| 引用本文: |
伍小玲, 宁为民, 李文昭, 郭炫, 刘发生, 赵湛.基于网络药理学与细胞实验探讨天麻健脑方调控GPER/EGFR信号轴改善缺血性脑卒中的作用机制[J].湖南中医药大学学报,2025,45(8):1443-1452[点击复制] |
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| 基于网络药理学与细胞实验探讨天麻健脑方调控GPER/EGFR信号轴改善缺血性脑卒中的作用机制 |
| 伍小玲,宁为民,李文昭,郭炫,刘发生,赵湛 |
| (广州中医药大学东莞医院, 广东 东莞 523000;广州中医药大学第一临床医学院, 广东 广州 510405) |
| 摘要: |
| 目的 探讨天麻健脑方治疗缺血性脑卒中（IS）的机制以及对天麻健脑方的活性成分进行筛选。方法 通过TCMSP数据库获取天麻健脑方的活性成分和靶点，在GeneCards、DisGeNET 、OMIM获取IS相关靶点，Venny在线平台获取天麻健脑方和IS的交集靶点并通过STRING数据库和Cytoscape 3.7获得PPI网络图，利用DAVID数据库对筛选靶点进行GO分析和KEGG通路富集分析，用Cytohub插件计算前10位靶基因。通过ELISA、RT-qPCR、Western blot验证天麻健脑方对海马神经元细胞HT22中G蛋白偶联雌激素受体（GPER）/表皮生长因子受体（EGFR）/环磷酸腺苷（cAMP）/蛋白激酶A(PKA)信号通路的调控作用。结果 天麻健脑方核心成分包括黄芩素、槲皮素、四氢鸭脚木碱等，主要作用于基质金属蛋白酶9(MMP-9)、肿瘤蛋白53(TP53)、蛋白激酶B1(Akt1)等关键靶点，参与对外源性刺激的反应、对缺氧的反应、脂多糖反应等生物学过程，调控缺氧诱导因子-1A(HIF-1A)、磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)、甲状腺、雌激素等信号通路。细胞实验证实，天麻健脑方可促进氧糖剥夺/复氧（OGD/R）损伤的HT22细胞增殖（P<0.05或P<0.01），降低缺血性脑卒中标志因子MMP-9和C反应蛋白（CRP）的表达水平（P<0.05或P<0.01），上调GPER/EGFR/cAMP/PKA信号通路相关蛋白的表达（P<0.05或P<0.01）。雌激素受体抑制剂G15和ICI182780作用后，天麻健脑方对HT22细胞的作用受到不同程度的抑制。结论 天麻健脑方治疗缺血性脑卒中可能是通过激活GPER受体介导雌激素非基因组效应实现的。 |
| 关键词: 缺血性脑卒中 网络药理学 天麻健脑方 雌激素信号通路 环磷酸腺苷/蛋白激酶A |
| DOI：10.3969/j.issn.1674-070X.2025.08.007 |
| 投稿时间：2025-02-05 |
| 基金项目:东莞市名中医专家传承工作室(5000074) |
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| Mechanism of action of Tianma Jiannao Formula in ischemic stroke rehabilitation by regulating the GPER/EGFR signaling axis based on network pharmacology and cell experiments |
| WU Xiaoling, NING Weimin, LI Wenzhao, GUO Xuan, LIU Fasheng, ZHAO Zhan |
| (Dongguan Hospital, Guangzhou University of Chinese Medicine, Dongguan, Guangdong 523000, China;The First Clinical Medical College, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong 510405, China) |
| Abstract: |
| Objective To investigate the mechanism of Tianma Jiannao Formula（TMJNF） in treating ischemic stroke（IS） and to screen its active components. Methods The active components and targets of TMJNF were obtained from the TCMSP database,while IS-related targets were retrieved from GeneCards, DisGeNET, and OMIM. The intersection targets of TMJNF and IS were obtained from the Venny online platform, and the protein-protein interaction（PPI） network diagram was obtained from the STRING database and Cytoscape 3.7. The DAVID database was used for Gene Ontology（GO） analysis and Kyoto Encyclopedia of Genes and Genomes（KEGG） pathway enrichment analysis of the screened targets, and the top 10 target genes were calculated using the Cytohub plugin. The regulatory effects of TMJNF on the G protein-coupled estrogen receptor（GPER）/epidermal growth factor receptor（EGFR）/cyclic adenosine monophosphate（c AMP）/protein kinase A（PKA） signaling pathway in hippocampal neuronal HT22cells were verified by ELISA, RT-qPCR, and Western blot. Results The core components of TMJNF, including baicalein, quercetin,tetrahydroplatyphylline, mainly act on key targets such as matrix metalloproteinase-9（MMP-9）, tumor protein p53（TP53）, and protein kinaseB（AKT1）. These components were involved in biological processes such as response to exogenous stimuli, hypoxia, and lipopolysaccharide, and regulated signaling pathways including hypoxia-inducible factor-1（HIF-1）, phosphatidylinositol 3-kinaseprotein kinase B（PI3K-Akt）, thyroid hormone, and estrogen. Cellular experiments confirmed that TMJNF can promote the proliferation of HT22 cells damaged by oxygen-glucose deprivation/reoxygenation（OGD/R）（P＜0.05 or P＜0.01）, reduce the expression levels of IS marker factors matrix metalloproteinase 9（MMP-9） and C-reactive protein（CRP）（P＜0.05 or P＜0.01）, and upregulate the protein expressions related to the GPER/EGFR/cAMP/PKA signaling pathway（P＜0.05 or P＜0.01）. After treatment with estrogen receptor inhibitors G15 and ICI182780, the effect of TMJNF on HT22 cells was suppressed to varying degrees. Conclusion TMJNF may treat IS by activating the GPER receptor and mediating the non-genomic effects of estrogen. |
| Key words: ischemic stroke network pharmacology Tianma Jiannao Formula estrogen signaling pathway cAMP/PKA |
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