The Effect of Leucine Supplementation on Inflammatory and Myogenic Genes at the Onset of Skeletal Muscle Regeneration in Aged Male Mice

Skeletal muscle regeneration supports the repair of damaged muscle tissue by coordinating the inflammatory responses following injury. However, with aging, the capacity for muscle regeneration declines, resulting in slower recovery from injuries in older individuals. Aging muscle has a prolonged inflammatory response during regeneration which contributes to an impaired regenerative response. Leucine, a branched-chain amino acid, has shown considerable ability to reduce inflammation associated with muscle injury in both young and older subjects. Nevertheless, the impact of leucine supplementation on age-related loss of muscle regenerative capacity remains insufficiently explored and warrants further investigation. The purpose of this study is to determine the effect of leucine supplementation on inflammatory and myogenic responses at the onset of muscle regeneration in the aged male mice. We hypothesized that leucine supplementation would attenuate inflammatory responses and enhance myogenesis at the onset of regeneration in aged male mice. Aged Male C57/BL6 mice (24 months) were divided into four experimental groups: uninjured, uninjured + leucine, injured, and injured +leucine. To induce muscle damage, bupivacaine was injected into the tibialis anterior (TA) muscles of mice, while the uninjured groups received phosphate-buffered saline (PBS) injections as a control. Following injection, the leucine groups were given drinking water with 1.5% leucine. Three days after injection, the TA was collected, and quantitative PCR was performed to analyze gene expression. A two-way ANOVA was conducted, and significance was established at p < 0.05.The ratio of TA mass to body weight (BW) revealed no significant differences (p > 0.05) across groups. A significant main effect was observed in injured groups, with a 15-fold increase in cyclin D1(p = 0.0064), a ~1.5-fold increase in MyoD (p = 0.0306), and a ~5-fold increase in MyoG (p = 0.0084), mRNA abundance compared to uninjured groups independent of leucine supplementation. No significant differences (p > 0.05) were observed in IL-1β and IL-6 mRNA abundance among the experimental groups. However, a significant main effect was observed in injured groups, with ~6-fold increase in TNF-α (p = 0.0306) mRNA abundance compared to uninjured groups independent of leucine supplementation. Genes related to myogenesis were upregulated in aged male mice despite leucine supplementation. However, inflammatory genes that are typically upregulated during regeneration were not, which could contribute to the delayed regenerative response associated with aged skeletal muscle. Leucine supplementation did not alter these responses. Further research is warranted to determine the efficacy of leucine supplementation as a therapeutic. This study was funded by the Arkansas Biological Institute, the major research component of the Arkansas Tobacco Settlement Proceeds Act of 2000. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.