Integrating spatial transcriptomics and single-nucleus RNA-seq revealed the specific inhibitory effects of TGF-β on intramuscular fat deposition.

Intramuscular fat (IMF) is a complex adipose tissue within skeletal muscle, appearing specially tissue heterogeneous, and the factors influencing its formation remain unclear. In conditions such as diabetes, aging, and muscle wasting, IMF was deposited in abnormal locations in skeletal muscle, damaged the normal physiological functions of skeletal muscle. Here, we used Longissimus dorsi muscles from pigs with different IMF contents as samples and adopted a method combining spatial transcriptome (ST) and single-nucleus RNA-seq to identify the spatial heterogeneity of IMF. ST revealed that genes involved in TGF-β signaling pathways were specifically highly enriched in IMF. In lean pigs, IMF autocrine produces more TGF-β2, while in obese pigs, IMF received more endothelial-derived TGF-β1. In vitro experiments have proven that porcine endothelial cells in a simulated high-fat environment released more TGF-β1 than TGF-β2. Moreover, under obesity mice, the addition of TGF-β after muscle injury abolished IMF production and slowed muscle repair, whereas TGF-β inhibition accelerated muscle repair. Our findings demonstrate that the TGF-β pathway specifically regulates these processes, suggesting it as a potential therapeutic target for managing muscle atrophy in obese patients and enhancing muscle repair while reducing IMF deposition.