免疫应激介导的何首乌“九蒸九晒”炮制减毒作用及代谢组学研究  

Metabolomic study on immunological stress-mediated hepatotoxicity of Polygonum multiflorum and its processed products of nine times steaming and nine times sunning

作　　者：李春雨[1,2] 何琴[2,3] 唐进法[2,4,5] 沙孟晨 涂灿[2,5] 张乐[2,5] 刘振兴[2] 王伽伯[2] 肖小河[2] 

机构地区：[1]中国医学科学院肿瘤医院,北京100021 [2]解放军302医院全军中医药研究所,北京100039 [3]湖南中医药大学药学院,湖南长沙410208 [4]河南中医药大学第一附属医院,河南郑州450000 [5]成都中医药大学药学院,四川成都611137

出　　处：《药学学报》2017年第7期1069-1076,共8页Acta Pharmaceutica Sinica

基　　金：国家公益性行业专项课题项目(201507004-04);中国博士后科学基金项目(2016M590065);北京市科技新星计划项目(Z16111000490000)

摘　　要：基于免疫应激模型,研究何首乌经"九蒸九晒"炮制后对特异质肝损伤大鼠的代谢作用机制。将何首乌和制首乌的50%乙醇提取物单独灌胃或联合无毒剂量的脂多糖(LPS)给予SD大鼠,检测血清丙氨酸氨基转氨酶(ALT)、天冬氨酸氨基转氨酶(AST),HE染色观察肝脏病理学改变,对比考察何首乌炮制前后单用或联合LPS的毒性差异。通过UPLC-QTOF/MS分析不同组别大鼠血浆样品的代谢轮廓谱,结合偏最小二乘判别法(OPLS-DA)的分析方法,研究何首乌经"九蒸九晒"炮制前后对特异质肝损伤大鼠血浆中内源性代谢物的影响,寻找何首乌炮制减毒的潜在生物标志物,将鉴定到的生物标志物输入KEGG数据库中构建代谢通路。结果显示,何首乌炮制前后单次灌胃(5.4 g·kg^(-1))对大鼠ALT和AST无显著影响,肝脏切片未见明显病理学改变;而相同剂量的何首乌和制首乌联合LPS给药后,何首乌组ALT和AST均显著升高(P<0.01),肝脏切片可见明显病理学改变,而制首乌组未出现肝损伤。主成分分析(PCA)结果显示,正常对照组、LPS组、LPS/制首乌组和LPS/何首乌组血清代谢物谱得到明显分离,发现并鉴定了10个与肝损伤相关的潜在生物标志物。推测这些生物标志物可能与鞘脂代谢、亚油酸代谢、甾类激素生物合成、半乳糖代谢、类固醇生物合成、细胞色素P450外源性物质代谢、嘧啶代谢、不饱和脂肪酸合成、初级胆汁酸合成,以及牛磺酸与亚牛磺酸代谢等10个代谢通路有关。故何首乌炮制减毒作用机制可能与调节这些代谢途径相关,以期为何首乌炮制减毒提供科学依据。It is investigated that the hepatotoxicity of Polygonum multiflorum（PM） was attenuated by its processed products of nine times steaming and nine times sunning（RPM） based on immunological stress-mediated animal model by using metabolomics method. Sprague-Dawley（SD） rats were intragastrically administered with（5.4 g crude drug per kg body weight） of 50% alcohol extracts of PM and its processed products of nine times steaming and nine times sunning respectively or co-treated with non-toxic dose of lipopolysaccharide（LPS, 2.8 mg·kg^（-1）） via tail vein injection. The plasma alanine aminotransferase（ALT） and aspartate aminotransferase（AST） activities were assayed and the isolated livers were evaluated for histopathological changes. Global metabolomics profiling, multivariate analysis and data base searching were performed to discover common differential metabolites for idiosyncratic liver injury. The results showed that co-treatment with non-toxic dose of LPS and PM could result in significant liver injury, indicated by the elevation of plasma ALT and AST activities, as well as obvious liver histologic damage; whereas RPM failed to induce detectable liver injury. Furthermore, 10 potential metabolomics biomarkers that differentially expressed in LPS/PM group compared with LPS/RPM without liver injury were identified by untargeted metabolomics, mainly involved ten pathways： sphingolipid metabolism, linoleic acid metabolism, taurine and hypotaurine metabolism, steroid hormone biosynthesis, galactose metabolism, steroid biosynthesis, metabolism of xenobiotics by cytochrome P450, pyrimidine metabolism, biosynthesis of unsaturated fatty acids, primary bile acid biosynthesis. This work illustrated the idiosyncratic hepatotoxicity of heshouwu and provided a metabolomic insight into diosyncratic liver injury of PM and RPM.

关 键 词：何首乌 免疫应激 特异质肝损伤 代谢组学 通路分析 

分 类 号：R285[医药卫生—中药学]