天津医药

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miR-122靶向调控心肌样细胞诱导分化过程中GATA4基因的表达

郭励京1,张志1,刘紫东2,付伟1,季州1   

  1. 1. 辽宁医学院
    2. 辽宁医学院附属第三医院心内科
  • 收稿日期:2013-10-11 修回日期:2013-11-27 出版日期:2014-04-15 发布日期:2014-04-15
  • 通讯作者: 郭励京

GATA4Expression During Induction of the Cardiomyocyte-Like Cells Differentiation Regulated by miR-122

  • Received:2013-10-11 Revised:2013-11-27 Published:2014-04-15 Online:2014-04-15

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摘要： 【摘要】目的 实现大鼠骨髓间充质干细胞（bMSCs）向心肌样细胞的诱导分化，并验证分化过程中miR-122靶向调控GATA4基因的表达。方法首先，分离培养bMSCs，应用5-氮杂胞苷（5-azacytidine）将其诱导分化为心肌样细胞。然后，采用靶基因预测软件miRanda和TargetScan对miR-122和GATA4基因的靶向匹配关系进行预测并通过双荧光素酶报告基因系统鉴定。qRT-PCR检测诱导分化过程中miR-122和GATA4的表达。结果诱导分化后的细胞平行排列，紧密相连，形态规则均一，免疫荧光化学显示有心肌肌钙蛋白I （cTnI）表达。生物信息学方法预测发现miR-122和GATA4基因两者匹配良好，通过双荧光素酶报告基因系统检测发现miR-122能结合到GATA4mRNA 的3′UTR并有效抑制其表达。qRT-PCR检测结果表明，miR-122的表达水平与GATA4表达呈负相关。结论miR- 122能调控心肌样细胞诱导分化过程中GATA4的表达。

关键词: 间质干细胞, 微RNAs, GATA4转录因子, 3'非翻译区, 计算生物学, 大鼠, Sprague-Dawley, miR-122

Abstract:

[Abstract] Objective To identify the miR-122which regulateing GATA4expression during the induction of rat bone marrow mesenchymal stem cells (bMSCs) differentiating into cardiomyocyte-like cells.Methods BMSCswere isolat? ed from bone marrow and induced to differentiate into cardiomyocyte-like cells using5-azacytidine. The miR-122which may regulate expression of GATA4were predicted using miRanda and TargetScan softwares and identified by dual luciferase report system. The expressions of miR-122and GATA4were determined using q-PCR during the differentiation of bMSCs into cardiomyocyte-like cells. Results The induced cells were completely in contacted with adjoining cells and uniform in shape and aligned parallelly. Cardiac troponin I (cTnI) expression was detected by immunofluorescence cytochemistry. Using dual luciferase reporter systemin vitro, miR-122were proved to be able to effectively inhibit GATA4expression by binding the 3′ UTR of GATA4mRNA. q-PCR results showed that the expression of miR-122is negatively correlated with that of GATA4mRNA transcription.Conclusion These results indicated that miR-122regulate the expression of GATA4during the induction of cardiomyocyte-like cells.

Key words: mesenchymal stem cells, MicroRNAs, GATA4 transcription factor, 3' untranslated regions, computational biology, rats, Sprague-Dawley, miR-122