耐药性癫痫患儿外周血microRNAs的表达差异分析  

Differential expression of microRNAs profile in the peripheral blood of children with drug-resistant epilepsy

作　　者：孔惠敏[1] 管刘娟 孙娟[2] 禚志红[1] 王怀立[2] Kong Huimin;Guan Liujuan;Sun Juan;Zhuo Zhihong;Wang Huaili(Department of Pediatrics,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450000,China;Pediatrics Intensive Care Unit,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450000,China)

机构地区：[1]郑州大学第一附属医院小儿内科,郑州450000 [2]郑州大学第一附属医院儿科重症监护室,郑州450000

出　　处：《中华实用儿科临床杂志》2022年第3期186-190,共5页Chinese Journal of Applied Clinical Pediatrics

基　　金：河南省医学科技攻关计划(201701008)。

摘　　要：目的研究耐药性癫痫患儿外周血microRNAs(miRNAs)的表达,为早期识别儿童耐药性癫痫寻找有诊断价值的生物标志物。方法回顾性研究。收集2018年1月至2019年6月在郑州大学第一附属医院儿科神经专科门诊的癫痫患儿组成的耐药性癫痫组(R组)、药物反应性癫痫组(F组)和同期儿童保健门诊体检健康的儿童组成的健康对照组(J组)的外周全血标本各5例,采用高通量测序法检测每个标本miRNAs的表达;以相同条件再次收集R′组(5例)、F′组(7例)和J′组(6例)儿童的外周全血标本,采用实时荧光定量聚合酶链反应(qPCR)验证11个候选miRNAs在3组中的表达;受试者工作特征曲线(ROC)分析7个目标miRNAs区分耐药性癫痫和药物反应性癫痫患儿的功效,在线数据库预测并筛选7个目标miRNAs的靶基因,并对候选靶基因进行基因组百科全书(KEGG)和基因功能(GO)分析。3组间比较采用Kruskal-Wallis秩和检验。结果与F组比较,R组共68个差异表达miRNAs,其中22个miRNAs表达上调,46个miRNAs表达下调。qPCR结果显示,11个候选miRNAs中有7个(let-7f、miR-99a-5p、miR-99b-5p、miR-125a-5p、miR-125b-5p、miR-142-5p、miR-100)表达和高通量测序结果一致。ROC曲线分析发现,当miR-99a-5p、miR-99b-5p、miR-125a-5p、miR-125b-5p、miR-142-5p、miR-100的临界值分别大于0.56、1.00、3.17、2.24、2.09、0.59时,这些miRNAs均有较高的曲线下面积(AUC)(≥0.871)、敏感性(≥80.0%)及特异性(≥85.7%),有望成为诊断儿童耐药性癫痫的生物学标志物,其中miR-125b-5p诊断耐药性癫痫最优。生物信息学分析发现miR-125b-5p可能参与调控的信号通路有缺氧诱导因子-1信号通路、胰岛素信号通路、调控干细胞多能性信号通路、丝裂原活化蛋白激酶信号通路、鞘磷脂信号通路、神经营养蛋白信号通路、哺乳动物类雷帕霉素靶蛋白信号通路。结论耐药性癫痫患儿外周全血中miRNAs的表达谱和药物反应Objective To study the profile of microRNAs(miRNAs)in the peripheral blood of children with drug-resistant epilepsy,and to find diagnostic biomarkers for early identification of drug-resistant epilepsy in children.Methods Retrospective study.Five peripheral blood samples were collected from children in drug-resistant epilepsy group(group R),drug-responsive epilepsy group(group F)composed of the children with epilepsy in pediatric neurology clinic of the First Affiliated Hospital of Zhengzhou University from January 2018 to June 2019 and healthy control group(group J)composed of healthy children who underwent physical examination in the children′s health care clinic at the same time for analyzing miRNA profiles by high-throughput sequencing.In addition,peripheral blood samples were collected from children in R′group(5 cases),F′group(7 cases)and J′group(6 cases)similarly for validating expression levels of 11 candidate miRNAs by quantificational real-time polymerase chain reaction(qPCR).Receiver operating characteristic curves(ROC)were plotted to analyze the diagnostic potential of 7 targeted miRNAs in distinguishing children with drug-resistant epilepsy from drug-responsive epilepsy.Target genes of the 7 validated miRNAs were predicted using online databases,which were then analyzed by Kyoto Encyclopedia of Genes and Genomes(KEGG)and gene ontology(GO).Kruskal-Wallis rank sum test was used for comparison among the three groups.Results High-throughput sequencing found that compared with group F,there were 68 differentially expressed miRNAs in group R,involving 22 up-regulated and 46 down-regulated miRNAs.qPCR results showed that,expression trends of 7 miRNAs(let-7f,miR-99a-5p,miR-99b-5p,and miR-125a-5p,miR-125b-5p,miR-142-5p,miR-100)were consistent with high-throughput sequencing results among the 11 selected miRNAs.ROC analysis found that when the cut-off values of miR-99a-5p,miR-99b-5p,miR-125a-5p,miR-125b-5p,miR-142-5p and miR-100 were greater than 0.56,1.00,3.17,2.24,2.09 and 0.59,respectively,their area

关 键 词：耐药性癫痫 儿童 全血 微小RNAS 

分 类 号：R742.1[医药卫生—神经病学与精神病学]