Aerobic Training Activates Nrf2-mediated Anti-oxidative Effect Of AMPK On Mouse Skeletal Muscle

Objective: AMPK not only plays an important role in the regulation of energy metabolism,AMPK can be activated by ROS,play an important role in the body's anti-oxidative stress response.In this study,cell and animal experiments were used to study the regulation of AMPK on the antioxidant activity mediated by Nrf2 in skeletal muscle of mice under aerobic training.Methods: The experiment of C2C12 cells in mouse skeletal muscle cells was performed in vitro.The effects of AMPK activator AICAR and AMPK inhibitor Compound C were investigated.Fluorescence inverted microscopy qualitative detection of ROS fluorescence intensity of C2C12 cells in each group.Fluorescence colorimetry was used to detect the level of ROS in C2C12 cells.The levels of anti-oxidase m RNA in C2C12 cells in each group were detected by RT-PCR.Western Blot method was used to detect AMPK,AMPK phosphorylation,Nrf2,Nrf2 phosphorylation and antioxidant enzyme protein expression in C2C12 cells.In vivo experiments,8-week-old AMPK?2 transgenic mice(TG mice),AMPK?2 knockout mice(KO mice)and wild-type mice(WT mice)were randomly divided into the quiet group and the training group Group of 10 each.Training group 6 days a week,a daily slope 0,12 m / min treadmill aerobic training 1 hour,a total of 4 weeks.Fluorescence colorimetry was used to detect the level of ROS in skeletal muscle of mice in each group.The levels of antioxidant enzymes m RNA in skeletal muscle of mice in each group were detected by RC-PCR.Western Blot method was used to detect AMPK,AMPK phosphorylation,Nrf2 phosphorylation and antioxidant enzyme protein expression in skeletal muscle cells.Results:(1)compared with the control group,p AMPK?2 /AMPK?2 ratio increased significantly in C2C12 cells of stimulant group;the expression of p Nrf2 protein significantly increased,the expression of NQO1 m RNA,HO-1m RNA and GSR m RNA was significantly increased;the protein expression of SOD1,GCLM,NQO1,HO-1,CAT,GCLc and GSR was significantly increased,and the level of fluorescence intensity of cell was significantly decreased.(2)compared with the control group,t NQO1 m RNA,HO-1m RNA,CATm RNA,SOD1 m RNA,Gpx-1m RNA and GCLcm RNA decreased significantly in the inhibitor group,and the protein expression of NQO1 and GCLM decreased significantly in C2C12 cells,and the level of cell ROS increased significantly.After 4 weeks of aerobic training(3)the expression of p Nrf2 protein in skeletal muscle increased significantly in wild type mice and AMPK?2 expressed high expression of transgenic mice.(4)the expression of HO-1m RNA in skeletal muscle of wild type mice increased significantly,while the expression of Gpx-1m RNA,HO-1m RNA and NQO-1m RNA in skeletal muscle of transgenic mice was significantly increased.Compared with wild rats,the expression of CATm RNA,NQO-1m RNA,HO-1m RNA,Gpx-1m RNA and SOD1 m RNA in the skeletal muscle of AMPK?2 gene knockout rats in the training group decreased significantly.(5)the protein expression of CAT,GSR,HO-1 and SOD1 in skeletal muscle of wild rats significantly increased,AMPK?2 transgenic rat skeletal muscle protein(CAT,Gpx-1,NQO-1)and protein(GCLc,GSR,HO-1,SOD1,SOD2)expression increased significantly.Compared with the training group of wild rats,protein expression of Gpx-1,GSR,HO-1,NQO-1,SOD1,SOD2 and GCLc in transgenic rat skeletal muscle of training group increased significantly.The expression of protein(HO-1 and SOD2)in AMPK?2 gene knockout mice was significantly lower than that in the training group.(6)the expression of ROS in skeletal muscle tissue of wild rat,AMPK?2 transgenic mice and AMPK?2 knockout mice,and no significant difference was found before and after training.Conclusions: The activation of AMPK in aerobic training can promote the expression of p Nrf2 in skeletal muscle,and then increase the expression of antioxidant enzymes and improve the antioxidant capacity of the body.