Therapeutic effects of raw rhubarb on gastrointestinal complications in ischemic stroke: An integrated analysis of gut microbiota, metabolomics, and network pharmacology

Gastrointestinal complications (GITC) are a major cause of increased morbidity and mortality in stroke patients, significantly impairing recovery by triggering systemic inflammation and hindering brain healing. Raw Rhubarb (RR) is a commonly used traditional Chinese medicine with significant potential in treating GITC of ischemic stroke (IS). However, its therapeutic mechanisms remain largely unknown. This study aims to investigate the therapeutic effects and potential mechanisms of RR on GITC in IS through an integrated analysis of gut microbiota, metabolomics, and network pharmacology. Male Sprague-Dawley rats were subjected to middle cerebral artery occlusion/reperfusion (MCAO/R) to induce IS. The rats were divided into six groups: sham, model, nimodipine-treated, and three different dose groups for RR. Neuroprotective efficacy was assessed using infarct size measurement, neurological scoring, and histopathological analysis. Gut microbiota composition was analyzed through 16S rRNA gene sequencing, while metabolomic profiling of brain and colon tissues was performed using UPLC-Q-Orbitrap HRMS/MS. Multivariate statistical methods were employed to identify the key metabolites and pathways affected by RR treatment. Correlation analysis was conducted to establish links between gut microbiota alterations and differential metabolites. Additionally, network pharmacology, molecular docking analysis, and Western blot assays were utilized to explore the molecular mechanisms underlying RR's treatment of GITC in IS. RR showed significant neuroprotective effects, reducing infarct volume, improving neurological scores, and restoring intestinal function compared to the model group. In addition, gut microbiota analysis revealed that RR administration reversed gut microbiota dysbiosis in MCAO/R rats by increasing the abundance of Bifidobacterium and Lactobacillus while decreasing the abundance of Escherichia-Shigella . Metabolomics analysis indicated that RR reversed the metabolic disturbances in MCAO/R rats by modulating arachidonic acid (AA) metabolism. Correlation analysis showed that AA and its metabolites, such as PGE2, were closely associated with Bifidobacterium and Lactobacillus . Combining metabolomics, network pharmacology, and molecular docking analysis suggested that RR might regulate AA metabolism through the PI3K/mTOR signaling pathway to treat GITC in IS. Finally, Western blot validation confirmed that RR modulates the PI3K/mTOR signaling pathway. These findings indicate that RR holds significant promise as a therapeutic strategy for addressing GITC of IS. The protective effects mediated by RR are associated with the improvement of gut microbiota dysbiosis and metabolic disturbances.