基于非靶向代谢组学的肝阴虚小鼠模型评价  

Evaluation of A Liver Yin Deficiency Mouse Model Based on Untargeted Metabolomics

作　　者：黄雪柠 赵夏 候梦雨 过至雨 叶耀辉 HUANG Xuening;ZHAO Xia;HOU Mengyu;GUO Zhiyu;YE Yaohui(Jiangxi University of Chinese Medicine,Nanchang 330004,China;Nanchang Medical College,Nanchang 330052,China)

机构地区：[1]江西中医药大学,南昌330004 [2]南昌医学院,南昌330052

出　　处：《中国实验方剂学杂志》2024年第5期118-125,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金地区科学基金项目(H2805-82060717)。

摘　　要：目的:基于超高效液相色谱-四极杆-飞行时间串联质谱法(UPLC-Q-TOF-MS),从非靶向代谢组学的角度评价甲状腺片混悬液联合10%四氯化碳(CCl_(4))建立小鼠肝阴虚模型的情况,为中医证候模型的建立奠定基础。方法:将24只雄性ICR小鼠随机平均分为空白组、模型组,模型组连续14 d灌胃甲状腺片混悬液(0.0032 g·kg^(-1))且每周1次腹腔注射10%CCl_(4)(5 mL·kg^(-1))建立肝阴虚模型,空白组仅腹腔注射等量橄榄油并灌胃等量蒸馏水,实验期间喂食正常饲料,造模完成后每组随机选取6只小鼠生化检测血清中丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、环磷酸腺苷(cAMP)、环磷酸鸟苷(cGMP)、白细胞介素(IL)-6、IL-10、肿瘤坏死因子-α(TNF-α)及肝脏中丙二醛(MDA)、超氧化物歧化酶(SOD)、总蛋白(TP)、羟脯氨酸(HYP)等指标的水平。取肝组织切片进行苏木素-伊红(HE)染色并观察病理变化。每组剩余6只小鼠采用UPLC-Q-TOF-MS结合主成分分析(PCA)和正交偏最小二乘法-判别分析(OPLS-DA)筛选肝阴虚小鼠模型的差异代谢物,利用京都基因与基因组百科全书(KEGG)数据库分析差异代谢物的相应代谢通路。结果:与空白组比较,模型组小鼠出现体质量减轻,疲倦易困,毛发杂乱无光泽,尿液增多,易怒等肝阴虚表现;生化指标ALT、cAMP/cGMP、IL-6、AST、MDA、cAMP、TNF-α明显升高(P<0.05,P<0.01),SOD、IL-10、cGMP明显降低(P<0.05,P<0.01),HYP、TP变化差异无统计学意义;模型组切片图可见肝脏脂肪变性,肝板细胞的放射状排列出现扭曲;内源性物质分离明显,血清样品中鉴别出252个差异代谢物,主要涉及嘌呤代谢、类固醇激素生物合成、嘧啶代谢等代谢途径;肝脏样品中鉴别出229个差异代谢物,主要涉及核苷酸代谢、嘌呤代谢、类固醇激素生物合成、嘧啶代谢、抗叶酸耐药、胰岛素抵抗、初级胆汁酸生物合成、前列腺癌症、硫中继系统、�Objective:Based on ultra performance liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry(UPLC-Q-TOF-MS),to evaluate the establishment of a mouse model of liver Yin deficiency by thyroid tablet suspension combined with 10%carbon tetrachloride(CCl_(4))from the perspective of non-targeted metabolomics,in order to lay the foundation for the establishment of a traditional Chinese medicine(TCM)syndrome model.Method:A total of 24 mice were randomly divided into blank group and model group.The model group was given thyroid tablet suspension(0.0032 g·kg^(-1))by gavage for 14 consecutive days,and 10%CCl(45 mL·kg^(-1))was intraperitoneally injected once a week to establish a liver Yin deficiency model,while the blank group was injected with an equal amount of olive oil intraperitoneally and gavaged with an equal amount of distilled water,and was fed with normal feed.After the modeling was completed,6 mice in each group were randomly selected,the levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),cyclic adenosine monophosphate(cAMP),cyclic guanosine monophosphate(cGMP),interleukin(IL)-6,IL-10,tumor necrosis factor-α(TNF-α)were measured in the mice serum,and malondialdehyde(MDA),superoxide dismutase(SOD),total protein(TP),hydroxyproline(HYP)and other indicators were measured in the mice liver.Liver tissue sections were taken for hematoxylin-eosin(HE)staining and observing pathological changes.The remaining 6 mice in each group were subjected to UPLC-Q-TOF-MS combined with principal component analysis(PCA)and orthogonal partial least squares-discriminant analysis(OPLS-DA)were used to screen differential metabolites in the liver Yin deficiency mouse model,Kyoto Encyclopedia of Genes and Genomes(KEGG)database was used to analyze the corresponding metabolic pathways of differential metabolites.Result:Compared with the blank group,mice in the model group showed liver Yin deficiency manifestations such as reduced body weight,fatigue and sleepiness,disheveled and lusterless hair,irritabi

关 键 词：肝阴虚 代谢组学 中医证候 模型评价 代谢途径 超高效液相色谱-四极杆-飞行时间串联质谱法(UPLC-Q-TOF-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]