microRNA-140 Targets RALA and Regulates Chondrogenic Differentiation of Human Mesenchymal Stem Cells by Translational Enhancement of SOX9 and ACAN
microRNA-140 Targets RALA and Regulates Chondrogenic Differentiation of Human Mesenchymal Stem Cells by Translational Enhancement of SOX9 and ACAN
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DOI：
10.1089/scd.2013.0209
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发表时间：
2014-02-01
影响因子：
4
通讯作者：
Brinchmann, Jan E.
中科院分区：
文献类型：
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作者：
Karlsen, Tommy A.;Jakobsen, Rune B.;Brinchmann, Jan E.
关节软骨损伤不能自愈。一种治疗策略是通过间充质干细胞（MSCs）的定向软骨分化在实验室中制造软骨。为了促进我们对软骨形成的分子控制的理解，我们比较了MSCs体外软骨形成过程中microrna （mirna）的变化与未培养和去分化关节软骨细胞（ACs）中观察到的变化。在MSCs的分化和ACs的去分化过程中，几种mirna表现出相互关系。miR-140-5p和miR-140-3p在体外软骨形成过程中变化最大，它们是组织工程软骨细胞中表达最高的mirna，也是未培养的ACs中表达最高的mirna。在分化的MSCs和未培养的ACs中，100个最高表达的mirna有57%的重叠，但对于其他mirna，表达模式则有很大不同。我们分别在分化的MSCs和去分化的ac中短暂稳定地抑制和过表达miR-140-5p和miR-140-3p，以描述全局效应并识别和验证新的靶点。令人惊讶的是，尽管mRNA水平不变，但在anti-miR-140转导的分化MSCs中，SOX9和聚集蛋白被发现下调。这表明miR-140在蛋白质水平上通过上调这些分子刺激体外软骨形成。RALA是一种小的GTPase，被确定为miR-140的靶标，敲低实验表明RALA在蛋白水平上调控SOX9。这些观察结果揭示了miR-140在体外和体内对软骨形成的影响。
Lesions of articular cartilage do not heal spontaneously. One treatment strategy would be to make cartilage in the laboratory by directed chondrogenic differentiation of mesenchymal stem cells (MSCs). To promote our understanding of the molecular control of chondrogenesis, we have compared the changes in microRNAs (miRNAs) during in vitro chondrogenesis of MSCs with those observed in uncultured and dedifferentiated articular chondrocytes (ACs). Several miRNAs showed a reciprocal relationship during the differentiation of MSCs and dedifferentiation of ACs. miR-140-5p and miR-140-3p changed the most during in vitro chondrogenesis, they were the miRNAs most highly expressed in tissue-engineered chondrocytes, and they were also among the miRNAs most highly expressed in uncultured ACs. There was a 57% overlap for the 100 most highly expressed miRNAs in differentiated MSCs and uncultured ACs, but for other miRNAs, the expression pattern was quite different. We transiently and stably inhibited and overexpressed miR-140-5p and miR-140-3p in differentiating MSCs and dedifferentiating ACs, respectively, to describe global effects and identify and validate new targets. Surprisingly, SOX9 and aggrecan proteins were found to be downregulated in anti-miR-140 transduced differentiating MSCs despite unchanged mRNA levels. This suggests that miR-140 stimulates in vitro chondrogenesis by the upregulation of these molecules at the protein level. RALA, a small GTPase, was identified as a miR-140 target and knockdown experiments showed that RALA regulated SOX9 at the protein level. These observations shed new light on the effect of miR-140 for chondrogenesis in vitro and in vivo.