摘要
通过网络药理学与分子对接技术探讨中风醒脑液干预脑出血微循环障碍的作用机制。中风醒脑液组分化学成分信息通过中药系统药理数据库和分析平台(Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform,TCMSP),其化学成分预测的靶点分别通过PubChem和SwissTargetPrediction获得,脑出血和微循环障碍的相关靶点收集源于GeneCards数据库,成分与疾病共同靶点经过DAVID数据库进行基因本体论(Gene Ontology,GO)和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集分析,使用Cytoscape软件将相关网络关系可视化,进一步筛选重要化学成分与疾病靶点进行分子对接预测。动物实验验证采用改良神经功能评分(modified neurological severity score,mNSS)、酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)、实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction,qRT-PCR)、免疫荧光和蛋白质印迹法(Western blot)检测中风醒脑液干预脑出血模型小鼠的作用情况。结果显示,中风醒脑液4个组分共有31个化学成分和856个靶点,脑出血微循环障碍靶点共有173个,疾病与成分共同靶点57个;富集分析显示共同靶点主要参与细胞增殖、凋亡等生物过程,肿瘤通路、病毒感染、磷脂酰肌醇-3-激酶/蛋白激酶B(phosphoinositide-3-kinase/protein kinase B,PI3K/AKT)信号通路、丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路等信号通路。分子对接结果发现大黄、三七、红参的共同成分β-谷甾醇与原癌基因酪氨酸蛋白激酶(proto-oncogene tyrosine-protein kinase,SRC)、信号转导和转录激活因子3(signal transducer and activator of transcription 3,STAT3)、磷脂酰肌醇-3-激酶催化亚基α基因(phosphoinositide-3-kinase catalytic alpha polypeptide gene,PIK3CA)、非受体型蛋白酪氨酸磷酸酶11(recombinant protein tyrosine phosphatase non receptor type 11,PTPN11)、AKT1、表皮生长因子受体(epidermal growth factor receptor,EGFR)、钙黏连相关蛋白β1(calcium adhesion-associated protein beta 1,CTNNB1)、血管内皮生长因子A(vascular endothelial growth factor A,VEGFA)和肿瘤蛋白p53(tumor protein p53,TP53),大黄的成分番泻苷E与MAPK1的靶点对接情况良好。实验结果发现中风醒脑液可减缓脑出血模型小鼠的神经损伤,降低S100钙结合蛋白B(S100 calcium-binding protein B,S100β)、神经元特异性烯醇化酶(neuron specific enolase,NSE)、基质金属蛋白酶9(matrix metalloproteinase 9,MMP9)、肿瘤坏死因子α(tumor necrsis factor α,TNF-α)、白细胞介素1β(interlenkin 1β,IL-1β)、SRC、EGFR、CTNNB1、VEGFA、TP53、胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)和白细胞分化抗原86(leukocyte differentiation antigen 86,CD86)的表达,增加p-PI3K、p-AKT和闭锁小带蛋白1(zona occludens 1,ZO-1)的表达。结果表明,中风醒脑液可能是通过PI3K/AKT/p53通路抑制神经元凋亡和炎症反应,保护血脑屏障从而减缓脑出血微循环障碍。
This study aimed to explore the mechanism of Zhongfeng Xingnao Decoction(ZFXN) in intervening microcirculatory di-sorders in cerebral hemorrhage by network pharmacology and molecular docking techniques.The information on the components of ZFXN was obtained through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP) database,and the predicted targets of chemical components were obtained from PubChem and SwissTargetPrediction.The relevant targets of cerebral hemorrhage and microcirculatory disorders were collected from the GeneCards database,and the common targets of the components and diseases were analyzed by the Database for Annotation,Visualization,and Integrated Discovery(DAVID) for Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analyses.Visualization of the correlation network was carried out using Cytoscape software to further screen important chemical components for molecular docking prediction with disease targets.The animal experiment validation was performed using modified neurological severity score(mNSS),enzyme-linked immunosorbent assay(ELISA),quantitative real-time polymerase chain reaction(qRT-PCR),immunofluorescence,and Western blot to detect the effects of ZFXN intervention in mice with cerebral hemorrhage.The results showed that there were 31 chemical components and 856 targets in the four drugs contained in ZFXN,173 targets for microcirculatory disorders in cerebral hemorrhage,and 57 common targets for diseases and components.The enrichment analysis showed that common targets were mainly involved in biological processes,such as cell proliferation and apoptosis,and signaling pathways,such as tumor pathway,viral infection,phosphoinositide-3-kinase/protein kinase B(PI3K/AKT) signaling pathway,and mitogen-activated protein kinase(MAPK) signaling pathway.Molecular docking results revealed that the common components β-sitosterol of Rhei Radix et Rhizoma,Notoginseng Radix et Rhizoma,and Ginseng Radix et Rhizoma Rubra showed good docking with proto-oncogene tyrosine-protein kinase(SRC),signal transducer and activator of transcription 3(STAT3),phosphoinositide-3-kinase catalytic alpha polypeptide gene(PIK3CA),recombinant protein tyrosine phosphatase non receptor type 11(PTPN11),AKT1,epidermal growth factor receptor(EGFR),calcium adhesion-associated protein beta 1(CTNNB1),vascular endothelial growth factor A(VEGFA),and tumor protein p53(TP53).Moreover,sennoside E of Rhei Radix et Rhizoma showed good docking with MAPK1.The results revealed that the ZFXN relieved the neural injury in mice with cerebral hemorrhage,decreased the expression of S100 calcium-binding protein B(S100β),neuron specific enolase(NSE),matrix metalloproteinase 9(MMP9),tumor necrosis factor α(TNF-α),interleukin 1β(IL-1β),SRC,EGFR,CTNNB1,VEGFA,TP53,glial fibrillary acidic protein(GFAP),and leukocyte differentiation antigen 86(CD86),and increased the expression of p-PI3K,p-AKT,and zona occludens 1(ZO-1).The results indicate that ZFXN may inhibit neuronal apoptosis and inflammatory response through PI3K/AKT/p53 pathway to protect the blood-brain barrier,thereby slowing down microcirculatory impairment in cerebral hemorrhage.
作者
钟晓琴
胡达峰
王宇
宁振求
邓敏贞
ZHONG Xiao-qin;HU Da-feng;WANG Yu;NING Zhen-qiu;DENG Min-zhen(Guangzhou University of Chinese Medicine,Guangzhou 510006,China;the Second Affiliated Hospital of Guangzhou University of Chinese Medicine/Guangdong Provincial Hospital of Chinese Medicine/Guangdong Provincial Academy of Chinese Medical Sciences,Guangzhou 510120,China)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2023年第22期6115-6127,共13页
China Journal of Chinese Materia Medica
基金
广州市科学技术局项目(2023A03J0744)
广东省中医院朝阳人才科研专项(ZY2022KY06)
国家自然科学基金项目(81904104)。
关键词
中风醒脑液
网络药理学
脑出血
微循环障碍
Z Zhongfeng Xingnao Decoction
network pharmacology
cerebral hemorrhage
microcirculatory disorders
