Background: Cerebral ischemia-reperfusion injury (CIRI) is a key problem to be solved urgently in the treatment of ischemic stroke, which seriously affects the prognosis of patients. As a natural flavonoid, Vitexin has been confirmed to alleviate CIRI, but the molecular network mechanism of its action has not been systematically analyzed. Methods: The HMC3 cells were subjected to OGD/R treatment to establish a CIRI cell model. Effect of Vitexin on cell viability, apoptosis, and levels of inflammatory factors were determined by CCK-8, TUNEL, and ELISA methods. The intersection of Vitexin target genes obtained from the SwissTargetPrediction website and I/R-related genes predicted from the GeneCards website was displayed by Venn diagram, and the PPI network between them was constructed through the STRING website to screen key genes. Meanwhile, GO and KEGG analyses of these genes were performed on the DAVID 6.8 database. PPI network of Vitexin-key genes-signaling pathway was then constructed via STRING website. Molecular docking of Vitexin with key targets was achieved by AutuDock Vina software. Additionally, the expression of MAPK8 and MAPK signaling pathway-related proteins was monitored by Western blot. Besides, the in vivo effect of Vitexin on CIRI was evaluated via a MCAO model. Results: Vitexin could restore the cell viability of HMC3 cells impaired by OGD/R, and inhibited cell apoptosis and the levels of inflammatory factors. The Venn diagram identified 32 common genes betweenVitexin targets and I/R-related genes. Meanwhile, the top 5 genes (PIK3CA, MAPK8, ABL1, JAK2, and HDAC6) with the strongest interactions were screened via the PPI network. The GO and KEGG analyses of the 32 common genes revealed that they were mainly enriched in protein phosphorylation, ErbB signaling pathway, and MAPK signaling pathway. The Vitexin-Genes-Pathways PPI network indicated that the MAPK signaling pathway was sensitive to Vitexin. Molecular docking confirmed that vitexin had a close binding with five key genes. Moreover, Vitexin downregulated the expression of MAPK8 protein and curbed the MAPK signaling pathway. Vitexin could effectively alleviate CIRI, including cerebral infarction and inflammatory response in rats. Conclusion: Through the application of network pharmacology and molecular docking techniques, this study confirmed that Vitexin could strongly bind to the MAPK8 protein, thereby inhibiting its expression and blocking the MAPK signaling pathway to effectively alleviate CIRI.
Dian et al. (Thu,) studied this question.