hsa-miR-143在人脂肪细胞诱导分化过程中的表达变化及调控因素分析

  本文关键词：hsa-miR-143在人脂肪细胞诱导分化过程中的表达变化及调控因素分析，由笔耕文化传播整理发布。

        目的（1）探讨hsa-miR-143在脂肪细胞分化前后的表达变化规律；（2）对hsa-miR-143进行系统的生物信息学分析，预测hsa-miR-143的靶基因和可能参与的功能；（3）FFAs（free fatty acids，游离脂肪酸）、TNF-（tumor necrosisfactor-alpha，肿瘤坏死因子alpha）、IL-6（interleukin6，白介素6）、leptin（瘦素）、resistin（抵抗素）分别对人成熟脂肪细胞hsa-miR-143表达的影响。方法（1）利用Real time PCR技术，检测脂肪细胞分化前后hsa-miR-143的表达水平；（2）应用miRbase、NCBI Mapviewer、UCSC Genome Browser等在线工具获取并分析hsa-miR-143的序列特征；应用TargetScan，PicTar及miRanda预测hsa-miR-143的靶基因，取三个预测结果的交集，并合并DIANALAB-TarBase6.0数据库的已验证靶标，对所得靶基因集分别进行功能富集分析（Gene Ontology, GO）和信号转导通路富集分析（Pathway Enrichment）；应用pubmed、google等信息搜索工具综述hsa-miR-143已有功能研究；（3）利用Realtime PCR技术检测FFAs、TNF-、IL-6、leptin、resistin干预成熟脂肪细胞前后hsa-miR-143的表达水平的变化。结果（1）分化成熟的人脂肪细胞hsa-miR-143的表达量较分化前显著上调（P<0.05）；（2）hsa-miR-143在各物种之间具有高度保守性，功能注释显示预测靶基因主要富集于肌动蛋白细胞骨架组织合成、Rac蛋白信号转导的正调、突触传递的调控等生物学过程（P<0.001）；（3）Pathway分析显示预测靶基因显著富集于肌动蛋白细胞骨架调节、黑色素瘤、细胞缝隙连接、MAPK（Mitogen-activated protein kinase，丝裂原激活蛋白激酶）信号通路等（P<0.05）；（4）FFAs, leptin及resistin显著下调人成熟脂肪细胞hsa-miR-143的表达（P<0.05），但TNF-及IL-6对其表达量影响较小（P>0.05）。结论（1）hsa-miR-143在成熟人脂肪细胞中，其表达量达到相对较高的水平，说明在脂肪细胞分化成熟过程中，其有一定的调控作用，生物信息学分析提示hsa-miR-143可能与细胞分化、代谢密切相关；（3）FFAs、leptin及resistin能显著下调成熟人脂肪细胞hsa-miR-143的表达，TNF-及IL-6对成熟人脂肪细胞hsa-miR-143的表达无明显作用。研究结果提示，脂肪细胞hsa-miR-143的表达，可能受FFAs、leptin及resistin因素的调节，并推论，，FFAs、leptin及resistin对miR-143的表达可能存在有一个负反馈调节机制，这一推论仍需验证。本研究阐明了hsa-miR-143在人脂肪前体细胞诱导分化过程中的表达规律，提示了FFAs、Leptin、Resistin等因素对成熟人脂肪细胞hsa-miR-143的表达具有一定的调控作用。研究结果为深入分析hsa-miR-143与肥胖、胰岛素敏感性之间的关系提供了新的线索，并为肥胖的早期干预及肥胖相关并发症的防治提供新的靶点。

    Objective （1） The different expression of hsa-miR-143in the process ofdifferentiation in human adipocytes was investigated.（2）To predict target gene andfunctions of hsa-miR-143by bioinformatic analysis.（3） The alteration ofhsa-miR-143expression in matured human adipocytes were studied after intervenedby FFAs and some adipokines such as TNF-, IL-6, leptin and resistin.Methods The levels of hsa-miR-143expression in the human preadipocytes thathad differentiated into mature adipocytes were analyzed using quantitative real timePCR. MiRBase, NCBI Mapviewer, UCSC Genome Browser database were used toget the sequence of hsa-miR-143. The target gene were predicted by TargetScan,PicTar and miRanda, and the intersection of results of three database with knowntarget gene were analyzed by Gene Ontology and pathway analysis. Pubmed andgoogle were used to review previous studies of hsa-miR-143. And furthermore, theexpression of hsa-miR-143were measured in mature adipocytes treated with FFAs,TNF-, IL-6, leptin and resistin were analyzed using quantitative real time PCR.Results （1）During the conversion of cultured human preadipocytes to matureadipocytes, the expression of hsa-miR-143was upregulated (P<0.05).（2）Hsa-miR-143was highly conserved among species. By gene ontology analysis, thetarget genes were enriched in actin cytoskeleton organization and biogenesis,positively regulating Rac protein signal transduction, synaptic transmission, and otherbiological processes (P<0.001).（3）By pathway analysis, the target genes weremainly involved in regulation of actin cytoskeleton, Melanoma, Gap junction, MAPKsignaling pathway, and others (P<0.05).（4）The expression of hsa-miR-143were downregulated by FFAs, leptin and resistin in human adipocytes (P<0.05), but not byTNF-and IL-6(P>0.05).Conclusions （1）During the conversion of cultured human preadipocytes to matureadipocytes, the expression of hsa-miR-143is upregulated. Bioinformatic Analysisresults show hsa-miR-143is closely related to cell differentiation and metabolism.（2）hsa-miR-143is inhibited by FFAs,leptin, and resistin,but not by TNF-and IL-6in cultered human adipocytes. These findings indicate that FFAs, leptin, and resistin,may inhibit the expression of miR-143via a negative feedback which requires furtherstudy. In conclusion, we identify the regularity of miR-143expression during theconversion of human preadipocytes into mature adipocytes and several regulativefactors on the miR-143expression in cultured human adipocytes. These data providefurther evidence of the partial involvement of hsa-miR-143in the regulation ofinsulin sensitivity mediated by certain adipokines. Our findings may bring newinsights into the mechanisms of obesity-associated insulin resistance.

hsa-miR-143在人脂肪细胞诱导分化过程中的表达变化及调控因素分析

中文摘要5-7Abstract7-8前言9-12第一部分 hsa-miR-143 在人脂肪细胞诱导分化过程中的表达变化及生物信息学分析12-37    材料与方法12-23    结果23-34    讨论34-37第二部分 hsa-miR-143 在成熟人脂肪细胞中调控因素的分析37-49    材料与方法37-40    结果40-46    讨论46-49参考文献49-55中英文缩略语55-56综述56-65    参考文献62-65已撰写和发表文章65-66致谢66

  本文关键词：hsa-miR-143在人脂肪细胞诱导分化过程中的表达变化及调控因素分析，由笔耕文化传播整理发布。