Melatonin Activates AMPK Pathway to Regulate the Regeneration of Slow Muscle Fibers in Skeletal Muscle Injury Repair

The process of skeletal muscle regeneration entails alterations in the relative composition of muscle fiber types, yet the regulatory mechanisms remain incompletely understood. This study aimed to investigate the role of melatonin in regulating the regeneration of slow muscle fibers during skeletal muscle repair and its underlying mechanisms. Using a tibialis anterior muscle frostbite model in 6-8-week-old male C57BL/6J mice, in vivo experiments revealed that intraperitoneal melatonin administration significantly increased Myh7/Myh2 protein expression while reducing Myh1/Myh4 levels. In vitro, melatonin-treated C2C12 myoblasts exhibited elevated oxygen consumption, mitochondrial mass, mitochondrial respiratory chain complexes activity, ATP production, mtDNA content, and membrane potential, alongside reduced LDH activity and ROS levels. Transcriptional upregulation of genes linked to mitochondrial complexes assembly, oxidative phosphorylation, and ATP synthesis was observed. Mechanistically, melatonin activated the AMPK/PGC-1α pathway, as evidenced by Compound C (AMPK inhibitor) pretreatment reversing these effects, decreasing p-AMPK/AMPK ratios, PGC-1α, and slow fiber markers (Myh7/Myh2), while increasing ROS and fast fiber marker (Myh4). The results indicate that melatonin facilitates the formation of slow-twitch fibers during muscle repair by augmenting mitochondrial function through the AMPK/PGC-1α signaling pathway. Consequently, these findings imply that melatonin improves mitochondrial function via AMPK/PGC-1α signaling, thereby promoting the regeneration of slow muscle fibers and facilitating the repair of skeletal muscle damage.