世界中医药

作者	单位
李鑫1，于丽红2，王兴臣2，胡晓洁2，郑浩1	1 山东中医药大学，济南，250013； 2 山东中医药大学第二附属医院，济南，250001

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中文摘要:

目的：基于网络药理学及动物实验分析化浊行血汤治疗脑缺血耐受（CIT）的机制。方法：通过中药系统药理学数据库与分析平台（TCMSP）、中医药百科全书数据库（ETCM）、中药分子机制的生物信息学分析工具（BATMAN-TCM）数据库及文献筛选化浊行血汤的有效成分及其靶点；通过基因数据库(GeneCards)、在线人类孟德尔遗传病数据库(OMIM)获取CIT靶点，绘制韦恩图获得化浊行血汤调治CIT的作用靶点；利用STRING平台构建靶点相互作用网络。利用Metascape对关键靶点进行基因本体（GO）通路富集分析与京都基因与基因组百科全书（KEGG）通路富集分析。通过大鼠实验进行验证，将SD大鼠用线栓法行脑缺血造模，灌服中药化浊行血汤及缺血预适应干预，横木行走试验（BWT）评价运动功能恢复情况、TTC染色法计算脑梗死体积、酶联免疫吸附试验法检测一氧化氮合酶不同亚型（内皮型一氧化氮合酶、神经元型一氧化氮合酶、诱生型一氧化氮合酶）的浓度观测缺血耐受程度。结果：网络药理学发现化浊行血汤中槲皮素、山柰酚、植物甾醇、木犀草素等化合物协同发挥产生缺血耐受的作用，涉及磷脂酰肌醇-3-激酶-蛋白激酶B（PI3K-AKT）、缺氧诱导因子1（HIF-1）、环腺苷酸（cAMP）、核因子κB（NF-κB）等信号通路及蛋白激酶B（AKT1）、肿瘤坏死因子（TNF）、白细胞介素-6（IL-6）、抑癌基因53（TP53）等靶点蛋白。动物实验预缺血+缺血组与中药+缺血组内皮型一氧化氮合酶、神经元型一氧化氮合酶、诱生型一氧化氮合酶的浓度以及脑梗死体积差异无统计学意义（P>0.05），预缺血+缺血组、中药+缺血组脑梗死体积小于缺血组（P<0.05），内皮型一氧化氮合酶的浓度明显高于缺血组（P<0.05）。结论：研究阐述了化浊行血汤多靶点、多通路产生缺血耐受的机制，证实了化浊行血汤能够影响内皮型一氧化氮合酶的表达，验证了网络药理学结果，为深入研究化浊行血汤产生缺血耐受提供了依据。

English Summary:

To explore the mechanism of Huazhuo Xingxue Decoction in inducing cerebral ischemic tolerance(CIT) based on network pharmacology and animal experiment.Methods:The effective components and targets of Huazhuo Xingxue Decoction were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP)，Encyclopedia of Traditional Chinese Medicine(ETCM)，Bioinformatics Analysis Tool for Molecular mechANism of Traditional Chinese Medicine(BATMAN-TCM)，and literature.The targets of CIT were obtained from GeneCards and Online Mendelian Inheritance in Man(OMIM).Venn diagram was established to identify the targets of Huazhuo Xingxue Decoction in regulating CIT and STRING was employed to build the protein-protein interaction network.Metascape was used for gene ontology(GO) annotation and Encyclopedia of Genes and Genomes(KEGG) for the key targets.The model of cerebral ischemia was established by the Longa thread bolt method in SD rats，which were then intervened with Huazhuo Xingxue Decoction and ischemic preconditioning.Beam walking test(BWT) was employed to evaluate the recovery of motor function，2,3,5-triphenyltetrazolium chloride(TTC) staining to measure the volume of cerebral infarction，and enzyme-linked immunosorbent assay to determine the concentrations of endothelial nitric oxide synthase(eNOS)，neuronal nitric oxide synthase(nNOS)，and inducible nitric oxide synthase(iNOS).Results:Quercetin，kaempferol，phytosterol，luteolin and other compounds in Huazhuo Xingxue Decoction play a synergistic role in inducing ischemic tolerance，involving phosphatidylinositol 3-kinase/protein kinase B(PI3K-AKT)，hypoxia-inducible factor-1(HIF-1)，cyclic adenosine monophosphate(cAMP)，and nuclear factor-κB(NF-κB) signaling pathways and the targets of AKT1，tumor necrosis factor(TNF)，interleukin-6(IL-6)，and tumor protein p53(TP53).Animal experiments showed that the ischemic preconditioning group and Huazhuo Xingxue Decoction group had no differences in the concentration of eNOS，nNOS，and iNOS and the volume of cerebral infarction(P>0.05).The two intervention groups had smaller volume of cerebral infarction and higher concentration of eNOS than the model group(P<0.05).Conclusion:Huazhuo Xingxue Decoction induces CIT in a multi-target and multi-pathway manner and regulates the expression of eNOS.The results of animal experiments verify the results obtained based on network pharmacology，which provides a basis for in-depth study of the CIT induced by Huazhuo Xingxue Decoction.

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