代谢综合征肾损害患者尿液外泌体微小RNA差异表达谱的生物信息学分析  

Bioinformatics analysis of differentially expressed miRNA in urine exosomes of metabolic syndrome patients with renal damage

作　　者：张勇[1,2] 吴庭坤 李舒静 陈小青 谢扬彬[1] 郭秀丽 赖素仁 杨玲婷 庄永泽 王丽萍 ZHANG Yong;WU Tingkun;LI Shujing;CHEN Xiaoqing;XIE Yangbin;GUO Xiuli;LAI Suren;YANG Linting;ZHUANG Yongze;WANG Liping(Fuzong Clinical Medical College of Fujian Medical University,Fuzhou,Fujian 350025,China;Division of Nephrology,The 900th Hospital of Joint Logistic Support Force,PLA)

机构地区：[1]福建医科大学福总临床医学院,福建福州350025 [2]联勤保障部队第九〇〇医院肾脏病科

出　　处：《中华高血压杂志》2023年第8期751-758,共8页Chinese Journal of Hypertension

基　　金：国家自然科学基金面上项目(81373837);福建省自然科学基金面上项目(2018J01184,2019J01526,2021J011277);福建省免疫性慢性肾病临床医学研究中心(2021Y016)。

摘　　要：目的 探讨代谢综合征(MS)肾损害患者尿液外泌体中的微小RNA(miRNA)差异表达谱,探究miRNA在MS肾损害发病中的作用。方法 选取2018年5月至2019年12月在联勤保障部队第九〇〇医院确诊为MS肾损害和MS无肾损害的患者,以及健康体检者。利用实时定量PCR技术及高通量测序技术检测MS肾损害患者(MSK组,n=3)及健康体检者(对照组,n=3)尿液外泌体miRNA的表达情况。对差异表达miRNA进行生物信息学分析,探索靶基因的主要生物学功能。对差异表达的miRNA及其靶基因进行临床验证(MS肾损害患者、MS无肾损害患者、健康受试者各14例)。结果 经过高通量测序技术分析,MSK组和对照组之间存在33个差异表达的尿液外泌体miRNA(P<0.05)。经过小样本量的实时定量PCR检测,提取到11个差异表达的miRNA(P<0.05),7个高表达(miRNA-449a, miRNA-449b-5p, miRNA-449c-5p, miRNA-122-3p, miRNA-618,miRNA-217-5p, miRNA-2115-3p),4个低表达(miRNA-4662a-5p, miRNA-1-3p, miRNA-3114-3p, miRNA-95-3p)。最终选取差异有统计学意义且稳定表达的miRNA-449a进行临床验证。京都基因和基因组百科全书(KEGG)富集分析发现,miRNA-449a靶基因主要参与丝裂原活化蛋白激酶(MAPK)信号通路、转化生长因子β(TGF-β)信号通路、Wnt信号通路等信号转导途径。尿液外泌体miRNA-449a在MSK组、MS无肾损害患者组(MS组)和对照组三组间比较,差异存在统计学意义(0.70±0.32比0.42±0.28比0.06±0.06,F=26.062,P<0.001),MSK组的尿液外泌体miRNA-449a表达水平明显上调。尿液外泌体miRNA-449a表达与血肌酐、尿微量白蛋白与肌酐比值呈正相关(P<0.05)。MSK组尿液外泌体miRNA-449a下游靶基因Bcl-2 mRNA相对表达较MS组下调(0.02±0.01比0.06±0.01,t=102.756,P<0.001)。结论 MS肾损害患者尿液外泌体miRNA存在差异性表达。生物信息学分析提示差异表达Bcl-2等基因可能参与MS肾损害的发生和发展。Objective To detect differentially expressed microRNA(miRNA)in urine exosomes of metabolic syn-drome(MS)patients with renal damage,and to explore the role of miRNAs in the pathogenesis of renal damage in MS.Methods The patients who were diagnosed with MS renal damage and MS without renal damage at the 900th Hospital of Joint Logistic Support Force,PLA from May 2018to December 2019,as well as the health examinations were selected.Real-time quantitative polymerase chain reaction(PCR)and high-throughput sequencing were used to detect the expression of urinary exosomal miRNAs in MS renal damage patients(MS renal damage group,n=3)and health examinations(control group,n=3).Bioinformatics analysis of differentially expressed miRNAs was per-formed to explore the main biological functions of the target genes.The differentially expressed miRNAs and their target genes were verified in 14MS patients with renal damage,14MS patients without renal damage,and 14health examinations.Results High-throughput sequencing analysis showed that 33differentially expressed miRNAs in uinary exosome were present between the MS renal damage group and the control group(P<0.05).Real-time quan-titative PCR with a small sample size showed that there were 11differentially expressed miRNAs(P<0.05),among which 7were up-regulated(miRNA-449a,miRNA-449b-5p,miRNA-449c-5p,miRNA-122-3p,miRNA-618,miRNA-217-5p,miRNA-2115-3p),4were down-regulated(miRNA-4662a-5p,miRNA-1-3p,miRNA-3114-3p,miRNA-95-3p).Then miRNA-449awas selected for further clinical validation due to its significant difference and stable ex-pression(P<0.05).Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis showed that target genes of differentially expressed miRNA may participate in the progression of MS renal damage through mito-gen-activated protein kinase(MAPK)signal pathway,transforming growth factorβ(TGF-β)signal pathway,Wnt signal pathway and other pathway.MiRNA-449ain urine exosomes was significantly different among MS renal dam-age group,MS without renal damage

关 键 词：代谢综合征 肾损害 尿液外泌体 小分子核糖核酸 生物信息学分析 

分 类 号：R692[医药卫生—泌尿科学] R589[医药卫生—外科学]