南方医科大学学报

南方医科大学学报 ›› 2021, Vol. 41 ›› Issue (1): 10-19.doi: 10.12122/j.issn.1673-4254.2021.01.02

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白藜芦醇治疗阿尔茨海默病的作用机制：基于网络药理学方法

方迎艳，苏振宏，司文霞，刘圆呈，李 洁，曾 鹏   

  • 出版日期:2021-01-26 发布日期:2021-01-25

Network pharmacology-based study of the therapeutic mechanism of resveratrol for Alzheimer's disease

  • Online:2021-01-26 Published:2021-01-25

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摘要：

目的 基于网络药理学方法探讨白藜芦醇（RES）治疗阿尔茨海默病（AD）的作用机制。方法 利用PubChem、BATMAN-TCM、Genecards、AD、TTD、String 11.0、AlzData、SwissTargetPrediction、Metascape等数据库，获得RES所有作用靶点和人类AD相关的靶点，使用Venny 2.1工具取交集，获得RES治疗AD的靶点。构建靶点蛋白质相互作用网络，基因本体论（GO）与KEGG通路富集分析，使用Cytoscape 3.7.1构建RES治疗AD的“靶点-信号通路”网络图。使用SwissDock（http://www.swissdock.ch/docking）服务器进行分子对接验证。通过蛋白免疫印迹检测RES处理AD细胞模型（293Tau细胞）的pS396、pS199、Tau5、CDK5、糖原合酶激酶3β（GSK3β）和p-GSK3β的蛋白水平。结果 分析得到RES作用靶点182个，AD相关靶点525个，RES治疗AD的靶点36个，其中34.6%的靶点为蛋白质修饰酶、27.7%的靶点为代谢物相互转化酶、13.8%的靶点为基因特异性转录调节因子、10.3%的靶点为转运蛋白。RES治疗AD的核心关键靶点为INS、APP、ESR1、MMP9、IGF1R、CACNA1C、MAPT（即微管相关蛋白Tau）、MMP2、TGFB1与GSK3B。GO生物学过程富集分析显示，RES治疗AD主要涉及对β-淀粉样蛋白的反应、转移酶活性的正调节、跨膜受体蛋白酪氨酸激酶信号通路、行为、学习或记忆、老化、跨膜转运的调控等多个生物功能。KEGG通路富集分析表明富集最显著的信号通路为阿尔茨海默病通路、PI3K-AKT信号通路、cGMP-PKG信号通路、MAPK信号通路等。分子对接结果显示，RES与ESR1、GSK3B、MMP9、IGF1R、APP与INS结合较好。蛋白免疫印迹结果显示，50 μmol/L RES处理12 h可以通过调控CDK5、核心靶点GSK3β活性显著降低293Tau细胞pS396和pS199水平（P<0.001）。结论 RES可以通过Aβ、Tau病理过程治疗AD，可通过调控CDK5与GSK3β改善AD病理改变。

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Abstract:

Objective To investigate the therapeutic mechanism of resveratrol (RES) for Alzheimer's disease (AD) in light of network pharmacology. Methods We searched PubChem, BATMAN-TCM, Genecards, AD, TTD, String 11.0, AlzData, SwissTargetPrediction, Metascape and other databases for the therapeutic targets of RES and human AD-related targets. The intersection was determined using Venny 2.1 to obtain the therapeutic targets of RES for AD. The protein-protein interaction (PPI) network was constructed, the gene ontology (GO) was enriched and the Kyoto Encyclopedia of Genes and Genomes pathway (KEGG pathway) were analyzed. Cytoscape 3.7.1 software was used to construct a target-signaling pathway network of RES in the treatment of AD. Molecular docking verification was carried out on SwissDock (http://www.swissdock.ch/docking). We examined a 293Tau cell model of AD for changes in protein levels of pS396, pS199, Tau5, CDK5, glycogen synthase kinase 3β (GSK3β) and p-GSK3β in response to RES treatment using Western blotting. Results We obtained 182 targets of RES, 525 targets related to AD, and 36 targets of RES for AD treatment, among which 34.6% of the targets were protein-modifying enzymes, 27.7% were metabolite invertase, 13.8% were gene-specific transcriptional regulators, and 10.3% were transporters. The core key targets of RES in the treatment of AD included INS, APP, ESR1, MMP9, IGF1R, CACNA1C, MAPT (microtubule- associated protein Tau), MMP2, TGFB1 and GSK3B. Enrichment analysis of GO biological process suggested that the biological function of RES in AD treatment mainly involved the response to β-amyloid protein, positive regulation of transferase activity, the transmembrane receptor protein tyrosine kinase signaling pathway, regulation of behavior, learning or memory, aging, and transmembrane transport. KEGG pathway enrichment analysis showed that the most significantly enriched signaling pathways were AD pathway, PI3K-AKT signaling pathway, cGMP-PKG signaling pathway, and MAPK signaling pathway. Molecular docking results showed that RES had strong binding with ESR1, GSK3B, MMP9, IGF1R, APP and INS. In the cell model of AD, treatment with 50 μmol/L RES for 12 h significantly reduced the levels of pS396 and pS199 by regulating CDK5 and GSK3β activity (P<0.001). Conclusion RES produces
therapeutic effects on AD by acting on multiple targets and affecting multiple signaling pathways and improves AD-associated pathologies via a direct action on Aβ and Tau pathological processes.

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