脓毒症外源性ARDS关键基因及信号通路的转录组学分析  

Transcriptomics analysis of key genes and signaling pathways in sepsis-related exogenous acute respiratory distress syndrome

作　　者：谢永鹏[1] 骆继业[1] 王言理[1] 胡文霞[1] 芦雨 张倩 李小民[1] Xie Yongpeng;Luo Jiye;Wang Yanli;Hu Wenxia;Lu Yu;Zhang Qian;Li Xiaomin(Department of Emergency and Critical Care Medicine,the First Pepple's Hospital of Lianyungang,the Institute of Emergency Medicine of Lianyungang,Lianyungang 222000,Jiangsu,China)

机构地区：[1]连云港市第一人民医院急危重症医学部,连云港市急诊医学研究所,江苏连云港222000

出　　处：《中华危重病急救医学》2022年第11期1154-1160,共7页Chinese Critical Care Medicine

基　　金：江苏省科技厅社会发展面上项目(BE2020670);江苏省卫生健康委科研项目(H2019109);江苏省连云港市卫生健康委青年科研项目(QN1804)。

摘　　要：目的分析脓毒症外源性急性呼吸窘迫综合征(ARDS)大鼠的差异表达基因(DEG),从转录组水平探讨脓毒症ARDS的早期诊断及防护机制。方法将12只6~8周龄雄性SD大鼠按随机数字表法分为脂多糖(LPS)致脓毒症ARDS模型组(模型组,腹腔注射LPS 15 mg/kg)与对照组(腹腔注射等量生理盐水),每组6只。分别提取两组大鼠左肺组织RNA,采用illumina Hiseq测序平台的双端测序模式进行高通量测序。采用DESeq2软件以|log_(2)差异倍数(FC)|≥3且P<0.001筛选DEG。对DEG进行基因本体(GO)功能富集分析和京都基因与基因组百科全书数据库(KEGG)通路富集分析。使用STRING在线数据库和CytoScape软件构建蛋白质-蛋白质相互作用(PPI)网络并筛选关键基因。分离并提取2021年3月至11月连云港市第一人民医院急危重症医学部收治的20例脓毒症患者及20例年龄匹配的同期健康体检者的外周血单个核细胞(PBMC),采用实时荧光定量聚合酶链反应(RT-qPCR)对关键基因进行验证。结果共筛选出DEG 286个,其中上调基因202个,下调基因84个。GO富集分析表明,DEG主要参与了中性粒细胞趋化迁移、抗菌体液反应、宿主免疫应答及体液免疫反应等生物学过程;KEGG富集分析显示,DEG主要通过参与白细胞介素-17(IL-17)信号通路、肿瘤坏死因子(TNF)信号通路及趋化因子信号通路发挥生物学作用。PPI分析共筛选出节点蛋白262个,相互作用关系852条边;筛选出前15位关键基因,分别为IL-6、TNF、IL-10、IL-1β、CXC趋化因子配体1(CXCL1)、CXCL10、CXC趋化因子受体3(CXCR3)、CXCR2、CXCL9、CC趋化因子配体7(CCL7)、CXCL11、CCL1、CXCL13、CCL12、CCL22。采用RT-qPCR对脓毒症ARDS患者与健康对照者PBMC进行差异基因测序验证,结果显示,脓毒症患者5个代表性关键基因表达明显高于健康对照者〔IL-6 mRNA(2^(-ΔΔCt)):2.803±1.081比0.951±0.359,TNF mRNA(2^(-ΔΔCt)):2.376±0.799比1.150±0.504,CXCL10 mRNA(2^(-ΔObjective To analyze the differentially expressed gene(DEG)in rats with sepsis-induced exogenous acute respiratory distress syndrome(ARDS)and explore the early diagnosis and protective mechanism of sepsis-induced ARDS at the transcriptome level.Methods Twelve 6 to 8 weeks old male Sprague-Dawley(SD)rats were randomly divided into lipopolysaccharide(LPS)induced sepsis-induced ARDS model group(model group,intraperitoneal injection of LPS 15 mg/kg)and control group(intraperitoneal injection of the same volume of normal saline),with 6 rats in each group.RNA was extracted from the left lung tissue of the two groups,and the paired-end sequencing mode of the illumina Hiseq sequencing platform was used for high-throughput sequencing.The DESeq2 software was used to screen DEG with|log_(2)(fold change,FC)|≥3 and P<0.001.Gene ontology(GO)function enrichment analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis were performed on DEG.STRING and CytoScape software were used to construct a protein-protein interaction(PPI)network and screen key genes.The peripheral blood mononuclear cell(PBMC)of 20 septic patients admitted to the emergency and critical care medical department of Lianyungang First People's Hospital from March to November 2021 and 20 age-matched healthy people in the same period were isolated and extracted,and the key genes were verified by real-time fluorescent quantitative polymerase chain reaction(RT-qPCR).Results A total of 286 DEG were screened,including 202 up-regulated genes and 84 down-regulated genes.GO enrichment analysis showed that DEG was mainly involved in biological processes such as neutrophil chemotaxis migration,antibacterial humoral response,host immune response,and humoral immune response.KEGG analysis showed that DEG mainly played a biological role through interleukin-17(IL-17)signaling pathway,tumor necrosis factor(TNF)signaling pathway,and chemokine signaling pathway.In PPI analysis,a total of 262 node proteins were screened,and the interaction relationship

关 键 词：脓毒症 急性呼吸窘迫综合征 转录组学 生物信息学 关键基因 

分 类 号：R459.7[医药卫生—急诊医学]