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microRNA在胚胎干细胞分化的心肌细胞中的表达变化

陈斐, 陈忠炎, 杨黄恬^*

上海交通大学医学院/中国科学院上海生命科学研究院健康科学研究所，中国科学院干细胞生物学重点实验室，上海 200031

摘要

胚胎干细胞(embryonic stem cells, ESCs)具有自我更新和多向分化的特性。近年研究显示转录后调控包括microRNAs (miRNAs)的调控在ESC心肌细胞分化(ESC-derived cardiomyocytes, ESCM)命运决定中起着重要作用。然而对决定ESC分化命运的miRNAs的了解还非常有限。为了进一步认识miRNAs对心肌细胞分化的调控作用，本研究采用经典的悬滴法诱导小鼠ESC (mESC)分化成心肌细胞，采用安捷伦8 × 15k小鼠miRNAs芯片(miRbase V16.0)比较了富含跳动心肌细胞区域与非跳动区域miRNAs表达谱的异同，发现与非跳动区域相比，跳动区域中有19个miRNAs发生了5倍以上的表达变化(n = 3, P < 0.05)，其中5个miRNAs表达上调、14个miRNAs表达下调。采用定量RT-PCR进一步分析了miRNAs芯片中差异倍数大于10的miRNAs，证明miR-196a，miR-196b和miR-467e在心肌细胞跳动区域的表达丰度明显低于非跳动区域(n = 3, P < 0.05)。用TargetScan数据库对miR-196a和miR-196b进行靶基因预测，发现其可能与心肌细胞分化呈负相关，提示其在ESC分化命运决定中可能具有一定的调控作用。这些结果为进一步明确miRNAs在ESC分化的心肌细胞中的作用提供了新线索。

关键词： 小鼠胚胎干细胞; microRNA; 心肌细胞; microRNA芯片; 分化

分类号：Q28

[Expression profile of microRNAs in the cardiomyocytes derived from mouse embryonic stem cells.] [Article in Chinese]

CHEN Fei, CHEN Zhong-Yan, YANG Huang-Tian^*

Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Jiao Tong University School of Medicine (SJTUSM) &； Shanghai Institutes for Biological Sciences (SIBS), Chinese Academy of Sciences (CAS), Shanghai 200031, China

Abstract

Embryonic stem cells (ESCs), derived from the inner cell mass of blastocysts, are self-renewing and pluripotent cells with the ability to differentiate into all derivatives of three primary germ layers, including cardiomyocytes. Recent studies have revealed that posttranscriptional regulations of lineage specific genes by microRNAs (miRNAs) emerge as a new class of cell fate and lineage determinants of ESCs. However, the miRNAs that control ESC differentiation are still largely unexplored. In the present study, we aimed to identify miRNAs that might be involved in cardiac differentiation of ESCs. Using a hanging drop technique, mouse ESCs (mESCs) were differentiated into cardiomyocytes. We then used the Aligent miRNAs chip (miRbase V16.0) to evaluate miRNA expression levels between the ESC-derived beating area enriched with cardiomyocytes and non-beating area. The expression levels of 19 miRNAs changed over 5-fold between two areas (n = 3, P < 0.05). Among them, 5 miRNAs were upregulated and 14 miRNAs were downregulated in the beating area compared with the non-beating area (P < 0.05). Then quantitative real-time-PCR was used to analyze the miRNAs with the differentiated expression level over 10-fold seen in the Aligent miRNAs chip. miR-196a, miR-196b and miR-467e were confirmed to be significantly lower in the beating area than those in the non-beating area (n = 3, P < 0.05). TargetScan analysis further suggested that miR-196a and miR-196b might be negatively related to the cardiomyocytes differentiation. Our findings provide a new clue for exploring roles of miRNAs in cardiac lineage commitment of mESCs.

Key words: embryonic stem cell; microRNA; cardiomyocyte; microRNA array; differentiation

收稿日期：2014-04-13　　录用日期：2014-05-14

通讯作者：杨黄恬　　E-mail: htyang@sibs.ac.cn

引用本文：

陈斐, 陈忠炎, 杨黄恬. microRNA在胚胎干细胞分化的心肌细胞中的表达变化[J]. 生理学报 2014; 66 (6): 702-708.

CHEN Fei, CHEN Zhong-Yan, YANG Huang-Tian. [Expression profile of microRNAs in the cardiomyocytes derived from mouse embryonic stem cells.] [Article in Chinese]. Acta Physiol Sin 2014; 66 (6): 702-708 (in Chinese with English abstract).