Mechanism of the Active Ingredients of Ephedra sinica Stapf Dried Grass Stems in the Treatment of Vasospasm Based on Network Pharmacology
[Objective]The experiment was to explore the potential mechanism of the active ingredients of Ephedra sinica Stapf dried grass stems in the treatment of vasospasm by integrating network pharmacology and in vitro cell experiments.[Method]The main active ingredients and targets of Ephedra sinica Stapf dried grass stems were obtained from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),SwissTargetPrediction and PharmMapper databases.The disease targets were obtained from DrugBank,GeneCards and OMIM databases.Then the ingredient-disease intersection targets were obtained.STRING database was used for protein-protein interaction(PPI)analysis,and Cytoscape 3.9.1 software was used to screen the core targets and core ingredients.Metascape platform was used for GO function and KEGG pathway enrichment analysis,and AutoDock Vina platform was used for molecular docking.Subsequently,the effect of active ingredients of Ephedra sinica Stapf dried grass stems on the proliferation of vascular smooth muscle cells(vSMCs)induced by angiotensin Ⅱ was detected by MTT method.[Result]The active ingredients of Ephedra sinica Stapf dried grass stems compound mainly included luteolin,quercetin,beta-quercetin,eriodictyol,naringenin,stigmasterol,kaempferol,etc.The main targets of these drug ingredients were 488,and 394 were related with vasospasm.Ephedra sinica Stapf dried grass stems had 67 prediction targets for vasospasm,mainly including matrix metallopeptidase Ⅸ(MMP9),MMP2,angiotensin Ⅱ receptor type 1(AGTR1),mitogen-activated protein kinase 1(MAPK1),vascular endothelial growth factor A(VEGFA),epidermal growth factor receptor(EGFR)and nitric oxide synthase 2(NOS2).Enrichment analysis of the above 67 targets found that they were related to biological processes such as positive regulation of cell migration,response to xenobiotic stimulus,response to hypoxia and positive regulation of angiogenesis,cell components such as nucleus,cytoplasm,extracellular space,plasma membrane,postsynaptic membrane,presynaptic membrane,dendrite,membrane raft and axon terminus,molecular functions such as RNA polymerase Ⅱ transcription factor activity,ligand-activated sequence-specific DNA binding,serine-type endopeptidase activity,endopeptidase activity,enzyme binding and zinc ion binding,and calcium signaling pathway,vascular smooth muscle contraction,relaxin signaling pathway,phosphatidylinositol 3-kinase/protein kinase B pathway(PI3K-Akt),sphingolipid signaling pathway and other pathways.The results of molecular docking showed that luteolin and quercetin,the main active ingredients of the Ephedra sinica Stapf dried grass stems compound,had excellent binding activity with the key targets MMP2 and MMP9.The in vitro test results showed that luteolin might inhibit angiotensin Ⅱ-induced proliferation and phenotype transformation of vSMCs through inactivating MAPK1 signaling pathways.[Conclusion]Various active ingredients of Ephedra sinica Stapf dried grass stems might play a role in the treatment of vasospasm by acting on targets,such as MMP2,MMP9,angiotensin Ⅰ converting enzyme(ACE),NOS2,etc.The study provided a scientific basis for revealing the mechanism of active ingredients of Ephedra sinica Stapf dried grass stems in the treatment of vasospasm.
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