Astragaloside IV suppresses neuroinflammation via PI3K/Akt/NF-κB to ameliorate cerebral ischemia-reperfusion injury based on network pharmacology analysis and experimental validation

Aim This study aimed to systematically investigate the therapeutic effects and underlying mechanisms of ASIV against cerebral ischemia-reperfusion injury (CIRI) using an integrated approach combining network pharmacology and experimental validation. Methods Potential targets of Astragaloside IV (ASIV) were predicted using SwissTargetPrediction, PharmMapper and Comparative Toxicogenomics Database (CTD). Ischemic stroke-related targets were collected from GeneCards, DisGeNET and DrugBank. Overlapping targets were used to construct a protein-protein interaction (PPI) network via STRING and visualized in Cytoscape. Functional enrichment analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways was performed using DAVID. The neuroprotective effects of ASIV were evaluated in a mouse model of middle cerebral artery occlusion/reperfusion (MCAO/R), assessing neurological function, infarct volume, blood brain barrier (BBB) integrity, inflammatory markers, and PI3K/Akt/NF-κB pathway activity. In vitro , LPS-stimulated BV-2 microglia were used to examine the effects of ASIV on cell viability, inflammatory cytokine expression, and PI3K/Akt/NF-κB signaling. Results Network pharmacology analysis identified 166 overlapping targets, and enrichment analysis emphasized the PI3K/Akt pathway as a key mechanism. In MCAO/R mice, ASIV significantly improved neurological function, reduced infarct volume and neuronal apoptosis, decreased Evans blue leakage, attenuated MMP-9 expression, and restored ZO-1 and Occludin levels. ASIV also suppressed the mRNA levels of IL-6, IL-1β, and TNF-α both in vivo and in LPS-stimulated BV-2 cells. Additionally, ASIV effectively upregulated the expression of Anti-inflammatory cytokines (IL-10 and Arg-1) in LPS-induced BV2 cells. Mechanistically, ASIV suppresses NF-κB activation through the stimulation of the PI3K/Akt signaling pathway. Conclusion ASIV exerts neuroprotective effects against CIRI by inhibiting neuroinflammation and preserving BBB integrity likely through modulation of the PI3K/Akt/NF-κB signaling pathway.