McKiernan Eadaoin, O'Driscoll Lorraine, Kasper Michael, Barron Niall, O'Sullivan Finbarr, Clynes Martin
National Institute for Cellular Biotechnology, Dublin City University, Dublin, Ireland.
Tissue Eng. 2007 Oct;13(10):2419-30. doi: 10.1089/ten.2006.0373.
The derivation of definitive endoderm and in particular endocrine cell types from undifferentiated embryonic stem (ES) cells remains difficult to achieve. In this study, we investigated the potential to regulate the differentiation of ES cells into endodermal derivatives using extracellular factors previously associated with various aspects of pancreatic development. Feeder-free-cultured mouse ESD3 cells were manipulated to form embryoid bodies (EBs) in the presence of retinoic acid (RA). RA-treated EBs were subsequently exposed to sodium butyrate (SB), betacellulin (BTC) or activin A (AA). A comparative analysis was performed on these models of directed differentiation in parallel with a model of spontaneous differentiation. Lineage differentiation was determined by profiling multilineage marker transcript expression (neuronal, myogenic, exocrine and endocrine pancreas, extraembryonic and apoptotic) and subsequent protein expression within ES-derived cultures. Using a two-stage differentiation protocol developed during this study, we successfully demonstrated the derivation of an intermediate multipotential population (RA_EBs) from undifferentiated ES cells that preferentially gives rise to pancreatic endocrine insulin-expressing cell types in the presence of SB, and neuronal- and glial-like cell types in the presence of AA or BTC.
从未分化的胚胎干细胞（ES细胞）中获得确定的内胚层，尤其是内分泌细胞类型，仍然很难实现。在本研究中，我们利用先前与胰腺发育各方面相关的细胞外因子，研究了调控ES细胞分化为内胚层衍生物的潜力。在无饲养层培养的小鼠ESD3细胞在视黄酸（RA）存在的情况下被诱导形成胚状体（EBs）。随后，用丁酸钠（SB）、β细胞素（BTC）或激活素A（AA）处理经RA处理的EBs。对这些定向分化模型与自发分化模型进行了比较分析。通过分析多谱系标记转录本表达（神经元、肌源性、外分泌和内分泌胰腺、胚外和凋亡相关）以及ES细胞来源培养物中的后续蛋白质表达，来确定谱系分化。使用本研究中开发的两阶段分化方案，我们成功地从未分化的ES细胞中获得了一个中间多能群体（RA_EBs），在SB存在的情况下，该群体优先产生表达胰岛素内分泌胰腺细胞类型，在AA或BTC存在的情况下，则产生神经元样和神经胶质样细胞类型。
