基于转录组学探讨滋肾清肝方改善2型糖尿病糖、脂代谢的作用机制  

Mechanism of Zishen Qinggan Prescription in Improving Glucose andLipid Metabolism in Type 2 Diabetes Based on Transcriptomics

作　　者：王景存 田春雨 张帆 喇孝瑾 吴范武 朱亮 马雷雷 WANG Jingcun;TIAN Chunyu;ZHANG Fan;LA Xiaojin;WU Fanwu;ZHU Liang;MA Leilei(North China University of Science and Technology,Tangshan 063210,China;Hebei Key Laboratory of Integrated Traditional Chinese and Western Medicine for Diabetes and Its Complications,Tangshan 063210,China;Tangshan Key Laboratory of Chinese Medicine Pharmacology,Tangshan 063210,China)

机构地区：[1]华北理工大学,河北唐山063210 [2]河北省中西医结合防治糖尿病及其并发症重点实验室,河北唐山063210 [3]唐山市中药药理重点实验室,河北唐山063210

出　　处：《中国实验方剂学杂志》2024年第8期109-117,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：河北省自然科学基金项目(H2020209235);唐山市科技计划项目(22130219H)。

摘　　要：目的:基于网络药理学及转录组学探讨滋肾清肝方(ZSQGF)改善2型糖尿病(T2DM)模型大鼠糖、脂代谢的作用机制。方法:基于网络药理学分析ZSQGF与T2DM的差异基因,进行基因本体(GO)富集分析与京都基因与基因组百科全书(KEGG)通路富集分析,并通过分子对接分析对接能,验证成分与靶点结合。通过高脂喂养及注射链脲佐菌素的方法建立T2DM大鼠模型,并随机分为正常组、模型组、二甲双胍组(二甲双胍72 mg·kg^(-1))和ZSQGF高、中、低剂量组(分别折合生药量4.8、2.4、1.2 g·kg^(-1)的药液);监测大鼠的生活状态;检测大鼠血清总胆固醇(TC)、总甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)、天冬氨酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT)含量;对肝脏组织进行苏木素-伊红(HE)染色和油红O染色;通过肝脏转录组学分析差异基因、进行GO与KEGG富集分析,绘制蛋白质-蛋白质相互作用(PPI)网络图筛选核心靶点;结合网络药理学与转录组学分析结果进行蛋白通路验证,通过蛋白免疫印迹法(Western blot)检测肝脏核转录因子-κB(NF-κB)、基质金属蛋白酶(MMP-1、MMP-9)的蛋白表达量,通过实时荧光定量聚合酶链式反应(Realtime PCR)验证B细胞淋巴瘤(Bcl)-2修正因子(BMF)、还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶4(NOX4)、脂肪酸合酶(FASN)的mRNA表达,通过免疫荧光染色检测MMP-1、MMP-9在肝脏的表达情况。结果:转录组学与网络药理学分析ZSQGF可能通过糖脂代谢通路、炎症通路作用于糖脂代谢、保护肝脏。实验证明给药8周后大鼠体质量、血糖、血清指标、病理染色结果均有改善,Western blot结果表明ZSQGF组肝脏NF-κB、MMP-1、MMP-9蛋白相对表达量降低,Real-time PCR结果表明ZSQGF高剂量组BMF、NOX4、FASN转录表达降低,免疫荧光结果ZSQGF组肝脏MMP-1、MMP-9蛋白表达量降低。结论:ZSQGF可以抑制FASN的表Objective:Based on network pharmacology and transcriptomics,the mechanism of Zishen Qinggan prescription(ZSQGF)in improving glucose and lipid metabolism in type 2 diabetes(T2DM)model rats was explored.Method:Based on network pharmacology analysis of the differential genes between ZSQGF and T2DM,gene ontology(GO)analysis and Kyoto encyclopedia of genes and genomes(KEGG)analysis were conducted,and molecular docking analysis was used to verify the binding between components and targets.A T2DM rat model was established by high-fat feeding and injection of streptozotocin(STZ).The rats were randomly divided into the control group,model group,metformin(Met,72 mg·kg^(-1))group,and ZSQGF high-,medium-,and low-dose groups(ZSQGF-H,ZSQGF-M,and ZSQGF-L,with 4.8,2.4,and 1.2 g·kg^(-1) raw drug in the solution).The living status of rats was monitored and the levels of total cholesterol(TC),total triglycerides(TG),high density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),aspartate aminotransferase(AST),and alanine aminotransferase(ALT)in rat serum were detected.The liver tissues were subjected to Hematoxylin eosin(HE)staining and oil red O staining.The differential genes were analyzed through transcriptomics,GO and KEGG analysis,and the protein-protein interaction(PPI)network was obtained to screen key targets.With network pharmacology and transcriptomics analysis results,the protein pathways were identified.The expression levels of nuclear factor-κB(NF-κB),matrix metalloproteinase(MMP)-1 and MMP-9 proteins in liver tissues were detected by Western blot.The mRNA expression of B-cell lymphoma-2(Bcl-2)modifying facto(r BMF),nicotinamide adenine dinucleotide phosphate(NADPH)oxidase 4(NOX4),and fatty acid synthase(FASN)was detected by real-time polymerase chain reaction(Real-time PCR).The expression of MMP-1 and MMP-9 in the liver was detected by immunofluorescence staining.Result:Transcriptomics and network pharmacology analysis suggested that ZSQGF may protect the liver through the glucose and lipid

关 键 词：滋肾清肝方 2型糖尿病 糖脂代谢 网络药理学 转录组学 

分 类 号：R284[医药卫生—中药学] R285[医药卫生—中医学] R289R287R22R2-031R33R24