新生与成年大鼠心脏成纤维细胞来源外泌体蛋白质组学差异分析

Differential proteomic analysis of cardiac fibroblast-derived exosomes in neonatal and adult rats

目的幼体心脏具有再生修复功能,通过分析新生大鼠与成年大鼠心脏成纤维细胞来源外泌体的差异蛋白,从分子水平上探讨心肌受损后可能的治疗靶点。方法分别从新生和成年大鼠心脏成纤维细胞中分离出外泌体,并提取蛋白质进行高效液相色谱法(high per-formance liquid chromatography,HPLC)和液相色谱串联质谱法(liquid chromatography coupled to tandem mass spectrometry,LC-MS/MS)分析,鉴定差异表达的蛋白质。使用DAVID数据库对差异表达蛋白基因进行基因本体论(Gene Ontology,GO)和京都基因与基因组百科全书(kyoto encyclopedia of genes and genomes,KEGG)通路分析。并进一步使用STRING数据库和Cytoscape软件进行蛋白质相互作用网络(protein-protein interaction,PPI)分析,以筛选出关键基因。结果在新生大鼠中共发现241个差异表达蛋白,其中上调表达的蛋白有117个,下调表达的蛋白有124个。GO分析显示上调表达基因主要涉及整合素介导的信号通路和对缺氧的反应,而对内肽酶活性负调控的基因下调表达。KEGG分析结果表明上调表达基因主要富集于Hippo信号通路,而下调表达基因主要涉及细胞外基质与受体相互作用。通过PPI分析得出有6个关键基因(Lamc1、Agrn、Itih3、Tfrc、Bgn和Atp5pd)下调,有1个关键基因(Rp17a)上调。结论通过蛋白组学和生物信息学方法,筛选出了新生大鼠和成年大鼠心脏成纤维细胞来源外泌体的差异蛋白,以及相关的信号通路和关键基因,为心肌受损精准治疗提供了潜在的靶点。

Objective This study investigated the regenerative repair function of the juvenile heart by analyzing differential pro-teins found in cardiac fibroblast-derived exosomes in neonatal rats and adult rats.The aim of this research was to identify possible thera-peutic targets at the molecular level following myocardial damage.Methods We isolated exosomes from neonatal and adult rat cardiac fibroblasts and extracted proteins for analysis using HPLC and LC-MS/MS.We identified differentially expressed proteins and further analyzed their gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways using the DAVID database.To identify key genes,we conducted a protein interaction network(PPI)analysis using the STRING database and Cytoscape software.Re-sults In neonatal rats,a total of 241 differentially expressed proteins were identified,with 117 up-regulated and 124 down-regulated.The GO analysis showed that the up-regulated genes were mostly involved in integrin-mediated signalling pathways and responses to hypoxia.On the other hand,the down-regulated genes were involved in negative regulation of endopeptidase activity.The KEGG analy-sis indicated that the up-regulated genes were mainly enriched in the Hippo signaling pathway,while the down-regulated genes were primarily involved in ECM-receptor interaction.PPI analysis revealed that one key gene(Rp17a)was up-regulated,while six key genes(Lamc1,Agrn,Itih3,Tfrc,Bgn and Atp5pd)were down-regulated.Conclusions In this study,proteomics and bioinformatics techniques were employed to identify differential proteins in exosomes derived from cardiac fibroblasts of neonatal and adult rats.The study also investigated the associated signaling pathways and key genes,which could serve as potential targets for precise treatment of myocardial damage.