A novel combination of factors, termed SPIE, which promotes dopaminergic neuron differentiation from human embryonic stem cells.
A novel combination of factors, termed SPIE, which promotes dopaminergic neuron differentiation from human embryonic stem cells.
复制标题
DOI：
10.1371/journal.pone.0006606
复制
发表时间：
2009-08-12
期刊：
影响因子：
3.7
通讯作者：
Freed WJ
中科院分区：
文献类型：
--
作者：
Vazin T;Becker KG;Chen J;Spivak CE;Lupica CR;Zhang Y;Worden L;Freed WJ
基质衍生诱导活性 (SDIA) 是从胚胎干细胞 (ESC) 生成多巴胺能 (DA) 神经元的最有效方法之一。可以通过将 ESC 与小鼠基质细胞系 PA6 或 MS5 共培养来实现 DA 神经元诱导。这种被称为“SDIA”的效应的分子性质迄今为止尚不清楚。最近，我们发现PA6细胞分泌的因子提供谱系特异性指令来诱导人ESC（hESC）的DA分化。在本研究中，我们将 PA6 细胞与缺乏 SDIA 效应的各种细胞系进行比较，并采用基因组表达分析来识别差异表达的信号分子。 PA6 细胞高表达且已知与 CNS 发育相关的因子包括基质细胞衍生因子 1 (SDF-1/CXCL12)、多效素 (PTN)、胰岛素样生长因子 2 (IGF2) 和肝配蛋白 B1 (EFNB1)。当这四种因子（其组合被称为 SPIE）应用于 hESC 时，它们在体外诱导分化为 TH 阳性神经元。 RT-PCR 和蛋白质印迹分析证实了受这四种分子影响的培养物中中脑特异性标记物的表达，包括 engrailed 1、Nurr1、Pitx3 和多巴胺转运蛋白 (DAT)。电生理记录表明，用 SPIE 处理 hESC 可诱导神经元分化，这些神经元能够产生动作电位并形成功能性突触连接。 SDF-1、PTN、IGF2 和 EFNB1 的组合模拟了 SDIA 的 DA 表型诱导特性，并且足以促进 hESC 分化为功能性中脑 DA 神经元。这些发现提供了一种分化 hESC 形成 DA 神经元的方法，无需使用动物源细胞系或产品。
Stromal-Derived Inducing Activity (SDIA) is one of the most efficient methods of generating dopaminergic (DA) neurons from embryonic stem cells (ESC). DA neuron induction can be achieved by co-culturing ESC with the mouse stromal cell lines PA6 or MS5. The molecular nature of this effect, which has been termed “SDIA” is so far unknown. Recently, we found that factors secreted by PA6 cells provided lineage-specific instructions to induce DA differentiation of human ESC (hESC). In the present study, we compared PA6 cells to various cell lines lacking the SDIA effect, and employed genome expression analysis to identify differentially-expressed signaling molecules. Among the factors highly expressed by PA6 cells, and known to be associated with CNS development, were stromal cell-derived factor 1 (SDF-1/CXCL12), pleiotrophin (PTN), insulin-like growth factor 2 (IGF2), and ephrin B1 (EFNB1). When these four factors, the combination of which was termed SPIE, were applied to hESC, they induced differentiation to TH-positive neurons in vitro. RT-PCR and western blot analysis confirmed the expression of midbrain specific markers, including engrailed 1, Nurr1, Pitx3, and dopamine transporter (DAT) in cultures influenced by these four molecules. Electrophysiological recordings showed that treatment of hESC with SPIE induced differentiation of neurons that were capable of generating action potentials and forming functional synaptic connections. The combination of SDF-1, PTN, IGF2, and EFNB1 mimics the DA phenotype-inducing property of SDIA and was sufficient to promote differentiation of hESC to functional midbrain DA neurons. These findings provide a method for differentiating hESC to form DA neurons, without a requirement for the use of animal-derived cell lines or products.
登录
查看更多内容
2001 年 8 月 9 日之前提取的三种人类胚胎干细胞系的核型稳定性、基因分型、分化、无饲养维持和基因表达取样
影响因子：
4
作者：
Brimble, SN;Zeng, XM;Schulz, TC
通讯作者：
Schulz, TC
影响因子：
5.8
作者：
Al-Shahrour, F;Díaz-Uriarte, R;Dopazo, J
通讯作者：
Dopazo, J
影响因子：
3
作者：
Castelo-Branco, Goncalo;Arenas, Ernest
通讯作者：
Arenas, Ernest
影响因子：
--
作者：
Bhattacharya B;Cai J;Luo Y;Miura T;Mejido J;Brimble SN;Zeng X;Schulz TC;Rao MS;Puri RK
通讯作者：
Puri RK
影响因子：
5.2
作者：
Chiba, Shunmei;Lee, Young Mook;Freed, Curt R.
通讯作者：
Freed, Curt R.