Combined Histidine and Proline Supplementation (HISPRO) Enhances Oxidative and Mitochondrial Function in Skeletal...

Amino acids are key regulators of metabolism, their coordinated effects on skeletal muscle signaling and metabolic remodeling under physiological conditions remain incompletely understood. Here, we investigated whether 14 weeks of combined histidine and proline supplementation (HISPRO; 700 mg/kg) enhances skeletal muscle function through metabolic reprogramming in normal ICR mice. HISPRO significantly improved muscle performance compared with the control group, including grip strength, rota-rod, and treadmill. Histological and biochemical analyses revealed a shift toward oxidative muscle phenotype compared with the control group, with larger muscle fibers and succinate dehydrogenase-positive fibers. Consistently, HISPRO promoted mitochondrial biogenesis and oxidative metabolism compared with the control group, as evidenced by upregulation of mitochondrial regulatory genes, mitochondrial DNA copy number, citrate synthase activity, and oxidative phosphorylation (OXPHOS) complex levels in skeletal muscles. Mechanistically, HISPRO was associated with activation of the SIRT1-PGC1α-AMPK signaling axis compared with the control group, as evidenced by increased Nampt and Nmnat1 expression, an elevated NAD⁺/NADH ratio, and enhanced AMPK phosphorylation. SIRT1 inhibition markedly attenuated HISPRO-induced increases in mitochondrial biogenesis markers but did not fully suppress OXPHOS protein expression, suggesting the involvement of both SIRT1-dependent and -independent mechanisms. Notably, HISPRO also improved muscle function in dexamethasone-induced muscle atrophy model. It restored mitochondrial biogenesis and function, and suppressed atrophy-related markers compared with the dexamethasone-treated group. HISPRO may contribute to improving muscle quality through coordinated metabolic regulation and could represent a complementary nutritional for supporting muscle metabolic health.