基于液质联用代谢组学技术探索肝内胆汁淤积症的潜在血清代谢标志物

Exploring potential serum metabolite markers of intrahepatic cholestasis based on liquid chromatography-mass spectrometry metabolomics technology

目的通过液质联用代谢组学技术分析肝内胆汁淤积症(IHC)患者的血液差异代谢物,寻找潜在代谢靶点。方法采集30例IHC患者(IHC组)和30名健康个体(健康组)血清样本。进行代谢组学技术分析后,基于差异倍数和显著性进行初筛,对差异代谢产物进行KEGG富集分析,确定候选靶点,利用受试者操作特征曲线分析这些特征代谢产物的临床潜在应用价值。结果IHC组和健康组年龄差异无统计学意义(P>0.05),2组肝生物化学指标、血常规、凝血以及炎症指标差异均有统计学意义(P值均<0.05),与临床病情一致。进一步经血液代谢组学检测分析,筛选出与IHC相关的差异代谢产物99种,其中15种差异有统计学意义。差异代谢产物主要涉及多种葡萄糖、脂质和能量代谢。受试者操作特征曲线下面积值>0.9的包括1H-吲哚-3-甲醛、6-羟基-1H-吲哚-3-乙酰胺、苯丙氨酰色氨酸、1-甲基鸟酐、2-乙氧基-5-甲基吡嗪、对羟基苯甲醛、5-(2-氯苯基)-3,4-二氢-2H-吡咯、甲硫腺苷、丙氨酰异亮氨酸、安艾苦素、N-乙酰基-DL-组氨酸一水合物、N-甲基烟酰胺等12种代谢产物。根据差异代谢物的定量值计算对应的比值联合受试者操作特征曲线对潜在生物标志物的鉴别能力进行深入评价分析后,得出变化上调及下调倍数前15个代谢物且受试者操作特征曲线下面积值>0.9的代谢物包括苯丙氨酰色氨酸、苯丙氨酰丙氨酸、5-甲硫腺苷、安艾苦素、N-甲基烟酰胺。结论苯丙氨酰色氨酸、苯丙氨酰丙氨酸、5'-甲硫腺苷、安艾苦素、N-甲基烟酰胺可能作为区分健康对照人群与IHC患者的潜在代谢标志物,其中N-甲基烟酰胺作为潜在标志物的意义较大。

Objective To analyze the blood differential metabolites of patients with intrahepatic cholestasis(IHC)by liquid chromatography-mass spectrometry metabolomics technology so as to find potential metabolic target.Method Serum samples were collected from thirty patients with intrahepatic cholestasis and thirty healthy individuals after metabolomics analysis.The differential metabolites were initially screened based on the multiple differences and significance.KEGG enrichment analysis was performed on the differential metabolites to determine the candidate targets.The potential clinical application value of these characteristic metabolites was analyzed using the receiver operating characteristic curve.Result A total of thirty patients with intrahepatic cholestasis and thirty healthy adults were included.The age difference between the two groups was not statistically significant(P>0.05).The clinical condition was consistent with the statistically significant differences in liver biochemical indicators,blood routine,coagulation,and inflammatory indicators between the two groups(P<0.05).Furthermore,a blood metabolomics screening analysis revealed 99 differentially expressed metabolites associated with intrahepatic cholestasis.Of these,15 showed statistically significant differences.Glucose,lipid,and energy metabolisms were the various primary types of differential metabolites involved.The receiver operating characteristic curve>0.9 included the following twelve kinds of metabolites:1H-indole-3-carboxaldehyde,6-hydroxy-1H-indole-3-acetamide,phenylalanyl tryptophan,1-methylguanosine,2-ethoxy-5-methylpyrazine,p-hydroxybenzaldehyde,5-(2-chlorophenyl)-3,4-dihydro-2H-pyrrole,methylthioadenosine,alanylisoleucine,anabsinthin,N-acetyl-DL-histidine monohydrate,N-methylnicotinamide,and others.The fifteen metabolites that were previously identified and calculated according to the differential quantitative value of the metabolite corresponding ratio exhibited fold-changes in the upregulated and downregulated potential biomarkers(phenylalanine tryptophan,phenylalanine,5'-methylthioadenosine,anabsinthin,and N-methylnicotinamide)in combination with the area under the receiver operating characteristic curve>0.9.Conclusion Phenylalanyl tryptophan,phenylalanylalanine,5'-methylthioadenosine,anabsinthin,and N-methylnicotinamide may serve as potential metabolic markers to distinguish patients with cholestasis from healthy controls.N-methylnicotinamide,among them,is of great importance as a potential marker.