基于网络药理学的小檗碱治疗缺血性脑卒中作用机制探讨

目的 采用网络药理学方法探讨小檗碱治疗缺血性脑卒中的作用机制。方法 利用中药系统药理学数据库与分析平台（TCMSP）、BATMAN-TCM数据库及SwissTargetPrediction平台检索并预测小檗碱的靶点，通过DrugBank、OMIM、GeneCards、DisGeNET、TTD数据库获得缺血性脑卒中疾病的相关靶点，两者取交集即得小檗碱抗缺血性脑卒中作用靶点。采用Cytoscape 3.6.1软件构建作用靶点蛋白质-蛋白质相互作用（PPI）网络，筛选出关键靶点，并对其进行分子对接验证。采用DAVID数据库进行基因本体（GO）生物学过程和京都基因与基因组百科全书（KEGG）通路富集分析。细胞实验观察小檗碱对脑缺血再灌注损伤的作用及对关键靶基因细胞分裂周期蛋白42（CDC42）、肉瘤病毒蛋白（SRC）mRNA表达的影响。结果 共获得小檗碱抗缺血性脑卒中相关靶点44个，筛选出关键靶点6个。分子对接显示，关键靶点CDC42、SRC、丝裂原活化蛋白激酶14（MAPK14）、MAPK8及雌激素受体1（ESR1）与小檗碱亲和力较好。KEGG富集分析结果表明，小檗碱治疗缺血性脑卒中相关的信号通路有血管内皮生长因子（VEGF）信号通路、血小板活化、受体相关蛋白1（Rap1）信号通路、鞘脂信号通路、神经营养素信号通路等。细胞实验显示，与对照组相比，氧葡萄糖剥夺/再恢复模型组细胞活力明显下降（P<0.01），CDC42、SRC mRNA表达水平显著升高（P<0.01），而小檗碱中、高剂量能明显升高氧葡萄糖剥夺/再恢复细胞的活力（P<0.05，P<0.01），降低CDC42、SRC mRNA表达水平（P<0.05，P<0.01）。结论 小檗碱可能通过调节CDC42、ESR1、MAPK14、MAPK8和SRC等核心靶点，干预VEGF信号途径、血小板活化通路等，发挥治疗缺血性脑卒中的作用。

Potential Mechanism of Berberine in Treatment of Ischemic Stroke Based on Network Pharmacology

Objective To explore the mechanism of berberine in the treatment of ischemic stroke by network pharmacology.Methods The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), a Bioinformatics Analysis Tool for Molecular Mechanism of Traditional Chinese Medicine (BATMAN-TCM) database, and SwissTargetPrediction platform were used to retrieve and predict the targets of berberine. DrugBank, Online Mendelian Inheritance in Man (OMIM), GeneCards, DisGeNET, and Therapeutic Target Database (TTD) databases were used to obtain ischemic stroke related targets. The intersection is the targets of berberine against ischemic stroke. The target protein-protein interaction (PPI) network was constructed by Cytoscape 3.6.1 software. The key targets were screened out and were further verified by molecular docking. The gene ontology (GO) biological function annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were carried out by Database for Annotation, Visualization, and Integrated Discovery (DAVID). The effects of berberine on cerebral ischemia-reperfusion injury and the mRNA expression of key targets cell division cycle 42 (CDC42) and SRC were observed by cell experiment.Results We obtained 44 targets related to berberine against ischemic stroke, of which 6 key targets were screened. Molecular docking showed that the key targets CDC42, SRC, mitogen-activated protein kinase 14 (MAPK14), MAPK8, and estrogen receptor 1 (ESR1) had a good affinity to berberine. KEGG enrichment analysis demonstrated that the signaling pathways related to berberine against ischemic stroke included vascular endothelial growth factor (VEGF) signaling pathway, platelet activation, receptor-associated protein 1 (Rap1) signaling pathway, sphingolipid signaling pathway, and neurotrophin signaling pathway. In addition, the cell experiment exhibited that the cell viability of the oxygen-glucose deprivation/reoxygenation (OGD/R) model group was decreased (P<0.01) and the mRNA expressions of CDC42 and SRC were increased (P<0.01), as compared with the control group. Compared with the OGD/R model group, the medium- and high-dose berberine groups showed increased viability of OGD/R cells (P<0.05, P<0.01), and decreased mRNA expression of CDC42 and SRC (P<0.05, P<0.01).Conclusion Berberine may play a role in the treatment of ischemic stroke by regulating core targets such as CDC42, ESR1, MAPK14, MAPK8, and SRC and intervening in VEGF signaling pathway and platelet activation pathway.