摘要
目的构建膀胱尿路上皮癌(bladder urothelial carcinoma,BUC)尿液差异蛋白数据集,筛选核心差异蛋白并初步验证。方法于2015年1月至2016年11月在天津医科大学第二医院泌尿外科住院病房,采集膀胱上皮癌患者术前尿液标本78例,其中男性55人,女性23人,年龄(68.9±6.6)岁。同时收集51例健康志愿者尿液标本用于对照组分析,其中男性34人,女性17人,年龄(60.6±11.0)岁。采用同位素相对标记和绝对定量技术(isobaric tags for relative and absolute quantitation,iTRAQ)对BUC组混合尿液样本4例和对照组混合尿液样本2例,共6例尿液混合标本进行比较蛋白质组学研究,找出两组间的尿液差异蛋白,选取差异倍数前20的差异蛋白进行质谱多反应监测(multiple reaction monitoring,MRM)技术验证;应用R语言cluster profiler包等进行差异蛋白的京都基因与基因组百科全书(kyoto encyclopedia of genes and genomes,KEGG)富集分析、应用R语言的org.Hs.eg.db包等进行基因本体(Gene ontology,GO)分析,初步选取有意义的差异蛋白,构建BUC尿液差异蛋白数据集。应用R语言绘制蛋白间相互作用(protein protein interaction,PPI)图,筛选核心差异蛋白。用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)验证核心差异蛋白在尿液中的差异存在。结果通过分析得到101个BUC尿液差异蛋白,其中37个蛋白表达上调,64个蛋白表达下调。MRM检测到10个差异蛋白;KEGG分析显示差异蛋白显著富集在其他聚糖降解,补体和凝血级联这两个代谢通路,包含11个差异蛋白;GO分析显示差异蛋白主要参与细胞黏附分子结合、羧酸结合、有机酸结合、糖胺聚糖结合、肽链内切酶激活等生物过程,涉及54个差异蛋白;三者共同构成BUC尿液差异蛋白数据集,包括69个蛋白。通过蛋白互作分析筛选出APOE和APOA4为核心差异蛋白。ELISA结果显示:BUC组和对照组的APOE平均浓度分别为0.55和0.30 pg/mL。BUC组和�
Objective To construct a urinary differential proteins dataset for bladder urothelial carcinoma(BUC)so as to screen the core differentially expressed proteins(DEPs)and then validate them.Methods Preoperative urine samples were collected from 78 hospitalized BUC patients(55 males,23 females;68.9±6.6 years old)in the Urology Department of the Second Hospital of Tianjin Medical University from January 2015 to November 2016.At the same time period,urine samples from 51 healthy volunteers were also collected for control group analysis(34 males,17 females;60.6±11.0 years old).Using isobaric tags for relative and absolute quantitation(iTRAQ)technology,comparative proteomics study was conducted on a total of 6 mixed urine samples,including 4 cases in the BUC group and 2 cases in the control group,to find out the DEPs between the 2 groups.The top 20 DEPs were selected for multiple reaction monitoring(MRM)verification;Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis was carried out with R package,clusterProfiler,and Gene Ontology(GO)enrichment analysis was subsequently performed with org.Hs.eg.db package to preliminarily select the meaningful differential proteins for BUC urinary differential protein dataset.Then the Protein-Protein Interaction(PPI)map was plot using R language to screen the core differential proteins.Furthermore,enzyme-linked immunosorbent assay(ELISA)was applied to verify the difference of core differential proteins in the urine samples.Results A total of 101 BUC urinary DEPs were obtained,of which 37 proteins were up-regulated and 64 down-regulated.MRM detected 10 DEPs.KEGG analysis found that there were 11 DEPs significantly enriched in 2 metabolic pathways,namely other chitosan degradation and complement and blood coagulation cascade.GO analysis displayed that totally 54 DEPs were mainly involved in cell adhesion molecule binding,carboxylic acid binding,organic acid binding,glycosaminoglycan binding,endopeptidase activity and other biological processes.Combining the 3 results,the BUC
作者
王琳瑶
李常颖
李建民
李宏杰
WANG Linyao;LI Changying;LI Jianmin;LI Hongjie(Hebei Provincial Key Laboratory of Chronic Diseases,College of Basic Medical Sciences,North China University of Science and Technology,Tangshan,Hebei Province,063000;Department of Tumor Immunology,Tianjin Institute of Urology,Second Hospital of Tianjin Medical University,Tianjin,300211,China)
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2021年第10期982-988,共7页
Journal of Third Military Medical University
基金
河北省省级科技计划(H2015209164)。
关键词
ITRAQ
MRM
R语言
KEGG富集分析
GO富集分析
蛋白质相互作用
isobaric tags for relative and absolute quantitation
multiple reaction monitoring
R language
Kyoto Encyclopedia of Genes and Genomes enrichment analysis
Gene Ontology enrichment analysis
Protein-Protein interaction
