Purpose: This study aimed to investigate the effects of downhill treadmill running on mitochondrial structure/function and expression levels of mitophagy-related proteins in rat skeletal muscle.
Methods: A total of 48 male adult Sprague-Dawley rats were randomly divided into a control group (C, n = 8) and an exercise group (E, n = 40). Rats in the E group were exercised on a treadmill down a 16° decline at 16 m·min for 90 min and were further divided into 0 h (E0), 12 h (E12), 24 h (E24), 48 h (E48), and 72 h (E72) postexercise subgroups (n = 8 each). At each time point, the soleus muscle was collected under full anesthesia. Mitochondrial ultrastructural changes in skeletal muscle were observed by a transmission electron microscope. The content of quantitative enzyme citrate synthase and the activities of mitochondrial respiratory chain complex II and complex IV were measured by enzyme-linked immunosorbent assay. Protein expressions of skeletal muscle cytochrome c oxidase subunit 1 (COX1), PTEN-induced putative kinase 1 (PINK1), and mitochondrial Parkin microtubule-associated protein 1 light chain 3 (LC3) were determined by Western blot. Mitochondrial colocalizations with Parkin, ubiquitin (Ub), p62/sequestosome 1 (p62), and LC3 were measured by the immunofluorescence double labeling technique.
Results: After downhill treadmill running, the skeletal muscle mitochondrial structure changed dramatically, and a large amount of mitophagosomes were observed; the citrate synthase content and complex II activity were significantly lower (P < 0.05), whereas complex IV activity and COX1 protein level remained unchanged; the expression levels of PINK1, Parkin, Ub, p62, and LC3 were significantly higher than those in the C group (P < 0.05 or P < 0.01).
Conclusion: A session of downhill treadmill running activated the PINK1/Parkin pathway and facilitated mitochondrial colocalizations with Ub, p62, and LC3, causing mitophagy and mitochondrial damage within the skeletal muscle.