Lipidomics and single-cell RNA sequencing reveal lipid and cell dynamics of porcine glycerol-injured skeletal muscle regeneration model

Intramuscular fat (IMF) infiltration and extracellular matrix (ECM) deposition are characteristic features of muscle dysfunction, such as muscular dystrophy and severe muscle injuries. However, the underlying mechanisms of cellular origin, adipocyte formation and fibrosis in skeletal muscle are still unclear. Pigs were injected with 50 % glycerol (GLY) to induce skeletal muscle injury and regeneration. The acyl chain composition was analyzed by lipidomics, and the cell atlas and molecular signatures were revealed via single-cell RNA sequencing (scRNA-seq). Adipogenesis analysis was performed on fibroblast/fibro-adipogenic progenitors (FAPs) isolated from pigs. The porcine GLY-injured skeletal muscle regeneration model was characterized by IMF infiltration and ECM deposition. Skeletal muscle stem cells (MuSCs) and FAP clusters were analyzed to explore the potential mechanisms of adipogenesis and fibrosis, and it was found that the TGF-β signaling pathway might be a key switch that regulates differentiation. Consistently, activation of the TGF-β signaling pathway increased SMAD2/3 phosphorylation and inhibited adipogenesis in FAPs, while inhibition of the TGF-β signaling pathway increased the expression of PPARγ and promoted adipogenesis. GLY-induced muscle injury and regeneration provides comprehensive insights for the development of therapies for human skeletal muscle dysfunction and fatty infiltration-related diseases in which the TGF-β/SMAD signaling pathway might play a primary regulatory role.