黄芪甲苷调控细胞焦亡减轻肠缺血再灌注损伤的分子作用机制  

Molecular Mechanism of Action of AstragalosideⅣin Modulating Pyroptosis to Attenuate Intestinal Ischemia-reperfusion Injury Based on Experimental Validation

作　　者：常利娅 冷玉芳[2] 赵紫岑 王玉 邢阳 李东斌 CHANG Liya;LENG Yufang;ZHAO Zicen;WANG Yu;XING Yang;LI Dongbin(The First Clinical Medical College of Lanzhou University,Lanzhou 730000,China;The First Hospital of Lanzhou University,Lanzhou 730000,China)

机构地区：[1]兰州大学第一临床医学院,兰州730000 [2]兰州大学第一医院,兰州730000

出　　处：《中国实验方剂学杂志》2024年第19期116-123,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(82260381);兰州大学医学教育创新发展项目(lzuyxcx-2022-108)。

摘　　要：目的:采用网络药理学和体内实验结合的方法研究黄芪甲苷(AS-Ⅳ)调控细胞焦亡(Pyroptosis)减轻肠缺血再灌注损伤(IRI)的作用机制。方法:首先从中药系统药理数据库和分析平台(TCMSP)数据库和Swiss Target Prediction数据库中检索获得AS-Ⅳ的对应靶点基因,并从GeneCards数据库中检索得到肠IRI和Pyroptosis相关的靶点基因,通过解螺旋网站绘制Venn图,获得三者的共同靶点基因。通过STRING数据库、Cytoscape软件构建蛋白-蛋白相互作用(PPI)网络筛选共同靶基因并导入Cytoscape软件获得核心靶基因。微生信平台用于基因本体(GO)和京都基因与基因组百科全书(KEGG)富集分析并预测AS-Ⅳ调控Pyroptosis减轻肠IRI的作用机制。然后采用随机数字表法,将C57/BL6J小鼠随机分为5组,正常组、模型组、给药组(IR+AS-Ⅳ)、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)激动剂NSS组(IR+AS-Ⅳ+NSS)、NLRP3抑制剂MCC950组(IR+AS-Ⅳ+MCC950)。模型组、给药组、NLRP3激动剂NSS组、NLRP3抑制剂MCC950组采用夹闭肠系膜上动脉45 min恢复灌注2 h建立肠IRI模型,正常组仅分离血管不夹闭。给药组、NLRP3激动剂NSS组、NLRP3抑制剂MCC950组于造模前连续3 d灌胃溶于0.1%二甲基亚砜的黄芪甲苷(50 mg·kg^(-1)),末次灌胃时间为造模前2 h,其余2组灌胃等量的生理盐水。NLRP3激动剂NSS组于造模前1 h腹腔注射4 mg·kg^(-1)的NSS,NLRP3抑制剂MCC950组于造模前1 h腹腔注射10 mg·kg^(-1)的MCC950。再灌注2 h处死小鼠,获取肠组织。采用酶联免疫吸附测定法(ELISA)检测白细胞介素(IL)-18,IL-1β水平,蛋白免疫印迹法(Western blot)检测硫氧还蛋白互作蛋白(TXNIP)、NLRP3、胱天蛋白酶(Caspase)-1、焦孔素D(GSDMD)蛋白表达,Chiu's评分评估小鼠肠组织病理学变化。结果:网络药理学分析表明肠IRI的靶点有1599个,AS-Ⅳ的作用靶点有199个,Pyroptosis的靶点有197个,三者共同靶点有20个。核心靶点有10个,�Objective:To investigate the mechanism of astragaloside-Ⅳ(AS-Ⅳ)in regulating pyroptosis to alleviate intestinal ischemia-reperfusion injury(IRI)by combining network pharmacology and in vivo experiments.Method:Firstly,the corresponding target genes of AS-Ⅳ were obtained from TraditionalChineseMedicineSystemsPharmacology(TCMSP)database and Swiss Target Prediction database,and the target genes related to intestinal IRI and Pyroptosis were obtained from GeneCards database,and the common target genes of the three were obtained by drawing Venn diagrams through unspiralized website.Proteinprotein interaction(PPI)network was constructed by STRING database and Cytoscape software to screen common target genes and imported into Cytoscape software to obtain core target genes.Microbiotics platform was used for gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)enrichment analysis and prediction of the mechanism of action of AS-Ⅳ in regulating Pyroptosis to alleviate intestinal IRI.Then C57/BL6J mice were randomly divided into 5 groups normal group,model group(IR),drug administration group(IR+AS-Ⅳ),nucleotide-binding oligomerization structural domain-like receptor protein 3(NLRP3)agonist NSS group(IR+AS-Ⅳ+NSS),and NLRP3 inhibitor MCC950 group(IR+AS-Ⅳ+MCC950)by using a randomized numerical table method.The intestinal IRI model was established by clamping the superior mesenteric artery for 45 min and resuming perfusion for 2 h in the model group,the drug administration group,the NLRP3 agonist NSS group,and the NLRP3 inhibitor MCC950 group,and the normal group was only separated from the vessels without clamping.The administration group,the NLRP3 agonist NSS group,and the NLRP3 inhibitor MCC950 group were gavaged with astragaloside dissolved in 0.1%dimethylsulfoxide(50 mg·kg^(-1))for 3 consecutive days before modeling,with the last gavage 2 h before modeling,and the remaining two groups were gavaged with equal amounts of saline.The NLRP3 agonist NSS group was injected intraperitoneally with 4 mg·kg^

关 键 词：黄芪甲苷 肠缺血再灌注损伤 硫氧还蛋白结合蛋白(TXNIP)/核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)信号通路 细胞焦亡 网络药理学 

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