Abstract Objective: Ischemic stroke is a leading cause of disability and mortality worldwide, and thrombosis plays a crucial role in its pathogenesis. Cooked Rhubarb (CR) has been traditionally used in stroke treatment, but its mechanisms of action remain unclear. This study aims to investigate the therapeutic effects of CR on IS from a thrombosis-related perspective, and using a rat model, we employ multiomics approaches to investigate its potential mechanisms. Materials and Methods: This study employed an integrative approach combining differential compound analysis, weighted gene co-expression network analysis (WGCNA), network pharmacology, metabolomics, gut microbiota analysis, and in vivo experiments using a middle cerebral artery occlusion/reperfusion (MCAO/R) model. Ultra-high-performance liquid chromatography coupled with quadrupole-Orbitrap high-resolution mass spectrometry identified key bioactive compounds in CR. Bioinformatics analysis, including WGCNA and network pharmacology, was conducted to predict CR-related targets and pathways in IS. The therapeutic efficacy of CR was evaluated through behavioral assessments, histopathological examination, coagulation function tests, and metabolomic profiling of brain and colon tissues. In addition, 16S rRNA sequencing was performed to explore CR’s influence on gut microbiota composition. Results: Wine cooking significantly increased the content of five anthraquinone compounds (emodin, aloe emodin, rhein, rhein methylester, and hydroxyl-emodin) and two others (p-coumaric-6-glucoside and ethyl gallate 4-glucuronide) in Rhubarb. WGCNA identified key stroke-associated modules, which were further analyzed through network pharmacology to predict that CR exerts its anti-IS effects primarily through the MAPK signaling pathway, sphingolipid signaling pathway, PI3K-Akt signaling pathway, and FoxO signaling pathway. In vivo experiments demonstrated that CR improved neurological function, reduced infarct volume, and alleviated histopathological damage in MCAO/R rats. Furthermore, CR significantly modulated coagulation function, reducing hypercoagulability and thrombosis-related biomarkers. Metabolomic analysis revealed that CR regulated sphingolipid and glycerophospholipid metabolism in brain tissue and arachidonic acid metabolism in colon tissue, suggesting a role in lipid homeostasis. Gut microbiota analysis indicated that CR intervention significantly enriched Akkermansia and Romboutsia while reducing Escherichia-Shigella abundance, highlighting its potential impact on the gut–brain axis. Integrative multiomics analysis revealed significant correlations between gut microbiota, lipid metabolism, and thrombus. Notably, Escherichia–Shigella abundance was closely correlated with lipid metabolism, particularly arachidonic acid, sphingolipid, and glycerophospholipid metabolism, suggesting a key role of the gut microbiota–metabolite axis in CR’s antithrombotic effects. Conclusions: This study systematically evaluated the pharmacological effects of CR in IS treatment, revealing that its protective effects are primarily mediated through the regulation of lipid metabolism, coagulation function, and gut microbiota composition. These findings expand the understanding of CR’s potential mechanisms in IS therapy and provide important theoretical support for future intervention strategies.
Cao et al. (Fri,) studied this question.