关白附与禹白附对中风沙鼠影响差异的尿液代谢组学分析  

Urinary Metabolomics Aanlysis of Differences in Effect of Aconiti Coreani Radix and Typhonii Rhizoma on Gerbils with Stroke

作　　者：周丽婷 曾琬婷 贾茹 徐慧颖 丁奕辉 董昊 马浩文 曲扬[1] 才谦[1] ZHOU Liting;ZENG Wanting;JIA Ru;XU Huiying;DING Yihui;DONG Hao;MA Haowen;QU Yang;CAI Qian(College of Pharmacy,Liaoning University of Traditional Chinese Medicine,Dalian 116600,China)

机构地区：[1]辽宁中医药大学药学院,辽宁大连116600

出　　处：《中国实验方剂学杂志》2024年第8期157-166,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：“兴辽英才”计划科技创新领军人才(特聘教授)项目(XLYC2002004)。

摘　　要：目的:采用非靶向代谢组学技术探究关白附与禹白附对中风沙鼠尿液代谢产物的影响差异,进而明确二者作用机制及其异同。方法:将24只沙鼠随机分成空白组、模型组、关白附组、禹白附组。除空白组外,其余各组均采用右侧单侧结扎沙鼠颈动脉构建缺血性脑卒中模型。各给药组分别灌胃对应的药材全粉混悬液(0.586 mg·g^(-1)),共给药14 d。给药第0、3、7、14天,采用Longa评分法对各组沙鼠进行神经功能缺损评分。给药结束后收集各组沙鼠血清、脑组织及尿液,采用2,3,5-氯化三苯四氮唑(TTC)染色法检测脑梗死率,利用酶联免疫吸附测定法(ELISA)对血清和脑组织中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、谷胱甘肽(GSH)、丙二醛(MDA)、超氧化物歧化酶(SOD)、一氧化氮(NO)的水平进行检测。采用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法(UPLC-Q-Orbitrap-MS)对各组沙鼠进行尿液代谢组学分析,并采用多元统计分析对数据进行处理,以第一主成分的变量重要性投影(VIP)值>1且t检验P<0.05筛选差异代谢物。运用京都基因与基因组百科全书(KEGG)数据库结合MetaboAnalyst 5.0软件对筛选出的差异代谢物进行代谢通路分析。结果:与空白组比较,模型组神经功能缺损评分明显升高(P<0.05),与模型组比较,关白附和禹白附组在给药7 d和14 d后神经功能缺损评分明显降低(P<0.05);与空白组比较,模型组脑梗死率明显升高(P<0.05),与模型组比较,关白附和禹白附组脑梗死率明显降低(P<0.05);与空白组比较,模型组沙鼠血清和脑组织中IL-6、TNF-α、MDA水平明显升高(P<0.05),SOD、GSH、NO水平明显降低(P<0.05),与模型组比较,关白附和禹白附组均能不同程度下调IL-6、TNF-α、MDA水平并上调SOD、GSH、NO水平。尿液代谢组学共筛选出112个内源性差异代谢物,其中关白附和禹白附分别回调了其中16个和26个代谢物,可分别Objective:To investigate the effects of Aconiti Coreani Radix and Typhonii Rhizoma on the urinary metabolites of gerbils with stroke by non-targeted metabolomics technique,and then to clarify the mechanism of the two,as well as their similarities and differences.Method:Twenty-four gerbils were randomly divided into control group(CG),model group(MG),Aconiti Coreani Radix group(RA)and Typhonii Rhizoma group(RT).Except for the CG,ischemic stroke model was constructed using right unilateral ligation of gerbil carotid artery in the remaining groups.Except for the CG and MG,rats in the other groups received whole powder suspension(0.586 mg·g^(-1))was administered for 14 days.The neurological deficit in each group was scored by Longa scoring on days 0,3,7 and 14.After the end of administration,the serum,brain tissue and urine of gerbils in each group were collected,and the rate of cerebral infarction was detected by 2,3,5-triphenyltetrazolium chloride(TTC),and the levels of interleukin(IL)-6,tumor necrosis factor(TNF)-α,malondialdehyde(MDA),superoxide dismutase(SOD),glutathione(GSH),and nitric oxide(NO)in serum and brain tissue were determined by enzyme-linked immunosorbent assay(ELISA).The urine metabolomics of gerbils in each group was studied by ultra performance liquid chromatography-quadrupole-electrostatic field orbitrap high resolution mass spectrometry(UPLC-Q-Orbitrap-MS),and the data were processed by multivariate statistical analysis,and differential metabolites were screened based on value of variable importance in the projection(VIP)of the first principal component>1 and t-test P<0.05.Metabolic pathway analysis of the screened differential metabolites was performed using Kyoto Encyclopedia of Genes and Genomes(KEGG)database and Metaboanalyst 5.0.Result:Compared with the CG,the neurological deficit score was significantly increased in the MG(P<0.05),compared with the MG,the neurological deficit scores in the RA and RT were significantly reduced after 7 d and 14 d(P<0.05).Compared with the CG,the rate of cer

关 键 词：关白附 禹白附 代谢组学 缺血性脑卒中 多元统计分析 生物标志物 超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法(UPLC-Q-Orbitrap-MS) 

分 类 号：R22[医药卫生—中医基础理论] R28[医药卫生—中医学] R969.1[理学—分析化学] O657[理学—化学]