益气凉血生肌方抗心肌缺血再灌注损伤作用机制的网络药理学分析及实验验证OA

Objective To predict the mechanism of Yiqi Liangxue Shengji Decoction against myocardial ischemia reperfusion injury(MIRI)based on network pharmacology and to validate it by cell experiments.Methods The main active components and potential targets of the Decoction were screened on the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP).The target names were standardized via the UniProt database.The Online Mendelian Inheritance in Man(OMIM)and GeneCards databases were combined to obtain targets related to the Decoction against MIRI.The protein-protein interaction(PPI)network model was constructed using the Search Tool for the Retrieval of Interacting Genes/Proteins(STRING)database.Gene ontology(GO)functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis were performed by the Metascape database to identify relevant pathways and targets.For in vitro experiments,AC16 myocardial cells were exposed to hypoxia/reoxygenation(H/R)to mimic MIRI and pretreated with different concentrations of Decoction-containing intestinal absorption solution.Cell viability was measured by the cell counting kit-8(CCK-8)assay.Inflammatory factor[interleukin-6(IL-6),interleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α)]levels in cells were measured by enzyme-linked immunosorbent assay(ELISA).mRNA expression of these inflammatory factors was examined by reverse transcription-polymerase chain reaction(RT-PCR).Results According to network pharmacology results,113 active components and 195 potential targets were identified from the Decoction,with 2480 MIRI-related gene targets.After intersection,120 targets were obtained against MIRI.GO functional enrichment analysis yielded 1966 GO entries,including 1707 biological processes,93 cellular components,and 166 molecular functions.KEGG pathway enrichment analysis identified 196 signaling pathways,primarily involving key target genes such as IL-6,IL-1β,and TNF,alongside key inflammatory pathways such as the TNF signaling pathway,the lipid and atherosclerosis pathway,and the nuclear factor-κB pathway.In cell experiments,compared to the control group,the model group exhibited significantly reduced cell viability,increased IL-6,IL-1β,and TNF-α expression levels,along with increased IL-6,IL-1β,and TNF-α mRNA expression levels(P<0.001).Compared to the model group,0.1%,0.25%,and 0.5%Decoction-containing intestinal absorption solution showed significantly elevated cell viability(P<0.05,P<0.01).The TCM groups demonstrated decreased IL-6,IL-1β,and TNF-α expression levels(P<0.001),along with decreased IL-6,IL-1β,and TNF-α mRNA expression levels(P<0.05,P<0.001).Conclusion The Decoction exhibits a multi-target,multi-pathway synergistic effect against MIRI,with underlying mechanisms related to inflammatory signaling pathways associated with TNF-α,IL-6,IL-1β,and others.