NCOA4-Mediated Ferritinophagy Contributes to Iron Overload-Driven Ferroptosis of Senescent Myoblasts in Mice

Sarcopenia is an age-related pathological syndrome characterized by progressive and generalized loss of skeletal muscle mass and function, with muscle atrophy representing its cardinal pathological hallmark. Ferroptosis, an iron-dependent regulated cell death, has been implicated in the pathogenesis of muscle atrophy; however, the precise role of iron dysregulation in sarcopenia remains incompletely understood. In the present study, we identified ferroptosis in D-galactose (D-gal)-induced senescent myoblasts, as evidenced by elevated intracellular iron levels and lipid peroxidation, increased malondialdehyde (MDA) content, and upregulated expression of prostaglandin endoperoxide synthase 2 (PTGS2), 4-hydroxynonenal (4-HNE), and long-chain acyl-CoA synthetase 4 (ACSL4), accompanied by diminished glutathione peroxidase 4 (GPX4), SLC7A11 (xCT), and glutathione (GSH) levels, as well as pronounced mitochondrial damage. Notably, treatment with the iron chelator deferoxamine (DFO) significantly attenuated senescence-associated ferroptosis. Moreover, D-gal-induced senescence in myoblasts was accompanied by reduced ferritin expression and elevated nuclear receptor coactivator 4 (NCOA4) levels, both of which were reversed by autophagy inhibition with 3-methyladenine (3-MA) or NCOA4 knockdown, suggesting that NCOA4-mediated ferritinophagy is involved in senescence-induced iron overload and ferroptosis. Furthermore, senescent myoblasts exhibited increased reactive oxygen species (ROS) generation and mitochondrial impairment, which were attributed to cytosolic iron overload-mediated upregulation of mitoferrin 2 (Mfrn2), thereby promoting mitochondria iron import. Finally, pharmacological inhibition of iron overload or ferroptosis by DFO or ferrostatin-1 (Ferr-1) effectively ameliorated skeletal muscle atrophy and functional decline in aged sarcopenia mice. Collectively, these findings elucidate the mechanistic basis of sarcopenia and highlight potential therapeutic avenues targeting iron dysregulation and ferroptosis.