Metformin Plays A Protective Effect On D-galactose-induced Aging Mice Through Parkin-mediate Mitophagy

Objective:Metformin(Met)was administered during the aging of mice induced by D-galactose(D-gal),in addition,mitochondrial autophagy inhibitor 3-methyladenine(3-MA)was also used in a part of mice.The study observed whether Met prevents mice from being susceptible to brain and liver during aging by activating Parkin-mediate mitochondrial autophagy.Methods:A total of 32 female 7-month-old Kunming mice were randomly assigned into 4groups(n=8)after 2 weeks of acclimatization: control group(Ctrl group),model group(D-gal replicative aging group,D group),Metformin administration group(replicative aging model given with Met,DM group)and mitochondrial autophagy inhibition group(induced aging mice given with both Met and mitochondrial autophagy inhibitor 3MA,DMM group).After consecutive administration for 16 weeks,the following tests were conducted:1.Behavioral experiments(Morris water maze and shuttle box)were used to test the learning and memory ability of mice in each group,and the effects of Met on learning and memory of mice were observed.2.Pathological methods(HE staining)were used to detect the histopathological sections of hippocampus and liver tissues in each group of mice,and the effects of Met on the morphological structure of hippocampus and hepatocytes were assessed.3.The levels of glutathione(GSH)and malondialdehyde(MDA)in brain and liver tissues of mice in each group were measured using oxidative stress assay kits,so as toassess the effects of Met of the level of oxidative stress in mice’s brain and liver tissues.4.Flow cytometry was used to detect the mitochondrial membrane potential in the liver of mice in each group,and the effects of Met on the mitochondrial membrane potential in the liver were also observed.5.Western blot was used to detect the expression of mitochondrial autophagy-related proteins in the brain and liver of mice in each group: Parkin,LC3-II/I and p62,so as to figure out the effects of Met on the levels of mitochondrial autophagy-related protein in mice’s brain and liver tissues.Results:1.The results of Morris water maze test showed that Met can significantly reduce the escape latency of mice and increase the swimming time of the mice in the target quadrant;shuttle box experiments showed that Met can significantly increase the frequency of active avoidance in mice;2.The results of HE staining in hippocampus and liver tissue in mice suggested that Met can reduce the pathological damage in dentate gyrus and hepatocyte to a certain extent;3.The detection results of oxidative stress level suggested that Met can increase the level of GSH and decrease the content of MDA in brain and liver tissues of mice.4.The results of flow cytometry suggested that Met can maintain the mitochondrial membrane potential and inhibit mitochondrial depolarization;5.The results of Western blot showed that Met can increase the level of Parkin and LC3-II/I proteins in mitochondria of brain and liver,and reduce the expression of p62 protein.Conclusion:Exogenous administration of Met is able to activate the Pakrin-mediated mitphagy in brain and liver tissues of mice,and remove their damaged or dysfunctionalmitochondria,stabilize mitochondrial membrane potential,inhibit mitochondrial depolarization,and reduce oxidative damage.Met possesses a function to enhance the ability of learning and memory in mice,thereby bringing a protective effect to the brain and liver tissues of mice during their aging process.