P0109The role and mechanism of aerobic exercise on improving DSS-induced intestinal inflammation by regulating mitophagy through AMPK-SIRT1-FOXO1 pathway

Abstract Background Studies have found that inflammatory bowel disease (IBD) is associated with mitochondrial dysfunction. Exercise is an effective behavioral therapy to promote mitochondrial health. This study explores the therapeutic value and potential mechanism of aerobic exercise in improving chronic intestinal inflammation. Methods Forty C57BL/6J mice were randomized into control, swim, DSS-colitis, and DSS-colitis+swim groups (n = 10). Chronic colitis was induced in DSS groups via cyclic 2% DSS administration. Swim groups underwent 8 weeks of aerobic swimming training. Colitis severity, inflammatory factors, oxidative stress, and mitochondrial function (ATP, membrane potential) were assessed. Mitophagy and mitochondrial damage were examined by electron microscopy. Protein levels of PINK1, Parkin, LC3B, AMPK, SIRT1, and FOXO1 were analyzed by Western blot/immunofluorescence. In vitro, an inflammatory model was established using HT-29 cells. AICAR, an AMPK agonist, was used to mimic aerobic exercise, while EX527 served as a SIRT1 inhibitor. Protein levels of AMPK, SIRT1, FOXO1, and mitophagy-related markers were analyzed by Western blot and immunofluorescence, along with assessments of inflammatory factors and oxidative stress. Results Aerobic exercise ameliorated DSS-induced colitis in mice, attenuating weight loss, colon shortening, splenomegaly, tissue injury, and inflammatory cell infiltration, while reducing systemic and colonic inflammatory factor expression. It also alleviated colonic oxidative stress by decreasing ROS and MDA levels and enhancing T-AOC, SOD, and GSH-PX activities. Furthermore, aerobic exercise improved mitochondrial membrane potential and ATP synthesis, restoring mitochondrial function. It enhanced mitophagy, evidenced by increased mitochondrial autophagosomes and upregulation of PINK1, Parkin, and LC3B proteins. The AMPK pathway was activated, with elevated SIRT1 and FOXO1 protein levels. The results of in vitro experiments showed that AMPK agonist AICAR improved the level of inflammation, oxidative stress damage and mitophagy in the single-layer cell inflammation model constructed by HT-29 cell line. After inhibition of SIRT1 activity, the protective effect was weakened, the level of cellular oxidative stress increased, the level of mitophagy decreased, and the level of inflammation increased again. Conclusion Aerobic exercise plays a therapeutic role in IBD by activating AMPK-SIRT1-FOXO1-mediated mitophagy, timely clearing damaged mitochondria and reducing the damage effects of oxidative stress on animals and cells. Conflict of interest: Dr. Xiao, Fang: No conflict of interest Xu, Jiaxin: No conflict of interest Li, Dongyan: No conflict of interest Liu, Haoying: No conflict of interest Shen, Yinxian: No conflict of interest Li, Junhua: No conflict of interest Chen, Liping: No conflict of interest Ke, Xiaoli: No conflict of interest Li, Juan: No conflict of interest