基于网络药理学和实验研究探...肪肝小鼠模型影响及作用机制_徐永妘.pdf

世界中西医结合杂志 2023 年第 18 卷第 1 期World Journal of Integrated Traditional and Western Medicine2023，Vol.18，No.1药物研究DOI:10 13935/j cnki sjzx 230112基金项目:上海市卫健委项目(2020JQ005)作者单位:1 上海中医药大学附属市中医医院内科，上海 200071;2 上海中医药大学附属曙光医院内科，上海201203;3 上海市虹口区欧阳路街道社区卫生服务中心 全科病房，上海 200081;4 上海中医药大学附属市中医医院肝病科，上海 200071;5 上海中医药大学附属曙光医院肝病科，上海 201203通信作者:卓蕴慧，Email:bengpifenm8c163 com基于网络药理学和实验研究探讨加味甘姜苓术汤对非酒精性脂肪肝小鼠模型影响及作用机制徐永妘1张磊2张宇3高司成4卓蕴慧5【摘要】目的甘姜苓术汤是金匮要略名方，增加丹参、小茴香二位药物形成加味方以提升疗效，本研究通过网络药理学、分子对接、动物实验证实其抗脂肪肝的影响和机制。方法通过数据库，寻找加味方和脂肪肝的共同靶点，做蛋白互作图，进行 GO 富集分析和 KEGG 通路预测。对主要化合物和主要蛋白做分子对接。动物实验中，C57BL6 小鼠 60%脂肪功能造模饲料造模。对照组无给药，低剂量、中剂量、高剂量加味甘姜苓术汤组采用加味甘姜苓术汤灌胃给药。原方组采用原方灌胃，阳性药组采用易善复灌胃，模型组采用同等剂量纯净水灌胃。测定肝指数，HE 染色、油红 O 染色切片观察肝组织形态学改变，ELISA 检测肝脏氧化还原水平，自动生化仪检测血脂水平。Western blot 验证网络药理学和分子对接的相关通路蛋白表达水平。结果网络药理学研究发现疾病与药物成分靶点共同靶点 98 个。蛋白互作 PPI 分析证实 JUN，AKT1，MAPK1，MAPK3，STAT3，HSP90AA1 等为核心基因。KEGG 通路预测了 HIF 1、MAPK、NOD、TNF、TL 等信号通路。分子对接证实主要化合物和蛋白之间连接紧密。动物实验证实，4 周给药后，加味方、原方、阳性药均减少小鼠体重，加味方中、高剂量组降低体质量的疗效高于原方组(P 0 05，P 0 001)，加味方高剂量组降低体质量的疗效高于阳性药组(P 0 01)。油红 O 染色证实，加味方促进肝组织脂滴减少。HE 染色证实，加味方改善肝细胞形态，减少了脂肪，与原方组比较，中剂量组NAS 评分更低(P 0 05)。肝功能及脂质代谢指标中，与模型组比较，加味方低、中、高剂量组、原方、阳性药均有效降低肝指数、ALT、AST、TC、TG、LDL C 含量(P 0 05，P 0 01，P 0 001)，提升了 HDL C 含量(P 0 01，P0 001)。与原方组比较，加味方中、高剂量组更能降低肝指数、ALT、AST、TC、TG、LDL C(P 0 05，P 0 01，P0 001);中、高剂量组更能升高 HDL C 指标(P 0 01，P 0 001);证实加味方应用提升了护肝、抗脂质代谢紊乱疗效。加味方低、中、高剂量组能促进肝组织中的 GSH 水平提升(P 0 05，P 0 001)，降低 TNF 、IL 6、IL1、MDA、OS 水平(P 0 001)，发挥抗炎、调节氧化还原水平的作用。Western blot 实验证实，加味方低、中、高剂量组 TL4 蛋白水平下降(P 0 001)。p P38 蛋白水平下降(P 0 05，P 0 01，P 0 001)，证实加味方抑制了肝组织 TL4 P38 MAPK 信号通路激活。结论加味甘姜苓术汤对非酒精性脂肪肝小鼠模型具有促进恢复血脂代谢平衡，调节氧化还原水平、减轻脂肪堆积、减轻体重的作用，其效果优于甘姜苓术汤原方。TL4 P38 信号通路参与了加味甘姜苓术汤对非酒精性脂肪肝的作用机制。【关键词】非酒精性脂肪肝;甘姜苓术汤;血脂;炎症;氧化还原【中图分类号】285 6【文献标识码】AEffect of Modified Ganjiang Lingzhu Decoction on Nonalcoholic Fatty Liver in Miceand Its Mechanism Based on Network Pharmacology and ExperimentsXU Yong yun1，ZHANG Lei2，ZHANG Yu3，GAO Si cheng4，ZHUO Yun hui5(1 Department of Internal Medicine，Municipal Hospital of Traditional Chinese Medicine，Shanghai University of Tradition-al Chinese Medicine，Shanghai 200071;2 Department of Internal medicine，Shuguang Hospital Affiliated to Shanghai Uni-versity of Traditional Chinese Medicine，Shanghai 201203;3 General Ward of Community Health Service Center in Ouyangoad Street of Hongkou District，Shanghai 200081;4 Department of Hepatology，Municipal Hospital of Traditional Chinese47世界中西医结合杂志 2023 年第 18 卷第 1 期World Journal of Integrated Traditional and Western Medicine2023，Vol.18，No.1Medicine，Shanghai University of Traditional Chinese Medicine，Shanghai 200071;5 Department of Hepatology，ShuguangHospital Affiliated to Shanghai University of Traditional Chinese Medicine，Shanghai 201203)【Abstract】ObjectiveGanjiang Lingzhu Decoction is a famous formula in Essentials from the Golden CabinetSalviae Miltiorrhizae adix et hizoma and Foeniculi Fructus were added to compose the modified formula and improve thecurative effect This study aimed to explore the effect of Ganjiang Lingzhu Decoction against fatty liver and its mechanismthrough network pharmacology，molecular docking，and animal experiments MethodsThrough databases，the common tar-gets of the modified Ganjiang Lingzhu Decoction and fatty liver were screened out，and the protein protein interaction(PPI)map was made Gene Ontology(GO)enrichment analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis were performed Molecular docking was performed between main compounds and proteins Inanimal experiments，C57BL6 mice were modeled with 60%fat functional modeling feed The control group was not givendrugs，and the low dose，medium dose，and high dose modified formula groups were given modified formula by gavageThe original formula group was perfused with the original formula，the positive drug group was perfused with Essentiale，andthe model group was perfused with the same dose of purified water Liver index was measured，and liver morphologicalchanges were observed by hematoxylin eosin(HE)staining and oil red O staining The liver redox levels were determinedby enzyme linked immunosorbent assay(ELISA)，and the blood liquid levels were determined by automatic biochemicalinstrument Western blot was used to explore the protein expression levels of pathways related to network pharmacology andmolecular docking esultsNinety eight common targets of diseases and drug components were found in the research ofnetwork pharmacology PPI analysis showed that JUN，protein kinase B1(AKT1)，mitogen activated protein kinase(MAPK)1，MAPK3，signal transducers and activators of transcription 3(STAT3)，and heat shock protein 90 alpha familyclass a member 1(HSP90AA1)were hub genes KEGG pathway enrichment analysis predicted hypoxia inducible factor 1(HIF 1)，MAPK，nucleotide binding oligomerization domain(NOD)，tumor necrosis factor(TNF)，Toll like receptor(TL)，and other signaling pathways Molecular docking demonstrated that the main compounds and proteins were closelylinked Animal experiments confirmed that after 4 weeks of administrat