摘要
目的采用代谢组学分析芫花酯甲抑制人肝细胞L02增殖的差异代谢物,并揭示其毒性机制。方法芫花酯甲0,2.5,5和10μmo.lL-1加入L02细胞孵育48 h后,采用超高效液相色谱串联四级杆飞行时间质谱(UPLC-QTOF/MS)分析细胞代谢物;运用偏最小方差判别分析模型区别对照组和芫花酯甲2.5,5和10μmo.l L-1作用L02后细胞代谢物的总体差异;计算各组细胞代谢物归一化后的峰面积与细胞毒性强度的相关系数,从而发现并鉴定芫花酯甲损伤L02细胞的差异标志物;采用DAVID数据库分析差异代谢标志物相关基因的信号通路。结果正常对照组及芫花酯甲2.5,5和10μmo.lL-1组间的L02细胞代谢物存在明显差异,共鉴定出5个细胞代谢物峰面积显著下调,与细胞毒性强度的相关系数为-0.53～-0.96;5个代谢物峰面积显著上调,与细胞毒性强度的相关系数为0.72～0.99。代谢物包括磷脂类和脂肪酸类等,主要涉及甘油磷脂代谢、鞘脂代谢、脂肪酸氧化、花生四烯酸代谢和有丝分裂原激活蛋白激酶细胞凋亡等相关信号通路。结论代谢组学方法能够鉴定芫花酯甲诱导人肝细胞L02损伤的差异标志物及其信号通路。
OBJECTIVE To analyze metabolites related to the damage to human L02 liver cells caused by yuanhuacine using metabonomics and to reveal the mechanism of yuanhuacine toxicity.METHODS L02 cells were cultured with yuanhuancine 0,2.5,5 and 10 μmol·L-1 for 48 h,and the cellular metabolites were analyzed by UPLC-QTOF/MS.The partial least squares-discriminant analysis（PLS-DA） model was used to distinguish normal control group from yuanhuacine 2.5,5 and 10 μmol·L-1 groups.Moreover,the correlation coefficient of normalized peak areas and cytotoxic intensity of yuanhuacine was calculated,and different markers of yuanhuacine which do harm to L02 cells were identified.Different biomarkers associated with gene signaling pathways were analyzed by DAVID database.RESULTS Significant difference in metabolite markers between normal control and yuanhuacine treated groups was observed.Normalized peak areas of five metabolites were significantly decreased,and the correlation coefficient was-0.53——0.96.Five metabolite normalized peak areas significantly increased,and the correlation coefficient was 0.72-0.99.These ten markers were phospholipids,fatty acids and the other intermediate products,mainly involved in glycerophospholipid metabolism,sphingolipid metabolism,fatty acid oxidation,arachidonic acid metabolism,and MAPK cell apoptosis signaling pathways.CONCLUSION This metabonomics approach is not only able to identify different markers and signaling pathways of yuanhuacine that induce damage to L02 liver cells,but helps reveal the mechanism of yuanhuacine toxicity.
出处
《中国药理学与毒理学杂志》
CAS
CSCD
北大核心
2013年第4期704-709,共6页
Chinese Journal of Pharmacology and Toxicology
基金
国家重点基础研究发展计划(973计划)(2011CB505300),国家重点基础研究发展计划(973计划)(2011CB505303)
国家自然科学基金(81274199)
江苏高校优势学科建设工程项目~~
关键词
芫花酯甲
肝细胞
毒性作用
串联质谱法
yuanhuacine
hepatocytes
toxic actions
tandem mass spectrometry
