基于网络药理学及分子对接发掘当归-川芎药对治疗肺栓塞的作用机制

Research of the pharmacological mechanism of drup pair Angelicae Sinensis Radix-Chuanxiong Rhizoma against pulmonary embolism based on network pharmacology and molecular docking

目的:通过网络药理学及分子对接技术发掘当归-川芎药对治疗肺栓塞的作用机制。方法:TCMSP查询当归-川芎药对的药物成分,使用Swiss Target Prediction预测药物成分的作用靶点;GeneCards网站在线获取肺栓塞的疾病靶点,Venny 2.1在线获取药物和疾病交集靶点。使用STRING进行蛋白-蛋白相互作用(PPI)分析并使用Cytoscape构建网络图,利用Metascape进行GO和KEGG分析。使用Cytoscape软件构建“药物-靶点-通路”网络图。PubChem、PDB及PyMoL、AutoDock软件进行分子对接。建立肺栓塞大鼠动物模型,治疗组予以中药当归-川芎药对灌胃治疗2周,对照组予以等量水灌胃,酶联免疫法检测两组大鼠血清相应靶点蛋白表达。结果:当归-川芎药对中的8个有效成分通过多条通路直接作用于25个疾病靶点治疗肺栓塞,其中β-谷甾醇、豆甾烯醇、杨梅酮、芎萘呋内酯等是核心成分,半胱氨酸蛋白酶3(caspase 3,CASP3)、雌激素受体1(estrogen receptor alpha,ESR1)、肉瘤病毒17癌基因(JUN)、前列腺素内过氧化物合酶2(prostaglandin-endoperoxide synthase 2,PTGS2)是核心靶点。GO富集分析结果显示,交集基因最可能相关的生物过程主要涉及细胞对形态发生、激素反应、有机环化合物的反应等,细胞组分主要涉及膜筏、膜微区、线粒体外膜等,分子功能主要涉及半胱氨酸内肽酶活性、泛素蛋白连接酶、蛋白二聚酶活性等。KEGG通路富集分析结果提示当归-川芎药对主要参与低氧诱导因子-1(HIF-1)、脂代谢、HCI、神经变性等信号通路。分子对接结果提示核心成分与重要靶点间的结合性较好。酶联免疫检测显示治疗组血清ESR1、JUN、PTGS2蛋白表达均高于对照组(t=-8.018、-10.370、-6.545,均P<0.01)。结论:当归-川芎主要通过调节HIF-1、脂代谢、HCI、神经变性等信号通路的ESR1、JUN、PTGS2等疾病靶点,干预酶的活性、脂代谢、神经变性等生物学过程,进而治疗肺栓塞。

Objective:To explore the mechanism of drup pair Angelicae Sinensis Radix-Chuanxiong Rhizoma in the treatment of pulmonary embolism through network pharmacology and molecular docking technology,and verify by animal experiments.Methods:TCMSP was used to screen the drug components of drup pair Angelicae Sinensis Radix-Chuanxiong Rhizoma,and the targets of drug components were predicted by the Swiss Target Prediction.Disease targets of pulmonary embolism were acquired online by Gene Cards,and intersection targets of drugs and diseases are acquired online by Venny 2.1.A protein-protein interaction(PPI)network was constructed by STRING and Cytoscape software,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were carried out by Metascape.The"drug-target-pathway"network was established by Cytoscape software.Molecular docking was carried out with PubChem,PDB,PyMoL and AutoDock software.The rat model of pulmonary embolism was established.The treatment group was given Angelica-Szechuan Lovage Rhizome pair by intragastric administration for 2 weeks,and the control group was given the same amount of water by intragastric administration for 2 weeks.The expression of corresponding target proteins in serum of the two groups were detected by the Enzyme linked immunosorbent assay.Results:It was found that 8 active components of drup pair Angelicae Sinensis Radix-Chuanxiong Rhizoma directly acted on 25 disease targets through multiple pathways in the treatment of pulmonary embolism,and the core components wereβ-sitosterol,stigmatenol,myricetin and ligustrafurlactone,and the most important targets were caspase 3(CASP3),estrogen receptor alpha(ESR1),JUN,prostaglandin-endoperoxide synthase 2(PTGS2).GO enrichment analyses found which biological processes mainly including cell morphogenesis,hormone reaction,organic compounds.Cellular component mainly included the membrane rafts,membrane micro zone,mitochondrial outer membrane,and molecular function mainly included cysteine in peptide protein enzymes,protein ubiquitin ligase,dimer enzyme activity.KEGG enrichment analyses found signaling pathways mainly including HIF-1,lipid metabolism,HCI,neurodegeneration,et al.The molecular docking results indicated that the core components and important targets had good binding properties.It was showed that the expression of ESR1,JUN and PTGS2 proteins in the treatment group were higher than those in the control group by the Enzyme linked immunosorbent assay(t=-8.018,-10.370,-6.545 respectively,all P<0.01).Conclusion:Drup pair Angelicae Sinensis Radix-Chuanxiong Rhizoma mainly regulates the targets of ESR1,JUN,PTGS2 and the signaling pathways of HIF-1,lipid metabolism,HCI,and neurodegeneration,and interferes with the biological processes of enzyme activity,lipid metabolism,and neurodegeneration to treat pulmonary embolism.