Abstract：Objective：To predict the therapeutic mechanism of Xuebijing Injection（XBJ）in acute kidney injury （AKI） based on network pharmacology and in vitro experiments. Methods： Active components of the five herbal medicines in XBJ were retrieved from literature and databases including the Integrative Pharmacology-based Network Computational Research Platform of Traditional Chinese Medicine（TCMIP）， Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform（TCMSP），and Traditional Chinese Medicine Immunology and Oncology Database（TCMIO）. Drug targets and disease-related targets were obtained from DrugBank， SwissTargetPrediction， Therapeutic Target Database，GeneCards，and OMIM. Venn diagrams were constructed to identify overlapping targets between XBJ and AKI. Gene Ontology（GO）functional annotation and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analyses were performed using the DAVID database and visualized. Protein-protein interaction（PPI） networks were constructed using Cytoscape 3.10.0 and the String database to screen core protein targets. A drug-component-target-pathway network was established to identify targets most relevant to disease pathways. Finally， in vitro experiments were conducted to investigate the effects of XBJ on the proliferation of human umbilical vein endothelial hybrid cells（EAhy926）and the expression of inflammatory factors. Results： A total of 114 active components，941 component targets，and 1 490 AKI-related targets were identified，with 252 overlapping drug-disease targets.Core therapeutic targets forAKIincluded serine/threonine kinase 1（AKT1），non-receptortyrosine kinase（SRC）， estrogen receptor 1（ESR1），mammalian target of rapamycin（mTOR），nuclear factor kappa B subunit 1（NF-κB1）， signal transducer and activator of transcription 3（STAT3）， B-cell lymphoma 2（BCL2）， and tumor necrosis factor （TNF）. GO enrichment analysis revealed that therapeutic targets were primarily involved in protein phosphorylation， negative regulation of apoptosis， epidermal growth factor receptor signaling pathway， and positive regulation of gene expression. KEGG pathway analysis highlighted key signaling pathways such as phosphatidylinositol 3-kinase-protein kinase B（PI3K-AKT）， Forkhead box O（FoxO）， TNF， and sphingolipid signaling pathway. In vitro experiments demonstrated that XBJ downregulated interleukin-6（IL-6）expression in lipopolysaccharide（LPS）-induced EAhy926 cells. Conclusion：XBJ may exert therapeutic effects on AKI through multiple active components acting on targets such as STAT3，NF-κB1，and mTOR，modulating inflammatory pathways including PI3K-AKT and TNF signaling pathway.