Age-related muscle atrophy (sarcopenia), characterized by reduced skeletal muscle mass and muscle strength, is becoming increasingly prevalent worldwide, which is especially true for older people, and can seriously damage health and quality of life in older adults. This study aims to investigate the beneficial effects of 5'-cytimidine monophosphate (CMP) on HO-induced muscular atrophy in C2C12 myotubes. C2C12 myotubes were treated with HO in the presence and absence of CMP and the changes in the anti-oxidation, mitochondrial functions, and expression of sarcopenia-related proteins were observed. Immunofluorescence analysis showed that CMP significantly increased the diameter of myotubes. We found that CMP could increase the activity of antioxidant enzymes and improve mitochondrial dysfunction, as well as reduce inflammatory cytokine levels associated with sarcopenia. RNA-seq analysis showed that CMP could relieve insulin resistance and promote protein digestion and absorption. Western blot analysis further confirmed that CMP could promote the activation of the IRS-1/Akt/S6K signaling pathway and decrease the expression of MuRF1 and Atrogin-1, which are important markers of muscle atrophy. The above results suggest that CMP protects myotubes from HO-induced atrophy and that its potential mechanism is associated with activating the IRS-1/Akt/S6K pathway to promote protein synthesis by improving mitochondrial dysfunction and insulin resistance. These results indicate that CMP can improve aging-related sarcopenia.