摘要
为了探讨甘草多糖(GPS)抗急性肝损伤(ALI)的作用靶点和机制,本试验采用网络药理学和体内试验相结合的方法进行研究,首先利用中药系统药理学数据库分析平台(TCMSP)、SwissTargetPrediction数据库收集GPS的靶点,利用GeneCards和DisGeNET数据库收集ALI的靶点,使用String数据库和Cytoscape软件构建GPS-ALI-靶点网络图,利用基因功能分类数据库(DAVID)和RStudio软件进行基因本体(GO)功能富集和京都基因与基因组百科全书(KEGG)通路分析,利用Autodock Tool分子对接技术验证GPS与靶蛋白的结合力;体内试验中,将48只C57BL/6J小鼠分为6个组:正常组、LPS组、阳性组、高糖组、中糖组和低糖组,高、中、低糖组小鼠分别以400、200、100 mg/(kg·bw·d)剂量GPS连续灌胃20 d,其余组灌胃等量生理盐水,第28天,除正常组外,其余组小鼠腹腔注射LPS[5 mg/(kg·bw·d)]建立小鼠ALI模型;2 h后阳性组小鼠灌胃地塞米松[5 mg/(kg·bw·d)],LPS处理8 h后,称重,收集小鼠肝脏组织和血清,酶联免疫吸附试验(ELISA)法检测生化指标,苏木精-伊红(H.E.)染色观察肝脏组织病理学变化,实时荧光定量聚合酶链式反应(qPCR)检测炎症因子及相关通路蛋白的基因表达水平。网络药理学分析结果显示,共获得27个GPS和ALI的共同靶点,10个关键靶点均富集于氧化应激分子机制和肿瘤坏死因子(TNF)通路中;且GPS与Toll样受体4(TLR4)和基质金属蛋白酶9(MMP9)结合较强。体内试验结果显示,ALI小鼠模型建立成功,GPS预处理可缓解肝脏组织损伤,与LPS组相比,阳性组、中糖组和高糖组小鼠血清中天冬氨酸氨基转移酶(AST)和谷氨酸氨基转移酶(ALT)活性显著降低(P<0.05),阳性组和高糖组小鼠肝脏组织中谷胱甘肽过氧化物酶(GSH-Px)活性显著升高(P<0.05),丙二醛(MDA)含量显著降低(P<0.05),阳性组、中糖组和高糖组小鼠肝脏组织中促炎因子肿瘤坏死因子(TNF-α)、白细胞介素-1β(IL-1β)和白细胞介素-6(IL-6)mRNA相对表达量均显著降低(P<0.05),MMP9 mRNA相对表达量均显著升高(P<0.05),TLR4表达量显著降低(P<0.05)。结果表明,GPS可能通过缓解氧化应激反应,介导MMP9/TLR4/TNF-α途径来减轻炎症反应,实现护肝作用。本试验可为后期以GPS为主要活性成分的保肝药物或功能食品的研发提供一定的参考依据。
To explore the therapeutic targets and mechanisms of Glycyrrhiza glabra polysaccharide(GPS)in combating acute liver injury(ALI),this study combined network pharmacology with in vivo experiments.Potential GPS targets were identified using the TCMSP and SwissTargetPrediction databases,and ALI-related targets were obtained from GeneCards and DisGeNET.A GPSALI-target interaction network was constructed using the String database and Cytoscape software.GO functional enrichment and KEGG pathway analysis were performed with DAVID and RStudio.Molecular docking was conducted using AutoDock Tools to assess the binding affinity between GPS and core target proteins.In the in vivo study,48 C57BL/6J mice were divided into six groups:Control,LPS model,Positive control,and High-,Medium-,and Low-dose GPS groups.Mice in the GPS groups were gavaged with 400,200,or 100 mg/(kg·bw·d)GPS for 20 consecutive days,while the others received normal saline.On day 28,all mice except those in the Control group were intraperitoneally injected with LPS[5 mg/(kg·bw·d)]to induce ALI.Two hours later,the Positive control group received dexamethasone[5 mg/(kg·bw·d)]by gavage.After 8 hours of LPS treatment,body weights were recorded,and serum and liver tissues were collected.Biochemical indicators were assessed via ELISA,histopathological changes were observed by H.E.staining,and qPCR was used to detect expression levels of inflammatory cytokines and pathway-related genes.Network pharmacology identified 27 common targets between GPS and ALI,with 10 key targets enriched in oxidative stress mechanisms and the TNF signaling pathway.GPS showed strong binding affinity with Toll-like receptor 4(TLR4)and matrix metalloproteinase 9(MMP9).In vivo results confirmed successful ALI modeling and demonstrated that GPS pretreatment alleviated liver injury.Compared with the LPS group,serum AST and ALT levels were significantly reduced in the Positive control,Medium-,and High-dose groups(P<0.05).GSH-Px activity was significantly increased and MDA levels decreased in the Positive and High-dose groups(P<0.05).The mRNA levels of pro-inflammatory cytokines TNF-α,IL-1β,and IL-6 were significantly downregulated in liver tissues of the Positive,Medium-,and High-dose groups(P<0.05),while MMP9 mRNA expression was significantly upregulated(P<0.05),and TLR4 expression was significantly reduced(P<0.05).These results suggest that GPS may exert a hepatoprotective effect by mitigating oxidative stress and modulating the MMP9/TLR4/TNF-αsignaling pathway to suppress inflammation.This study provides a scientific basis for the development of GPS-based hepatoprotective drugs or functional foods.
作者
张椰莉
李嘉晨
李娜
李美艳
董晓婷
武晓英
ZHANG Yeli;LI Jiachen;LI Na;LI Meiyan;DONG Xiaoting;WU Xiaoying(School of Biological Sciences and Technology,Taiyuan Normal University,Jinzhong 030619,China;Institute of Basic and Translational Medicine,Xi'an Medical University,Xi'an 710069,China)
出处
《中国兽医杂志》
北大核心
2025年第8期95-106,共12页
Chinese Journal of Veterinary Medicine
基金
山西省重点研发计划重点项目(201603D21109-1)
山西省重点研发计划项目(201603D221008-2)
山西省基础研究计划项目(202103021223327)。
关键词
甘草多糖
急性肝损伤
保护作用
网络药理学
Glycyrrhiza glabra polysaccharide
acute liver injury
protective effect
network pharmacology
