Small Molecule Mesengenic Induction of Human Induced Pluripotent Stem Cells to Generate Mesenchymal Stem/Stromal Cells
Small Molecule Mesengenic Induction of Human Induced Pluripotent Stem Cells to Generate Mesenchymal Stem/Stromal Cells
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DOI：
10.5966/sctm.2011-0022
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发表时间：
2012-02-01
影响因子：
6
通讯作者：
Fisk, Nicholas M.
中科院分区：
文献类型：
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作者：
Chen, Yen Shun;Pelekanos, Rebecca A.;Fisk, Nicholas M.
间充质干/基质细胞（MSC）的翻译潜力受到其在体器官中的稀有性、异质性和需要通过侵入性程序收获的限制。诱导多能干细胞（iPSC）可能是MSC的有利来源，但从多能细胞衍生MSC的尝试需要繁琐或不可翻译的技术，如共培养、物理操作、分选或病毒转导。我们设计了一种单步方法，使用小分子抑制剂指导人胚胎干细胞（ESC）和iPSC的间充质分化。首先，通过在含有转化生长因子-β途径抑制剂SB 431542的无血清培养基中培养ESC/iPSC来产生上皮样单层细胞。10天后，iPSC显示中胚层基因（MSX 2、NCAM、HOXA 2）的上调和多能性基因（OCT 4、LEFTY 1/2）的下调。然后通过将细胞转移到常规MSC培养基中完成分化。由此产生的MSC样形态的发展与基因表达的增加有关，反映了上皮细胞向间质细胞的转化。ESC和iPSC衍生的MSC均表现出典型的MSC免疫表型，表达高水平的波形蛋白和N-钙粘蛋白，并且在蛋白质水平上缺乏多能性标志物的表达。在体外ES-MSC和iPS-MSC中诱导了稳健的成骨和成软骨分化，而成脂分化是有限的，如原始胎儿MSC和通过其他方法衍生的ES-MSC所报道的。我们的结论是，在二维培养物中用SB 431542处理，然后培养诱导的上皮向间充质转化，导致人多能细胞的快速和均匀的MSC转化，而不需要胚状体形成或饲养细胞共培养，为从人iPSC产生MSC提供了一种稳健的、临床适用的和有效的系统。干细胞转化医学2012;1：83-95
The translational potential of mesenchymal stem/stromal cells (MSCs) is limited by their rarity in somatic organs, heterogeneity, and need for harvest by invasive procedures. Induced pluripotent stem cells (iPSCs) could be an advantageous source of MSCs, but attempts to derive MSCs from pluripotent cells have required cumbersome or untranslatable techniques, such as coculture, physical manipulation, sorting, or viral transduction. We devised a single-step method to direct mesengenic differentiation of human embryonic stem cells (ESCs) and iPSCs using a small molecule inhibitor. First, epithelial-like monolayer cells were generated by culturing ESCs/iPSCs in serum-free medium containing the transforming growth factor-beta pathway inhibitor SB431542. After 10 days, iPSCs showed upregulation of mesodermal genes (MSX2, NCAM, HOXA2) and downregulation of pluripotency genes (OCT4, LEFTY1/2). Differentiation was then completed by transferring cells into conventional MSC medium. The resultant development of MSC-like morphology was associated with increased expression of genes, reflecting epithelial-to-mesenchymal transition. Both ESC- and iPSC-derived MSCs exhibited a typical MSC immunophenotype, expressed high levels of vimentin and N-cadherin, and lacked expression of pluripotency markers at the protein level. Robust osteogenic and chondrogenic differentiation was induced in vitro in ES-MSCs and iPS-MSCs, whereas adipogenic differentiation was limited, as reported for primitive fetal MSCs and ES-MSCs derived by other methods. We conclude that treatment with SB431542 in two-dimensional cultures followed by culture-induced epithelial-to-mesenchymal transition leads to rapid and uniform MSC conversion of human pluripotent cells without the need for embryoid body formation or feeder cell coculture, providing a robust, clinically applicable, and efficient system for generating MSCs from human iPSCs. STEM CELLS TRANSLATIONAL MEDICINE 2012;1:83-95