Stigmasterol suppresses macrophage-mediated inflammatory amplification in acute pancreatitis through PPAR-γ-dependent...

Glycine max (L.) Merr. (G. max; soybean), a widely cultivated medicinal and edible plant, has been traditionally used in Chinese medicine and dietary practices for maintaining digestive health. Stigmasterol (STG), a naturally occurring dietary phytosterol widely distributed in G. max and other edible and medicinal plants, is investigated here as a bioactive dietary constituent rather than as an isolate from a documented traditional remedy specifically used for acute pancreatitis (AP). STG has attracted increasing attention due to its anti-inflammatory and immunomodulatory activities; however, its potential role and underlying mechanisms in AP remain unclear.

This study aimed to investigate the protective effects of STG against AP and elucidate its role in regulating macrophage-mediated inflammatory amplification.

Publicly available transcriptomic, spatial transcriptomic, and single-cell RNA sequencing datasets were downloaded and bioinformatically analyzed to identify key inflammatory pathways and cellular interactions in AP. The protective effects of STG were evaluated in AP mice and compared with ulinastatin (UTI). Network pharmacology, molecular docking, molecular dynamics simulations, and cellular thermal shift assays (CETSA) were applied to identify STG targets. RAW264.7 macrophages and AR42J pancreatic acinar cells were used for in vitro validation, including peroxisome proliferator-activated receptor gamma (PPAR-γ) knockdown and co-culture experiments.

Multi-omics analyses revealed that Toll-like receptor 4 (TLR4)-mediated nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling is a central inflammatory pathway in AP and is closely associated with macrophage-related high mobility group box 1 (HMGB1) signaling. STG treatment alleviated pancreatic injury, inhibited NF-κB activation, and regulated macrophage polarization with comparable efficacy to UTI. Integrated pharmacological analyses and CETSA supported PPAR-γ as a key functional mediator and candidate molecular target of STG. STG regulated macrophage polarization and reduced extracellular HMGB1 release from activated macrophages in a PPAR-γ-dependent manner, accompanied by attenuation of macrophage-mediated TLR4/NF-κB-associated inflammatory responses in pancreatic acinar cells.

Although further validation in additional experimental models is warranted, STG ameliorates AP through PPAR-γ-dependent remodeling of macrophage immune responses, accompanied by reduced extracellular HMGB1 release and attenuation of TLR4/NF-κB-associated inflammatory signaling.