Quantitative Proteomics Revealed the Molecular Regulatory Network of Lysine and the Effects of Lysine Supplementation on Sunit Skeletal Muscle Satellite Cells

Stimulating skeletal muscle satellite cells (SMSCs) with amino acids improves their proliferation and differentiation, enhancing skeletal muscle mass, thereby increasing lean meat rate. This study explored lysine (Lys)'s effects on SMSCs and their protein profiles in Sunit sheep. SMSCs were successfully isolated, assessing their survival and proliferation after Lys stimulation at varying concentrations using the CCK-8 assay. Western blotting revealed Lys-induced changes in myogenic differentiation protein expression, while immunocytochemistry detected α-Actinin and Myostatin within the SMSCs. TMT proteomics identified differentially expressed proteins, which underwent functional and interaction analyses, with RT-qPCR validating the corresponding gene expression. This study revealed that 4 mmol/L of Lys significantly boosted SMSC proliferation. A 24 h stimulation with this concentration reduced Myostatin expression, and increased MYOD1 and α-Actinin levels in the SMSCs. A proteomic analysis identified 577 differentially expressed proteins, primarily associated with lipoblast differentiation and muscle development, as highlighted by the GO enrichment analysis. A pathway analysis further demonstrated these proteins' involvement in the autophagy-lysosome and NOD-like receptor signaling pathways. Lys enhances SMSC proliferation, differentiation, and adipogenesis in Sunit sheep, exhibiting antioxidant properties and supporting muscle stability and amino acid metabolism. It may also have anti-inflammatory, anti-pyroptotic, and proteolysis-inhibitory effects, offering insights into muscle growth mechanisms through amino acid supplementation in ruminants.