper and nickel was significantly lower than single exposure（P<0.01）.The combined exposure of copper and nickel had no interaction on liver Fas、Fas L、FADD and Caspase-8 in mice.TNFR1,TRADD,and Cleaved-Caspase-8 protein expression levels in mouse liver interacted and showed synergistic effects.Conclusion:1.Cu-Ni compound exposure can damage the structure of liver cells and lead to oxidative stress injury in the liver,which in turn leads to liver injury.2.Cu-Ni combined exposure can induce hepatocyte apoptosis by activating the mitochondrial pathway and death receptor pathway,it also had a synergistic effect on TNFR1,TRADD,and Cleaved-Caspase-8 in the death receptor pathway.