Skeletal muscle ischemia-reperfusion (I/R) injury is a critical clinical condition. AMP-activated protein kinase (AMPK) during IR injury remains unclear. A hindlimb I/R model was established in Sprague-Dawley rats using vascular occlusion. AMPK activator Metformin and AMPK inhibitor Compound C were applied to investigate the effects of AMPK on hindlimb I/R injury. Differentiated C2C12 myotubes were subjected to hypoxia/reoxygenation to mimic I/R injury in vitro. Histopathology, apoptosis, and angiogenesis were observed by Hematoxylin-Eosin, TUNEL staining, and immunohistochemistry. Western blotting and Enzyme-Linked Immunosorbent Assay were performed to assess autophagy and ferroptosis-related indicators. Co-immunoprecipitation validated AMPK-ULK1 interactions. Furthermore, Unc-51-like kinase 1 (ULK1) was inhibited to explore its effects in vivo and in vitro. AMPK activation via Metformin reduced I/R-induced muscle damage, apoptosis, and endothelial dysfunction, while Compound C exacerbated these effects. Metformin upregulated p-AMPK and suppressed ferroptosis in vivo. In C2C12 cells, AMPK overexpression attenuated hypoxia/reoxygenation-induced ferroptosis (increased GPX4 and SLC7A11) and activated autophagy (increased LC3-II/LC3-I and decreased p62). Mechanistically, AMPK phosphorylated ULK1 at Ser555, promoting ATG7/ATG5-dependent autophagy. ULK1 inhibition suppressed autophagy and abolished AMPK's protective effects, exacerbating muscle injury and ferroptosis. AMPK activation protects against skeletal muscle I/R injury by enhancing ULK1/ATG-mediated autophagy and inhibiting ferroptosis.
Li et al. (2026) studied this question.