高脂饲料诱导的胰岛素抵抗小鼠肝脏组织差异表达microRNAs的鉴定及生物信息学分析  

Identification and bioinformatics analysis of differentially expressed microRNAs in mice liver tissues with a high fat diet-induced insulin resistance

作　　者：于曼丽[1] 王文栋[1] 饶小娇 郝敏[1] 吴亚柳 常晓彤[1] 

机构地区：[1]河北北方学院临床检验诊断学重点实验室,张家口075000

出　　处：《卫生研究》2016年第6期897-905,共9页Journal of Hygiene Research

基　　金：河北省自然科学基金(No.C2011405015);河北北方学院创新人才培育项目(No.CXRC1316)

摘　　要：目的采用茎环-逆转录实时荧光定量PCR方法,鉴定高脂饲料诱导的胰岛素抵抗小鼠肝脏组织中差异表达的mircoRNAs(miRNAs),即miR-1897-3p、miR-690和miR-7a-5p,并预测分析miRNAs调控的靶基因和功能,探讨胰岛素抵抗与差异表达的miRNAs的相关性。方法 30只雄性C57BL/6小鼠随机分为对照组和高脂饲料诱导的胰岛素抵抗模型组,设计茎环引物和实时荧光定量PCR特异引物,建立茎环-逆转录SYBR Green I实时荧光定量PCR方法,检测小鼠肝脏组织中miR-1897-3p、miR-690和miR-7a-5p的表达。统计学分析小鼠miR-1897-3p、miR-690和miR-7a-5p表达的差异。利用生物信息学软件预测miRNAs调控的靶基因,分析靶基因富集的基因功能(gene ontology,GO)和涉及到的信号转导通路及其靶基因蛋白的相互作用。结果经实时荧光定量PCR鉴定分析,与对照组相比,胰岛素抵抗组小鼠肝脏组织中miR-1897-3p、miR-690、miR-7a-5p表达下调(P<0.05)。生物信息学分析结果显示,有16种靶基因被两种差异表达的miRNAs调控,其中有8个靶基因可与4种以上的蛋白质相互作用,Rac1、Rhoa、Prkcz、Tgfbr2、Itch和Ube2d3蛋白位于网络的中心节点,且它们与胰岛素信号通路相关或存在交联。结论肝脏miR-1897-3p、miR-690和miR-7a-5p可能参与了胰岛素抵抗的病理生理过程,其机制可能通过调节靶基因的表达,影响了胰岛素信号通路的正常级联反应。Objective To identify the expression levels of miR-1897-3p,miR-690 and miR-7a-5p in mice liver tissues with a high fat diet-induced insulin resistance using a stem-loop reverse transcriptional real-time fluorescence quantitative PCR（ RT-q PCR）method,and predict microRNAs（ miRNAs）-regulated target genes and their functions to investigate the relationship between insulin resistance and differentially expressed miRNAs.Methods The total of 30 liver tissue samples were obtained from 15 normal control mice and 15 test mice with a high fat diet-induced insulin resistance.The stemloop reverse transcriptional primers and the primers of real-time PCR were designed to establish a RT-q PCR method,and then the expression levels of miR-1897-3p,miR-690 and miR-7a-5p of 30 liver tissue samples were detected and analyzed.Using bioinformatics methods, the target genes of differentially expressed miRNAs were predicted,and then enriched gene ontology（ GO）,related signal pathways and target gene protein-target gene protein interactions were analyzed,respectively.Results Compared with control group, the expression levels of miR-1897-3p, miR-690 and miR-7a-5p detected by RT-q PCR in the group with a high-fat diet-induced insulin resistance were significantly decreased（ P 〈 0.05）,which exhibited the similar pattern of down regulation to the previous microarray results.Bioinformatics analysis results showed that a total of 16 target genes were regulated by two differentially expressed miRNAs.Among of the 16 target gene protein,8 proteins had interactions with ≥4 proteins.Rac1,Rhoa,Prkcz,Tgfbr2,Itch and Ube2d3 protein were located in the central node of the network,and they were associated or cross-linking with insulin signaling pathway.Conclusion miR-1897-3p,miR-690 and miR-7a-5p in liver tissues may be involved in the physiopathologic process of insulin resistance,which may be affect the normal insulin signaling pathway cascade by regulating the expression of target genes.

关 键 词：胰岛素抵抗 微小RNAS 实时荧光定量PCR 生物信息学 

分 类 号：Q593.1[生物学—生物化学] Q578