Regulation Of Aerobic Exercise-activated AMPK Pathway Diet-induced Skeletal Muscle Fibrosis In Mice

Purpose Aerobic exercise indirectly activates AMPK to improve insulin resistance,reduce blood sugar and reduce lipid accumulation.Continuous skeletal muscle fibrosis in diabetic patients is associated with impairment of muscle repair,growth,and insulin sensitivity.We intends to explore whether aerobic exercise can improve insulin resistance through the AMPK pathway to reduce extracellular matrix deposition-induced fibrosis and explore new therapeutic targets for improving the health of obese or diabetic patients through high-fat diet-induced mouse models.Methods Large male C57bl/6J mice aged 6-8 weeks in 30 mice.Mouse feeding conditions were SPF grade.After 1 week of adaptive feeding,the mice were randomly divided into the general diet group(Ctr group,n=10)and the high-fat diet group(HF group,n=20).The two groups were fed simultaneously for 16 weeks of common and high-fat feed.All mice were tested for fasting glucose and glucose tolerance at 14 and16 weeks,and the results were recorded to test the success of diabetes modeling.Since then,the mice in the Ctr group have continued to feed,and the mice in the HF group have been randomly divided into the high-fat group(HF group)and the aerobic group(HF+E group).Each group of 10 mice have continued high-fat diet and the aerobic group has been given eight weeks of aerobic platform training intervention.After 8 weeks of aerobic exercise,the quadriceps and gastrocnemius tissues were collected.HE staining of quadriceps femoris and sirius staining were performed to observe tissue morphology and fibrosis degree.Western-blot was used to detect the expression of COL1,COL3,TGF?,Samd3,p-Samd3,AMPKa,p-AMPKa,MMP9,GLUT4 and PGC-1a in gastrocnemius tissues.Results1.Changes in body weight A model of obese mice was established after feeding for 16 weeks from 6 to 8weeks.Both general and high-fat diet mice had increased body weight,but the growth of mice in HF group was significantly greater than that in Ctr group(p <0.05).After eight weeks of exercise,the trend of weight gain in the Ctr group was stable,while the HF group continued to grow,but the HF+E group decreased slowly(p<0.05).2.Changes in fasting blood glucose There was no difference in fasting blood glucose in each group at the early stage of feeding,but in the later,the Ctr group was slightly lower than that in the HF group,but there was no significant difference.After 8 weeks of exercise,the HF group was slightly higher than the HF+E group and the Ctr group,but there was no significant difference.3.Changes in fasting glucose tolerance When modeling was completed,fasting glucose tolerance test(GTT)in HF group was significantly higher than that in Ctr group,and the area under the curve(AUC)was higher than that in Ctr group(p<0.05),indicating a significant decrease in insulin sensitivity in high-fat mice.After eight weeks of exercise,the GTT and AUC in the HF+E group were significantly lower than those in the HF group(p<0.05),but the GTT and AUC in the Ctr group were also significantly lower than those in the HF+E group(p<0.05),indicating that aerobic exercise did improve insulin resistance due to a high-fat diet,but short-term exercise did not return to normal insulin levels.4.Morphological changes of HE stained cells After he staining,compared with the Ctr group,the skeletal muscle cells of mice in the HF group were enlarged(p <0.05).The order of cells in the HF group was disordered,and the cells were also larger than those in the HF+E group,with a significant difference(p <0.05).5.Fibrosis changes after Sirius Red staining After Sirius red staining,it can be observed that the collagen deposition of mouse skeletal muscle cells in the HF group was significantly different from that in the Ctr group(p <0.05).Compared with the HF group,cell fibrosis was significantly reduced in the HF+E group(p<0.05),indicating that aerobic exercise could reverse skeletal muscle fibrosis caused by a high-fat diet.6.Protein changes after High-Fat diet modeling and aerobic exercise Compared with the Ctr group,western blot test results showed that the glucose transporter GLUT4 expression in the HF group was significantly reduced and significantly different(p<0.05).The expression of the related fibrosis protein COL1,COL3 was significantly increased(p<0.05),the expression of the upstream regulatory protein matrix metalloproteinase MMP9 protein of fibrosis was decreased,and the expression of TGF?;and phosphorylated signaling protein p-samd3 was increased.The amp-dependent protein kinase AMPKa pathway also changed,and the expression of phosphorylated AMPKa protein and peroxisome proliferator activator receptor gamma co-activator PGC-1a protein increased significantly.After 8 weeks of aerobic exercise,western blot test of HF+E group compared with HF group showed that GLUT4 protein expression in HF+E group was significantly higher and significantly different(p<0.05),whereas fibrosis protein COL1 and COL3 expression were all decreased.The expression of MMP9 protein was significantly increased(p<0.05),the expression of p-Smad3 protein was significantly decreased(p<0.05).Conclusion High-fat dietary feeding can cause insulin resistance and fibrosis of skeletal muscle in C57 mice;aerobic exercise can improve the TGF?/Smad3 pathway in skeletal muscle of obese mice with insulin resistance and affect AMPK-related proteins.