Updated project metadata. Sepsis remains a leading cause of mortality and long-term disability, with survivors frequently developing intensive care unit–acquired weakness (ICU-AW) as part of post-intensive care syndrome. To identify a nutritional therapy for ICU-AW, we investigated the mechanisms underlying sepsis- induced skeletal muscle dysfunction using a cecal slurry-induced sepsis mouse model. Although body weight and skeletal muscle mass recovered 14 days after sepsis induction, muscle strength remained impaired, accompanied by persistent mitochondrial abnormalities. Transcriptomic analysis revealed that the pathways termed the ‘sirtuin signaling pathway’ and ‘mitochondrial dysfunction’ significantly enriched with downregulation of Sirt3 which is a major mitochondrial nicotinamide adenine dinucleotide (NAD⁺)-dependent deacetylase. Biochemical analyses confirmed increased acetylated lysine of mitochondrial proteins in septic muscle tissue. Among these proteins, mass spectrometry identified complex I subunits as candidate substrates of Sirt3. Knockdown of Sirt3 in C2C12 myotubes impaired mitochondrial respiration, whereas treatment with β-nicotinamide mononucleotide (β-NMN) partially rescued energy production. In vivo, acute-phase administration of β-NMN preserved mitochondrial morphology and restored muscle strength without altering muscle mass. These findings demonstrate that sepsis induces mitochondrial dysfunction and persistent muscle weakness through Sirt3 downregulation, and highlights β-NMN supplementation as a promising NAD⁺-targeted therapeutic strategy for mitigating ICU-AW.