Endothelial lineage differentiation from induced pluripotent stem cells is regulated by microRNA-21 and transforming growth factor β2 (TGF-β2) pathways.
Endothelial lineage differentiation from induced pluripotent stem cells is regulated by microRNA-21 and transforming growth factor β2 (TGF-β2) pathways.
复制标题
DOI：
10.1074/jbc.m113.495531
复制
发表时间：
2014-02-07
期刊：
影响因子：
--
通讯作者：
Xu Q
中科院分区：
文献类型：
--
作者：
Di Bernardini E;Campagnolo P;Margariti A;Zampetaki A;Karamariti E;Hu Y;Xu Q
背景：诱导多能干细胞（iPSC）是再生医学的一个有吸引力的细胞来源。结果：MicroRNA-21在VEGF存在下介导源自iPSC的内皮分化。结论：MicroRNA-21和TGF-β2信号通路调控iPSC向内皮细胞分化。重要性：阐明microRNA-21调控iPSC分化的分子机制可能为血管疾病的干细胞治疗提供基础信息。为心血管再生寻找合适的内皮细胞来源是再生医学的一个挑战性问题。在本文中，我们描述了一种新的机制，调节诱导多能干细胞（iPSC）分化为EC，特别关注miRNA及其靶点。我们首先建立了一个使用胶原IV和VEGF来驱动iPSC向EC的功能分化的方案，并比较了分化和未分化细胞的miRNA特征。在分化细胞中过度表达的miRNA中，我们专注于microRNA-21（miR-21）并研究其在iPSC分化中的作用。miR-21在预分化iPSCs中的过表达诱导EC标志物上调以及体外和体内毛细血管形成;因此，抑制miR-21产生相反的效果。重要的是，miR-21过表达增加了TGF-β2 mRNA和分泌蛋白水平，与iPSC分化期间TGF-β2的强烈上调一致。事实上，用TGFβ-2处理iPSC诱导EC标志物表达和体外管形成。SMAD 3是TGFβ-2的下游效应子，抑制SMAD 3可显著降低VE-钙粘蛋白的表达。此外，TGFβ-2中和和敲低抑制miR-21诱导的EC标志物表达。最后，我们证实了PTEN/Akt通路是miR-21的直接靶点，并且我们表明PTEN敲低是miR-21介导的内皮分化所必需的。总之，我们阐明了一种新的信号通路，促进iPSC分化为适用于再生医学应用的功能性EC。
Background: Induced pluripotent stem cells (iPSCs) constitute an attractive source of cells for regenerative medicine. Results: MicroRNA-21 mediates endothelial differentiation derived from iPSCs in presence of VEGF. Conclusion: MicroRNA-21 and TGF-β2 signaling pathways regulate iPSC differentiation to endothelial lineage. Significance: Elucidation of the molecular mechanisms underlying microRNA-21-regulated iPSC differentiation might provide the basic information for stem cell therapy of vascular diseases. Finding a suitable cell source for endothelial cells (ECs) for cardiovascular regeneration is a challenging issue for regenerative medicine. In this paper, we describe a novel mechanism regulating induced pluripotent stem cells (iPSC) differentiation into ECs, with a particular focus on miRNAs and their targets. We first established a protocol using collagen IV and VEGF to drive the functional differentiation of iPSCs into ECs and compared the miRNA signature of differentiated and undifferentiated cells. Among the miRNAs overrepresented in differentiated cells, we focused on microRNA-21 (miR-21) and studied its role in iPSC differentiation. Overexpression of miR-21 in predifferentiated iPSCs induced EC marker up-regulation and in vitro and in vivo capillary formation; accordingly, inhibition of miR-21 produced the opposite effects. Importantly, miR-21 overexpression increased TGF-β2 mRNA and secreted protein level, consistent with the strong up-regulation of TGF-β2 during iPSC differentiation. Indeed, treatment of iPSCs with TGFβ-2 induced EC marker expression and in vitro tube formation. Inhibition of SMAD3, a downstream effector of TGFβ-2, strongly decreased VE-cadherin expression. Furthermore, TGFβ-2 neutralization and knockdown inhibited miR-21-induced EC marker expression. Finally, we confirmed the PTEN/Akt pathway as a direct target of miR-21, and we showed that PTEN knockdown is required for miR-21-mediated endothelial differentiation. In conclusion, we elucidated a novel signaling pathway that promotes the differentiation of iPSC into functional ECs suitable for regenerative medicine applications.