基于网络药理学的中药复方醒脑治瘫胶囊治疗气虚血瘀型中风的作用机制研究与实验验证

Study on the mechanism of action of traditional Chinese medicine Xingnao Zhitan Capsule in the treatment of blood stasis due to qi deficiency type of stroke based on network pharmacology and experimental validation

关键词:  醒脑治瘫胶囊  气虚  血瘀  中风  网络药理学  分子对接

Key words:  Xingnao Zhitan Capsule  qi deficiency  blood stasis  stroke  network pharmacology  molecular docking

摘要:

[目的] 中药复方醒脑治瘫胶囊在临床上被广泛用于治疗各种气虚血瘀型中风，但其整体药理学的作用机制研究相对较少。本文通过网络药理学、分子对接以及实验验证手段探索醒脑治瘫胶囊的整体药理作用机制。[方法] 检索中药系统药理学（TCMSP）数据库和中医药信息数据库（TCM-ID）中醒脑治瘫胶囊各味中药口服生物利用度大于30%，类药性大于0.18的有效化学成分及其对应的作用靶点；在DrugBank、GeneCards、OMIM、TTD等数据库中检索气虚血瘀型中风对应的靶点；使用Cytoscape软件构建醒脑治瘫胶囊中药-活性成分-气虚血瘀型中风潜在靶点网络；使用STRING软件构建蛋白-蛋白相互作用网络；使用DAVID软件进行靶点基因本体（GO）功能富集分析和京都基因与基因组百科全书（KEGG）通路富集分析；使用AutoDock Vina和Discovery Studio Visualizer软件进行分子对接和作用模式分析；评价醒脑治瘫胶囊提取物对氧糖剥夺/复糖复氧（OGD/R）诱导人源性神经母细胞瘤细胞（SH-SY5Y）生存率、乳酸脱氢酶（LDH）释放率和网络药理学分析出的部分相关基因的影响。[结果] 网络药理学分析显示PTGS2、PTGS1、HSP90AA1、NCOA2、ADRB2、CHRM1等靶点是醒脑治瘫胶囊发挥药效的潜在主要靶点；槲皮素、山柰酚、木犀草素等化合物是醒脑治瘫胶囊发挥药理作用的潜在有效成分。分子对接显示木犀草素与β2-肾上腺素能受体（ADRB2）、槲皮素与环氧合酶-2（PTGS2）、黄芩素与M1受体（CHRM1）、隐丹参酮与环氧合酶-1（PTGS1）具有潜在的结合可能。醒脑治瘫胶囊提取物可剂量依赖性地修复OGD/R对SH-SY5Y细胞的损伤，提高细胞生存率，降低LDH释放率，并能下调OGD/R引起的细胞PTGS2/PTGS1/HSP90AA1基因的mRNA和蛋白的高表达。[结论] 网络药理学研究结果、分子对接技术结合实验验证为阐明醒脑治瘫胶囊药效物质基础和整体药理机制提供了新的研究方向。

Abstract:

[Objective] The traditional Chinese medicine formula Xingnao Zhitan Capsule has been widely used in the treatment of various types of strokes with blood stasis due to qi deficiency，but its overall pharmacological mechanism of action has been relatively little studied. In this paper，we explored the overall pharmacological mechanism of Xingnao Zhitan Capsule using network pharmacology，molecular docking，and experimental validation. [Methods] It was searched in TCMSP and TCM-ID databases for active chemical components with oral bioavailability greater than 30% and drug-like properties greater than 0.18 and their corresponding targets of action for each flavor of traditional Chinese medicine in Xingnao Zhitan Capsule. It was searched in DrugBank，GeneCards，OMIM，TTD and other databases for targets corresponding to blood stasis due to qi deficiency type of stroke. A potential target network of Chinese medicine-active ingredient- blood stasis due to qi deficiency type of stroke was constructed using Cytoscape software for Xingnao Zhitan Capsule. A protein-protein interaction networks was constructed using STRING software. Target GO function enrichment analysis and KEGG pathway enrichment analysis were performed using the DAVID program. Software such as AutoDock Vina and Discovery Studio Visualizer were used to perform molecular docking and analyze patterns of action. The effects of the extract of Xingnao Zhitan Capsules on human neuroblastoma cells（SH-SY5Y） injury induced by oxygen-glucose deprivation/reoxygenation（OGD/R），such as cell survival rate，lactate dehydrogenase（LDH） release rate，and the regulation of some genes expression. [Results] Network pharmacological analysis showed that PTGS2，PTGS1，HSP90AA1，NCOA2，ADRB2、CHRM1 are the potential main targets for the efficacy of Xingnao Zhitan Capsules. Compounds such as quercetin，kaempferol，luteolin，and other compounds are the potential active ingredients in the pharmacological effect of Xingnao Zhitan Capsule. Molecular docking revealed potential binding possibilities of Luteolin to β2-adrenergic receptor（ADRB2），quercetin to cyclooxygenase-2（PTGS2），baicalein to M1 receptor（CHRM1），and cryptotanshinone to cyclooxygenase-1（PTGS1）. The extract of Xingnao Zhitan Capsules dose-dependently repaired OGD/R damage to SH-SY5Y cells，increased cell survival，decreased LDH release rate and down-regulated OGD/R-induced high expression of cellular PTGS2/PTGS1/HSP90AA1 mRNA and protein. [Conclusion] The results of network pharmacology research，molecular docking combined with experimental validation provide a new research direction for elucidating the material basis of the efficacy of Xingnao Zhitan Capsules and the overall pharmacological mechanism.