[Network pharmacology and experimental validation reveal therapeutic mechanism of Yuyang Jianpi Pills for gastric ulcer with spleen-stomach deficiency-cold syndrome by inhibiting inflammation and apoptosis via IL-6/JAK2/STAT3 pathway].

This study integrated network pharmacology, molecular docking, and animal experiments to investigate the therapeutic effects and mechanisms of Yuyang Jianpi Pills for gastric ulcer(GU) in rats modeled with spleen-stomach deficiency-cold syndrome. Active ingredients and targets of Yuyang Jianpi Pills were searched against TCMSP, BATMAN-TCM, SwissTargetPrediction, and relevant literature. The targets associated with GU and spleen-stomach deficiency-cold syndrome were screened from the GeneCards database and the TCMIP platform. The common targets shared by Yuyang Jianpi Pills and GU were identified as core therapeutic targets. Subsequent analyses included protein-protein interaction(PPI) network construction, Gene Ontology(GO) functional annotation, and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment. Molecular docking was performed to validate the binding interactions between key active ingredients and core targets. Male SPF-grade SD rats were randomized into a sham-operated group(8 rats) and a model group(52 rats). The GU model with spleen-stomach deficiency-cold syndrome in rats was established by a combination of bitter-cold purgation, irregular feeding, and direct acetic acid cauterization. After the model was successfully established, the rats were randomly allocated into model, omeprazole(3.6 mg·kg~(-1)·d~(-1)), and high-, medium-, low-dose(2.2, 1.1, 0.55 g·kg~(-1)·d~(-1), respectively) Yuyang Jianpi Pills groups(n=8). The drugs were administered continuously for 2 weeks. Food intake, body weight, rectal temperature, and symptoms of spleen-stomach deficiency-cold syndrome were monitored to evaluate modeling success and treatment efficacy. After gastric tissue collection, the pathological changes in the tissue were observed via hematoxylin-eosin(HE) staining. The enzyme-linked immunosorbent assay(ELISA) was used to quantify the levels of interleukin(IL)-17, prostaglandin E2(PGE2), and pepsin content. Western blot was employed to determine the expression levels of factors in the IL-6/janus kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3) signaling pathway, as well as B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), and cleaved cysteinyl aspartate-specific proteinase 3(cleaved caspase-3). immunohistochemical assay was adopted to detect the expression levels of IL-6, phosphorylated(p)-JAK2, and p-STAT3 in gastric mucosa. Gastric mucosal cell apoptosis was detected by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling(TUNEL) assay. A total of 1 094 drug targets and 1 104 disease targets were screened out, with an intersection of 205 core targets. The PPI, GO and KEGG enrichment analyses, and molecular docking indicated that the therapeutic mechanism of Yuyang Jianpi Pills for GU with spleen-stomach deficiency-cold syndrome was closely related to the IL-6/JAK2/STAT3 signaling pathway, inflammation, and apoptosis. The results of animal experiments confirmed that during modeling, compared with the sham group, the model group exhibited significantly decreased food intake, and rectal temperature, along with slower body mass growth and pronounced symptoms of spleen-stomach deficiency-cold syndrome. After administration, compared with the model group, the high-and medium-dose Yuyang Jianpi Pills groups showed significant increases in food intake, body mass, and rectal temperature, along with significant alleviation of spleen-stomach deficiency-cold syndrome symptoms. Each treatment group showed alleviated pathological damage of the gastric mucosa to varying degrees, declined levels of IL-17 and pepsin in the gastric tissue, elevated levels of PGE2, and down-regulated protein levels of IL-6, p-JAK2, and p-STAT3 in the gastric mucosa. TUNEL staining showed that the treatments reduced gastric mucosal cell apoptosis, down-regulated the protein levels of Bax and cleaved caspase-3 in the gastric tissue, while up-regulating the expression of Bcl-2. These findings demonstrate that Yuyang Jianpi Pills effectively treats GU in rats with spleen-stomach deficiency-cold syndrome, potentially through regulating the IL-6/JAK2/STAT3 signaling pathway, thereby suppressing gastric mucosal inflammation and inhibiting cell apoptosis.