基于生物信息学的非酒精性脂肪性肝病关键基因筛选及实验验证  

Screening and experimental validation of key genes in non-alcoholic fatty liver disease basedon bioinformatics

作　　者：王一涵 漆光紫 李文学[5] 岑育芳[1,3,4] 韦俊宏 唐玉航 肖荣清 何志雁 庞雅琴 Wang Yihan;Qi Guangzi;LiWenxue;Cen Yufang;Wei Junhong;Tang Yuhang;Xiao Rongqing;He Zhiyan;Pang Yaqin(School of Basic Medical Sciences,Youjiang Medical University for Nationalities,Baise 533000,China;School of Public Health and Management,Youjiang Medical University for Nationalities,Baise 533000,China;Key Laboratory of Environment and Population Health Research of Guangxi University Eco-Aluminum Industrial Base,Baise 533000,China;Key Laboratory of Environmental Pollution and Health Risk Assessment,Youjiang Medical University for Nationalities,Baise 533000,China;Toxicology and Biochemical Laboratory,Guangzhou Center for Disease Control and Prevention,Guangdong Province,Guangzhou 510000,China;School of Laboratory Medicine,Youjiang Medical University for Nationalities,Baise 533000,China)

机构地区：[1]右江民族医学院基础医学院,百色533000 [2]右江民族医学院公共卫生与管理学院,百色533000 [3]广西高校生态铝工业基地环境与人群健康研究重点实验室,百色533000 [4]右江民族医学院“环境污染与健康风险评价”重点实验室,百色533000 [5]广东省广州市疾病预防控制中心毒理与生化检验科,广州510000 [6]右江民族医学院医学检验学院,百色533000

出　　处：《广西医科大学学报》2023年第7期1092-1100,共9页Journal of Guangxi Medical University

基　　金：国家自然科学基金资助项目(No.81960595,No.81360438,No.81660549);广西自然科学基金资助项目(No.2019JJD140011);右江民族医学院2022年校级研究生创新计划项目(No.YXCXJH2022010)。

摘　　要：目的:探索非酒精性脂肪性肝病(NAFLD)潜在的关键基因和microRNAs(miRNAs)。方法:从美国国立生物技术信息中心(NCBI)公共基因芯片数据平台(GEO)数据库下载两个基因表达谱芯片(GSE96936和GSE62267),利用GEO2R在线工具筛选出NAFLD组与正常对照组的差异表达基因(DEGs),对DEGs进行GO和KEGG信号通路富集分析,进一步应用STRING数据库构建蛋白质—蛋白质相互作用(PPI)网络,用Cytoscape筛选出关键基因。构建NAFLD小鼠模型,通过实时荧光定量PCR(RT-qPCR)验证筛选出的关键基因,利用NetworkAnalyst构建关键基因的靶向miRNAs。结果:总共鉴定出192个DEGs,GO分析显示DEGs生物学功能主要涉及5个KEGG通路,包括类固醇激素生物合成、补体与凝血级联、胆汁分泌、化学致癌、视黄醇代谢相关信号通路,结合PPI网络和CytoHubba的结果,筛选出Tyrobp、Hck、Ctss、Aif1、Fcgr1、Cd68、Cd53、Ly86、Fyb和Alox5ap 10个关键基因,通过RT-qPCR检测,发现与正常肝组织相比,NAFLD肝组织Ly86的mRNA表达下调(P<0.05),NAFLD肝组织Hck、Ctss、Aif1、Fcgr1、Cd68、Cd53、Alox5ap和Fyb的mRNA表达上调(P<0.05),结合DEGs-miRNAs可视化发现4种miRNAs(mmu-miR-155-5p、mmu-miR-122-5p、mmu-miR-124-3p和mmu-miR-1a-3p)与关键基因链接紧密,预测为NAFLD的关键miRNAs。结论:研究结果将有助于确定NAFLD潜在的生物标志物和治疗的新策略。Objective:To explore potential key genes and microRNAs(miRNAs)in non-alcoholic fatty liver disease(NAFLD).Methods:Two gene expression profile chips(GSE96936 and GSE62267)were downloaded from the Gene Expression Omnibus(GEO)database of the National Center for Biotechnology Information(NCBI)in the United States.The differentially expressed genes(DEGs)in the NAFLD group and the control group were screened using GEO2R online tool,the GO and KEGG signaling pathway enrichment analysis was carried out on DEGs,and the protein-protein interaction(PPI)network was further constructed using STRING database.Key genes were screened out using Cytoscape to construct the NAFLD mouse model,the selected key genes were verified by realtime fluorescence quantitative polymerase chain reaction(RT-qPCR)and NetworkAnalyst was used to construct targeted miRNAs of key genes.Results:A total of 192 DEGs were identified.GO analysis showed that the biological functions of DEGs were mainly involved in five KEGG pathways,including signaling pathways related to steroid hormone biosynthesis,complement and coagulation cascade,biliary secretion,chemical carcinogenesis,and retinol metabolism.Combined with the results of PPI and CytoHubba,ten key genes,Tyrobp,Hck,Ctss,Aif1,Fcgr1,Cd68,Cd53,Ly86,Fyb and Alox5ap,were screened out RT-qPCR showed that compared with the normal liver tissues,the mRNA expression of Ly86 in the NAFLD liver tissues was down-regulated(P<0.05),while the mRNA expressions of Hck,Ctss,Aif1,Fcgr1,Cd68,Cd53,Alox5ap and Fyb in the NAFLD liver tissues were upregulated(P<0.05).According to the visualization of DEGs-miRNAs,four miRNAs(mmu-miR-155-5p,mmumiR-122-5p,mmu-miR-124-3p and mmu-miR-1a-3p)were closely linked to key genes,and were predicted to be the key miRNAs of NAFLD.Conclusion:The research findings will contribute to identifying potential biomarkers and new strategies for the treatment of NAFLD.

关 键 词：生物信息学 非酒精性脂肪性肝病 基因 MIRNAS 

分 类 号：R575.5[医药卫生—消化系统]