Exploring the molecular mechanism of Shenling baizhu formula in the treatment of ulcerative colitis via the cGMP-PKG signaling pathway.

SLBZ treatment markedly alleviated UC symptoms in DSS-induced colitis mice.

• A dextran sulfate sodium (DSS)-induced colitis model was established in C57BL/6 mice.
• Comprehensive evaluations included histological analysis, immunofluorescence staining, 16S rDNA sequencing, transcriptomic analysis, and Western blot validation.
• SLBZ treatment reduced clinical symptoms including persistent diarrhea characteristic of UC.

SLBZ restored intestinal barrier function through upregulation of tight junction proteins ZO-1 and Occludin.

• Tight junction protein expression was assessed via immunofluorescence staining.
• Western blot validation confirmed upregulation of ZO-1 and Occludin proteins.
• The restoration of tight junction proteins was identified as one of two dual mechanisms underlying SLBZ's barrier-restoring effects.

SLBZ modulated gut microbiota composition in DSS-induced colitis mice.

• Gut microbiota profiling was performed using 16S rDNA sequencing.
• Modulation of gut microbiota composition was identified as the second of two dual mechanisms by which SLBZ restores intestinal barrier function.
• Gut microbiota modulation was observed in colon tissues of treated mice.

Transcriptomic analysis identified the cGMP-PKG signaling pathway as a central therapeutic target of SLBZ.

• Transcriptomic analysis was conducted to assess gene expression patterns in treated vs. untreated colitis mice.
• The cGMP-PKG signaling pathway was pinpointed as a central therapeutic target among differentially expressed genes.
• This finding directed subsequent mechanistic investigations into the sGC-cGMP-PKG axis.

Soluble guanylate cyclase (sGC) was identified as the primary molecular target of SLBZ using an integrated computational approach.

• A multi-tiered strategy integrating molecular docking, machine learning (random forest algorithm), and deep neural network (graph neural network, GNN) analysis was employed.
• sGC was identified as the primary molecular target among candidates evaluated.
• Choerospondin was identified as a key bioactive component of SLBZ responsible for its therapeutic effects.
• In vitro experiments using Caco-2 cell monolayers were used to validate computational findings.

SLBZ robustly activated the sGC-cGMP-PKG signaling axis both in vivo and in vitro.

• Activation of the sGC-cGMP-PKG pathway was demonstrated in colon tissues (in vivo) and Caco-2 cells (in vitro).
• Activation of this pathway resulted in significant suppression of intestinal epithelial apoptosis.
• Western blot was used among the methods to validate key proteins in this signaling pathway.

An integrated molecular modeling and machine learning framework was used to systematically validate SLBZ's mechanism and identify bioactive compounds.

• The framework combined molecular modeling, machine learning algorithms, and graph neural network (GNN) approaches.
• In vitro experiments using Caco-2 cell monolayers complemented the computational strategy.
• This multi-tiered approach enabled both pathway validation and identification of key bioactive components such as Choerospondin.