基于非靶向代谢组学的儿童川崎病血浆生物标志物筛选研究

Plasma biomarker screening in pediatric Kawasaki disease based on non-target metabolomics

目的通过非靶向代谢组学方法探究川崎病(KD)儿童血浆中代谢物的变化。方法采用病例对照研究,选用2021年5月到2022年5月在南通大学附属常州儿童医院风湿免疫科确诊为KD的22例患儿为病例组,并以同期21例健康体检儿童作为对照组,采集入组儿童的血浆样本进行基于HPLC-MS/MS系统四极杆-静电场轨道阱Orbitrap质谱仪的代谢组学检测,并以正交偏最小二乘判别分析(OPLS-DA)进行建模,以模型的VIP值>1和t-test比较中P<0.05作为标准筛选KD儿童血浆样本中的特异性差异代谢物;并选用KEGG数据库对差异代谢物进行注解,用MetaboAnalyst软件对其进行富集分析和拓扑分析,以筛选KD儿童血浆中涉及的关键差异代谢通路。结果代谢组学分析结果表明:①OPLS-DA模型中,KD组和正常组儿童血浆代谢轮廓存在显著差异,组间存在393种差异表达的代谢物,上调的代谢物包括1-甲基腺苷(+85%)、二脂酰甘油基三甲基高丝氨酸(3∶0/20∶4)(+2234%)、嘌呤(+220%)等,下调的代谢物包括溶血磷脂酰胆碱(P-16∶0)(-71%)、异羟脱氧胆酸(-79%)等;②MetaboAnalyst结果显示,鞘脂代谢、咖啡因代谢和酪氨酸代谢通路是KD病理状态中改变最显著的差异代谢通路。结论KD病理状态可造成血浆代谢物谱的显著变化;鞘脂代谢、咖啡因代谢和酪氨酸代谢是改变最显著的差异代谢通路,提示KD会系统性地扰动患儿的生理代谢活动,本文从代谢组学角度较系统地揭示了KD儿童和健康儿童的差异代谢物和代谢通路,为KD的早期诊疗和对应的药物开发提供参考。

Objective To explore the metabolic changes in the plasma of children with Kawasaki disease(KD)with non-targeted metabonomics.Methods The case control study was conducted.Totally 22 children with KD diagnosed in our hospital from May 2021 to May 2022 were enrolled,and 21 healthy children served as the control group.Plasma samples of the children were collected for metabonomics detection based on HPLC-Q-Exactive Orbitrap mass spectrometry.Orthogonal projections to latent structures-discriminant analysis(OPLS-DA)model was used to discriminate the groups.The VIP value of the model>1 and P<0.05 in t-test comparison were used as criteria to screen the specific differential metabolites in plasma samples of KD children;KEGG database was used to annotate the differential metabolites,and MetaboAnalyst software was used to conduct the enrichment and topologic analysis and to screen the key differential metabolic pathways involved in the plasma of KD children.Results The metabonomics showed that:①OPLS-DA model showed significant differences in the plasma metabolic profiles between the KD group and the control group.There were 393 differentially expressed metabolites between groups.The up-regulated metabolites included 1-methyladenosine(+85%),DGTs(3∶0/20∶4)(+2234%),and purine(+220%);and the down-regulated metabolites included lysophosphatidylcholine(P-16∶0)(-71%)and isohydroxydeoxycholic acid(-79%);②MetaboAnalyst showed that sphingolipid metabolism,caffeine metabolism and tyrosine metabolism were the most significant differential metabolic pathways in the pathological condition of KD.Conclusion KD can cause significant changes in the plasma metabolome profile.The most significant differential metabolic pathways include sphingolipid metabolism,caffeine metabolism and tyrosine metabolism,suggesting that KD systematically disturbs the physiological metabolic processes in children.This study reveals the differential metabolites and metabolic pathways between the KD and healthy children by metabolomics,and provides reference for the early diagnosis,treatment and research of novel drugs for KD.