基于网络药理学、分子对接、代谢组学及实验验证分析葛根汤改善大脑缺氧减轻癫痫发作的机制  

Analysis of the Mechanism of Gegen Decoction in Improving Cerebral Hypoxia and Alleviating Epileptic Seizure Based on Network Pharmacology,Molecular Docking,Metabolomics and Experimental Verification

作　　者：王子格 任润雪 范文涛[1] WANG Zige;REN Runxue;FAN Wentao(Shaanxi University of Chinese Medicine,Xianyang 712056 Shaanxi,China)

机构地区：[1]陕西中医药大学,陕西咸阳712056

出　　处：《中药新药与临床药理》2024年第9期1389-1400,共12页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金项目(82274332);陕西省科技厅项目(2023-JC-YB-761)。

摘　　要：目的采用网络药理学、分子对接、代谢组学及实验验证探讨葛根汤对缺氧状态下癫痫发作的影响。方法运用网络药理学筛选葛根汤、癫痫、脑缺氧的核心靶点,分子对接验证配体与受体之间的关联度,再采用动物实验验证。用氯化锂-毛果芸香碱法建立癫痫模型,结扎单侧颈总动脉并放置在氧气含量为10%~15%的密闭箱中复制缺氧模型。60只大鼠随机分为空白组、模型组、阳性药苯妥英钠(26 g·kg^(-1))组及葛根汤高、中、低(10.62、5.31、2.655 g·kg^(-1))剂量组。治疗结束统计各组大鼠癫痫发作次数,评估大鼠神经功能恢复及癫痫发作情况;HE染色观测各组大鼠海马CA1区形态学改变;免疫组化法检测白细胞介素6(IL-6)和白细胞介素17(IL-17)的表达;靶向代谢组学检测分析大脑神经递质的改变。结果网络药理学共获得葛根汤有效成分128个,靶点228个;使用STRING构建药物-疾病蛋白互作(PPI)网络图,结果显示IL-6、STAT3、AKT1为核心靶标;GO富集在生物过程(Biological Process,BP)涉及对外来刺激的反应、氧水平、缺氧和活性氧的代谢过程的反应等。细胞组分(Cell Component,CC)方面主要对细胞膜、蛋白激酶复合物等有影响;分子功能方面(Molecular Function,MF)对泛素蛋白和泛素样蛋白连接酶结合等影响较大。KEGG分析显示葛根汤治疗脑部缺氧诱发癫痫的关键通路主要涉及脂质与动脉粥样信号通路、IL-17信号通路、Th-17信号通路等。实验验证结果发现,葛根汤治疗组大鼠的癫痫发作次数较模型组明显下降(P<0.01),大鼠海马区细胞形态结构均有不同程度改善,细胞空泡现象减少。各治疗组IL-17A、IL-6表达较模型组降低,其中葛根汤中剂量组的差异具有统计学意义(P<0.01,P<0.05)。脑部代谢组学检测,与模型组比较,葛根汤高剂量组单胺类神经递质5-羟基色氨酸(5-HTP)、去甲肾上腺素(NE)升高,色氨酸代谢产物犬尿氨�Objective Network pharmacology,molecular docking,metabolomics and experimental verification were used to investigate the effect of Gegen Decoction on epileptic seizure under cerebral hypoxia.Methods The core targets of Gegen Decoction,epilepsy and cerebral hypoxia were screened by network pharmacology,and the correlation between ligand and receptor was verified by molecular docking.Further animal experiments were carried out to verify the effect of Gegen Decoction on the rat model with epilepsy accompanied by cerebral hypoxia,which was established by lithium chlorine-pilocarpine method combined with unilateral common carotid artery ligation and placing in a closed chamber with oxygen content of 10%-15%.Sixty rats were randomly divided into blank group,model group,phenytoin sodium(26 g·kg^(-1))group,high-,medium-and low-dose(10.62,5.31,2.655 g·kg^(-1))Gegen Decoction groups.After treatment,the number of epileptic seizures rats in each group was counted,and the recovery of nerve function and epileptic seizures were evaluated.HE staining was used to observe the morphological changes of the CA1 region of hippocampus in rats.The expression of interleukin-6(IL-6)and interleukin-17(IL-17)were detected by immunohistochemistry.The changes of brain neurotransmitters were analyzed by targeted metabolomics.Results Network pharmacological results suggested that 128 active components and 228 targets were obtained.A PPI network map of drug-disease was constructed by STRING.The results showed that STAT3,IL-6 and AKT1 were the core targets.GO enrichment for biological processes(BP)involves responses to external irritant,oxygen levels,hypoxia and metabolic processes of reactive oxygen species.Cell components(CC)mainly involve in the impact on cell membrane and protein kinase complex,while molecular function(MF)involves in influence on the binding of ubiquitin-like protein and ubiquitin-like protein ligase.The key pathways of Gegen Decoction in the treatment of hypoxic-induced epilepsy in the brain,which were analyzed by KEGG,inv

关 键 词：癫痫 缺氧 葛根汤 网络药理 代谢组学 分子对接 实验验证 大鼠 

分 类 号：R285.5[医药卫生—中药学]