基于网络药理学和分子对接探究白杨素和柚皮素抗PEDV的作用机制及试验验证

doi:10.16431/j.cnki.1671-7236.2024.04.043

摘要/Abstract

摘要： 【目的】探讨白杨素和柚皮素抗猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)的作用靶点和机制,为进一步以白杨素和柚皮素开发新型抗PEDV的治疗药物提供理论依据。【方法】利用PharmMapper、TCMSP、SEA Search Server和STITCH在线数据库获得白杨素和柚皮素的潜在作用靶点,同时检索GeneCards数据库获得PEDV对宿主的作用靶点,利用在线程序Draw Venny Diagram获取白杨素、柚皮素与疾病的交集靶点。通过STRING数据库和Cytoscape 3.9.1软件构建靶蛋白互作(PPI)网络并筛选关键靶点,对靶点进行GO功能和KEGG通路富集分析。通过AutoDock Vina v 1.2.0软件对核心靶点及小分子进行分子对接,并分析白杨素和柚皮素与靶蛋白的结合能和结合模式,利用PyMOL v 2.5实现对接结果的可视化。采用实时荧光定量PCR和Western blotting检测白杨素和柚皮素对核心靶蛋白表达的影响。【结果】白杨素潜在抗PEDV的靶点有12个,其中白蛋白(ALB)、雌激素受体1(ESR1)、转化生长因子β-1蛋白(TGF-β1)可能是白杨素抗PEDV的核心靶点;柚皮素潜在抗PEDV的靶点有18个,其中ALB、胱天蛋白酶3(Caspase-3,CASP3)、过氧化物酶体增殖物激活受体γ(PPARG)可能是柚皮素抗PEDV的核心靶点。白杨素抗PEDV靶点涉及21种生物过程、5种细胞组分和6种分子功能,共获得11条信号通路;柚皮素抗PEDV靶点涉及31种生物过程、8种细胞组分和19种分子功能,共获得13条信号通路。核心靶点与两种天然化合物之间以氢键和疏水作用力为主,有较强的相互作用。白杨素和柚皮素能极显著降低Caspase-3表达量(P＜0.01),可能通过影响Caspase-3的表达和活化来颉颃PEDV诱导的细胞凋亡;极显著升高TGF-β1蛋白表达量(P＜0.01),可能通过影响TGF-β1的表达抗PEDV诱导的炎症反应而治疗PEDV的感染。【结论】本研究揭示了白杨素和柚皮素分别通过潜在核心靶点ALB、ESR1、TGF-β1和ALB、Caspase-3、PPARG,以及IL17、PI3K-Akt和AMPK信号通路发挥抗PEDV作用的机制,为新型抗PEDV药物的研发提供新的思路。

关键词: 猪流行性腹泻病毒(PEDV); 网络药理学; 分子对接; 白杨素; 柚皮素

Abstract: 【Objective】 This study was objective to explore the target and mechanism of chrysin and naringenin against Porcine epidemic diarrhea virus (PEDV),and provide theoretical basis for further developing new anti-PEDV therapeutic drugs with chrysin and naringenin.【Method】 Online databases of PharmMapper,TCMSP,SEA Search Server and STITCH were used to obtain the potential targets of chrysin and naringenin,at the same time GeneCards database was searched to obtain the targets of PEDV on the host,and Draw Venny Diagram online program was used to obtain the intersection targets of chrysin,naringenin and disease.The target protein-protein interaction (PPI) network was constructed by STRING database and Cytoscape 3.9.1 software,and the key targets were screened,and the GO function and KEGG pathway enrichment of the targets were analyzed.The molecular docking of core targets and small molecules was carried out by AutoDock Vina v 1.2.0 software,and the binding energy and binding mode of chrysin and naringenin with target proteins were analyzed,and the docking results were visualized by PyMOL v 2.5.The effects of chrysin and naringenin on the expression of core target proteins were detected by Real-time quantitative PCR.【Result】 The results showed that there were 12 potential anti-PEDV targets of chrysin,among which albumin (ALB),estrogen receptor 1 (ESR1) and transforming growth factor β-1 protein (TGF-β1) might be the core targets of chrysin against PEDV.Naringenin had 18 potential anti-PEDV targets,among which ALB,Caspase-3 (CASP3) and peroxisome proliferator-activated receptor γ (PPARG) might be the core targets of naringenin against PEDV.The anti-PEDV target of chrysin involved 21 biological processes,5 cellular components and 6 molecular functions and obtained 11 signal pathways,while naringenin anti-PEDV targets involved 31 biological processes,8 cellular components and 19 molecular functions, and obtained 13 signal pathways.There was a strong interaction between the core targets and the two natural compounds due to hydrogen bonding and hydrophobic interaction.Chrysin and naringenin could extremely significantly reduce the expression of Caspase-3 (P<0.01),which might antagonize the apoptosis induced by PEDV by affecting the expression and activation of Caspase-3,and they extremely significantly increased the expression of TGF-β1 protein (P<0.01),which might treat PEDV infection by affecting the expression of TGF-β1 and anti-PEDV-induced inflammation.【Conclusion】This study revealed the mechanism of anti-PEDV effects of chrysin and naringenin through potential core targets ALB,ESR1,TGF-β1 and ALB, Caspase-3,PPARG,respectively, and IL17,PI3K-Akt and AMPK signaling pathways, providing new ideas for the development of novel anti-PEDV drugs.

Key words: Porcine epidemic diarrhea virus (PEDV); network pharmacology; molecular docking; chrysin; naringenin

中图分类号:

S853.74

植玉鹏, 刘雨桐, 陈弟诗, 宫萌菲, 夏学妹, 任玉鹏. 基于网络药理学和分子对接探究白杨素和柚皮素抗PEDV的作用机制及试验验证[J]. 中国畜牧兽医, 2024, 51(4): 1757-1772.

ZHI Yupeng, LIU Yutong, CHEN Dishi, GONG Mengfei, XIA Xuemei, REN Yupeng. Study on the anti-PEDV Mechanism and Experimental Verification of Chrysin and Naringenin Based on Network Pharmacology and Molecular Docking[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(4): 1757-1772.

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