| Objective: Methylmercury(MeHg)is a highly neurotoxic,environmentally accumulating pollutant that can cross the blood-brain barrier and cause serious damage to the central nervous system.Mitochondria are the target organelles of MeHg,and MeHg can induce apoptosis by increasing ROS production and decreasing mitochondrial membrane potential,leading to mitochondrial damage.Meanwhile,QE is also an effective activator of SIRT1,and the increased activity of SIRT1 can alleviate mitochondrial damage.Therefore,whether QE can alleviate mitochondrial damage induced by MeHg and thus alleviate neurotoxicity remains to be studied.This study aims to clarify whether QE pretreatment can antagonize mitochondrial damage by activating SIRT1/PGC-1α pathway,thereby antagonizing cell apoptosis induced by MeHg,and provide experimental basis for the prevention and control of MeHg neurotoxicity.Methods: In this experiment,the mice were randomly divided into control group,MeHg exposure group,50 mg/kg QE group,12.5 mg/kg QE+MeHg group,25 mg/kg QE+MeHg group and 50 mg/kg QE+MeHg group according to their body weight by establishing the animal model of MeHg exposure and QE pretreatment.There were 14 mice in each group,84 mice in total,half male and half female.The control group was given physiological saline by gavage,the pretreatment group was given QE gavage treatment,and MeHg gavage was administered for 4 hours after QE gavage treatment,and the relevant indicators were tested.Cold atomic absorption method was used to detect the mercury content in the cerebral cortex of mice,gait test and claw grip test were used to detect the behavioral changes of mice,flow cytometry was used to detect ROS and apoptosis of cerebral cortex cells,JC-1 staining was used to detect mitochondrial membrane potential,the kit was used to detect SIRT1 deacetylase activity.The immunoprecipitation method was used to detect the acetylation level of PGC in the cerebral cortex of mice in each group,the RT-qPCR method was used to detect the mRNA level of SIRT1 and PGC-1α in the cerebral cortex of mice in each group,the qPCR method was used to detect the change of mitochondrial DNA copy number in each group,the double fluorescence staining method was used to detect the co-localization level of SIRT1 and PGC-1α,the co-immunoprecipitation method was used to detect the interaction between SIRT1 and PGC-1α protein in each group,and the Western blot method was used to detect SIRT1 and PGC-1α in the cerebral cortex of mice in each group,The relative expression level of mitochondrial biogenesis related proteins NRF1,TFAM,fusion related proteins OPA1,MFN1,fission related proteins DRP1,FIS1 and apoptosis-related proteins Bcl-2,Bax,cleaved caspase-3 and Cyt C.Results: After four weeks of MeHg exposure and QE pretreatment,the body weight of mice in the MeHg group decreased significantly,while the body weight of mice in the QE pretreatment group increased significantly.Mercury content in the cerebral cortex of mice treated with MeHg was significantly higher than that of the control group.The results of gait experiment showed that mice in the MeHg group had slower movement and decreased gait stability compared with mice in the control group.In the claw grip experiment,the claw grip of mice in the MeHg group was significantly weakened,which was alleviated by QE treatment.Compared with the control group,the ROS level of cerebral cortex cells in MeHg group was significantly increased,mitochondrial membrane potential was significantly decreased,mitochondrial DNA copy number was decreased,resulting in mitochondrial damage,and the apoptosis rate of cerebral cortex cells was increased.In addition,the levels of SIRT1 and PGC-1α protein and mRNA in MeHg group were significantly decreased,the activity of SIRT1 deacetylase was decreased,the deacetylation ability was weakened,and the acetylation level of PGC-1α was significantly increased.Dual fluorescence staining showed that the colocalization levels of SIRT1 and PGC-1α were weakened after MeHg exposure,and Pearson correlation coefficient was significantly reduced compared with the control group,while QE pretreatment could improve the co-localization level,and Pearson correlation coefficient was gradually increased.The co-immunoprecipitation results suggested that,compared with the control group,MeHg exposure significantly inhibited the interaction between SIRT1 and PGC-1α protein.Western Blot results showed that the relative expression levels of SIRT1,PGC-1α,mitochondrial biogenesis related proteins NRF1 and TFAM decreased.The relative expression levels of fission related proteins DRP1 and FIS1 were increased,and the relative expression levels of fusion related proteins OPA1 and MFN1 were decreased.The relative expression levels of apoptosis-related proteins Bax,cleaved caspase-3 and CytC were increased,and the relative expression levels of anti-apoptotic protein Bcl-2 were decreased.Compared with MeHg group,after QE pretreatment,SIRT1 deacetylase activity,SIRT1 mRNA and protein expression levels were significantly increased,the acetylation level of PGC-1α was significantly decreased,and the interaction between SIRT1 and PGC-1α protein was gradually increased,and the ROS level of cerebral cortex cells was decreased.Mitochondrial membrane potential increased,mitochondrial DNA copy number increased,mitochondrial damage was recovered,and the apoptosis rate of cerebral cortex cells decreased.The expression level of biogenesis related proteins increased,the expression level of fission related proteins decreased,and the expression level of fusion related proteins increased.The expression of apoptosis-related proteins decreased and the expression of anti-apoptosis-related proteins increased.These results suggest that QE pretreatment can antagonize mitochondrial damage induced by MeHg exposure and inhibit mitochondrial apoptosis pathway.Conclusion: QE can promote mitochondrial biogenesis and regulate mitochondrial dynamic balance by activating SIRT1/PGC-1α pathway,thus antagonizing mitochondrial damage and cell apoptosis induced by MeHg exposure. |
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