| Sumario: | Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by symptoms such as persistent diarrhea. Shenlingbaizhu Formula (SLBZ), a classical Chinese herbal formula with centuries of clinical application, has long been used to treat such gastrointestinal disorders. This historical efficacy provides a strong ethnopharmacological rationale for investigating its therapeutic potential and underlying mechanisms in UC. To elucidate the molecular mechanism by which SLBZ ameliorates UC by restoring the intestinal barrier via the sGC-cGMP-PKG signaling pathway. To systematically investigate the therapeutic effects of SLBZ on UC, we established a dextran sulfate sodium (DSS)-induced colitis model in 57BL/6 mice. Comprehensive evaluations were conducted to assess intestinal barrier function and elucidate underlying mechanisms, including histological analysis, immunofluorescence staining of tight junction proteins, 16S rDNA sequencing for gut microbiota profiling, transcriptomic analysis for gene expression patterns, and Western blot validation of key proteins. Furthermore, a multi-tiered experimental strategy was employed, integrating molecular modeling, machine learning algorithms, graph neural network (GNN) approaches and in vitro experiments using Caco-2 cell monolayers, to systematically validate the sGC-cGMP-PKG signaling pathway and identify bioactive compounds responsible for SLBZ's therapeutic effects. SLBZ treatment markedly alleviated UC symptoms in mice and restored intestinal barrier function through dual mechanisms: upregulation of tight junction proteins (ZO-1/Occludin) and modulation of gut microbiota composition. Transcriptomic analysis pinpointed the cGMP-PKG signaling pathway as a central therapeutic target. Through an integrated approach combining molecular docking, machine learning (random forest algorithm), and deep neural network analysis, we identified soluble guanylate cyclase (sGC) as the primary molecular target, with Choerospondin emerging as a key bioactive component. Mechanistic investigations demonstrated that SLBZ robustly activated the sGC-cGMP-PKG axis both in vivo (colon tissues) and in vitro (Caco-2 cells), culminating in significant suppression of intestinal epithelial apoptosis. SLBZ alleviates UC by activating the sGC-cGMP-PKG pathway to restore intestinal barrier function through tight junction upregulation and microbiota modulation. Our study both validates SLBZ's traditional use and identifies this pathway as a novel therapeutic target for UC. [Display omitted] • SLBZ restores mechanical, chemical & immune barriers in UC mice. • 16S rDNA shows SLBZ modulates gut microbiota & biological barrier. • Transcriptomics, ML and GNN reveal cGMP-PKG pathway as a core mechanism. • Choerospondin is identified as a key active compound activating sGC. • In vivo & in vitro tests confirm SLBZ activates the sGC-cGMP-PKG pathway.
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