Integrating proteomics and targeted metabolomics to reveal the material basis of liver-gallbladder damp-heat syndrome in chronic hepatitis B  

联合蛋白质组学和靶向代谢组学揭示慢性乙型肝炎中肝胆湿热证的物质基础

作　　者：LI Ni’ao GONG Yuefeng WANG Jia CHEN Qingqing SU Shibing ZHANG Hua LU Yiyu 李妮奥;龚跃峰;王佳;陈清清;苏式兵;张华;陆奕宇(上海中医药大学交叉科学研究院,上海201203;启东市中医院肝胆泌尿外科,江苏启东226200;上海中医药大学附属曙光医院肝病研究所肝肾疾病教育部重点实验室,上海201203)

机构地区：[1]Institute of Interdisciplinary Integrative Medicine Research,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China [2]Hepatobiliary and Urological Surgery,Qidong Hospital of Traditional Chinese Medicine,Qidong,Jiangsu 226200,China [3]Key Laboratory of Liver and Kidney Diseases(Ministry of Education),Institute of Liver Diseases,Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China

出　　处：《Digital Chinese Medicine》2024年第4期320-331,共12页数字中医药(英文)

基　　金：National Natural Science Foundation of China(82274183);Shanghai Municipal Health Commission’s special clinical research project in the health industry(202240243);Science and Technology Innovation Program of Science and Technology Commission of Shanghai Municipality(STCSM)(20ZR1453700).

摘　　要：Objective To elucidate the biological basis of liver-gallbladder damp-heat syndrome(LGDHS)within the framework of traditional Chinese medicine(TCM)as a complementary diagnostic and therapeutic approach in chronic hepatitis B(CHB).Methods CHB patients and healthy volunteers were enrolled from Shuguang Hospital Affili-ated to Shanghai University of Traditional Chinese Medicine between August 21,2018 and December 31,2020.They were divided into three groups:healthy group,LGDHS group,and latent syndrome(LP)group.Proteomic analysis using isobaric tags for relative and absolute quantitation(iTRAQ)was performed to identify differentially expressed proteins(DEPs).Metabolomic profiling via ultra-performance liquid chromatography-tandem mass spec-trometry(UPLC-MS/MS)was applied to serum samples to detect differentially regulated metabolites(DMs).Kyoto Encyclopedia of Genes and Genomes(KEGG)and Gene Ontology(GO)enrichment were employed to explore dysregulated pathways.Principal component analysis(PCA)and orthogonal partial least squares discriminant analysis(OPLS-DA)were utilized to visualize group separation and identify key metabolites and proteins contributing to LGDHS differentiation.Receiver operating characteristic(ROC)curve analysis evaluated the diagnostic performance of key biomarkers,while logistic regression models assessed their predictive accuracy.P values were corrected for multiple tests using the Benjamini-Hochberg method to control the false discovery rate(FDR).Validation of potential biomarkers was con-ducted using independent microarray data and real-time quantitative polymerase chain reac-tion(RT-qPCR).Results A total of 150 participants were enrolled,including healthy group(n=45),LGDHS group(n=60),and LP group(n=45).254 DEPs from proteomics data and 72 DMs from metabolomic profiling were identified by PCA and OPLS-DA.DEPs were mainly enriched in immune and complement pathways,while DMs involved in amino acid and energy metabolism.The integrated analysis identified seven key biomarkers:α1-acid glycoprote目的阐明肝胆湿热证(LGDHS)在中医框架内作为慢性乙型肝炎(CHB)辅助诊疗手段的生物学基础。方法本研究于2018年8月21日至2020年12月31日期间从上海中医药大学附属曙光医院招募CHB患者和健康志愿者,并将他们分为三组:健康组、LGDHS组和潜在症状(LP)组。使用基于同位素标记相对和绝对定量(iTRAQ)的蛋白质组学分析,以识别差异表达蛋白(DEPs)。通过超高效液相色谱串联质谱(UPLC-MS/MS)对血清样本进行代谢组学分析,鉴定差异代谢物(DMs)。采用京都基因与基因组百科全书(KEGG)和基因本体论(GO)富集分析,探讨LGDHS失调的生物通路。使用主成分分析(PCA)和正交偏最小二乘判别分析(OPLS-DA)进行组间分离和关键代谢物及蛋白的筛选。通过受试者工作特征(ROC)曲线分析评估关键生物标志物的诊断性能,使用逻辑回归模型评估其预测准确性。多重检验采用Benjamini-Hochberg方法对多重测试进行P值校正以控制伪发现率(FDR)。此外,利用独立的微阵列数据和实时定量聚合酶链反应(RT-qPCR)验证潜在生物标志物。结果研究共招募了150名参与者,包括健康组(n=45)、LGDHS组(n=60)和LP组(n=45)。通过PCA和OPLS-DA从蛋白质组学中共鉴定254个DEPs,代谢组学分析中发现72个DMs。DEPs主要富集于免疫和补体通路,DMs涉及氨基酸和能量代谢通路。综合分析筛选出7个关键生物标志物,包括类粘蛋白1(ORM1)、天冬酰胺合成酶(ASNS)、溶质载体家族27成员5(SLC27A5)、葡萄糖苷酶IIα亚基(GANAB)、己糖激酶2(HK2)、5-甲基四氢叶酸-同型半胱氨酸甲基转移酶(MTR)和麦芽糖酶-葡糖淀粉酶(MGAM)。微阵列验证显示这些基因在ROC曲线分析中的曲线下面积(AUC)值范围为0.536至0.759。其中,ORM1、ASNS和SLC27A5在区分LGDHS患者方面表现出显著的差异能力(分别为P=0.016、P=0.035、P<0.001),对应的AUC值分别为0.749、0.743、0.759。逻辑回归模型结合这三个基因�

关 键 词：Liver-gallbladder damp-heat syndrome(LGDHS) Chronic hepatitis B(CHB) PROTEOMICS Targeted metabolomics Molecular mechanism Biomarkers 

分 类 号：R73[医药卫生—肿瘤]