Drury-Stewart Danielle, Song Mingke, Mohamad Osama, Yu Shan Ping, Wei Ling
Emory University School of Medicine, Department of Anesthesiology, Woodruff Circle, Woodruff Memorial Research Building Suite 617, Atlanta, GA 30322 USA.
J Stem Cells. 2011;6(1):1-7.
While human embryonic stem (hES) and induced pluripotent stem (hiPS) cells offer exciting prospects in the fields of regenerative medicine and developmental biology, efficient directed differentiation of these cells is still difficult. Neural induction protocols often include suspension culture or co-culture with other cell types, introducing heterogeneity and complicating analysis. In addition, expensive recombinant factors are often used over processes that take weeks to complete, making such experiments financially difficult. We have developed a fully adherent and feeder free neural differentiation protocol using small molecules such as dorsomorphin and common medium supplements. Using this protocol, we obtain >90% of cells developing into neural precursors, as measured by nestin staining. Neurons derived from these precursors are electrophysiologically active. After three weeks of terminal differentiation, we obtain functional neurons which fire high-amplitude action potentials upon depolarization. A subset of neurons also fires repetitive trains. This protocol offers a simpler and less expensive method for investigations involving the differentiation of neural precursors and neurons in culture.
虽然人类胚胎干细胞（hES）和诱导多能干细胞（hiPS）在再生医学和发育生物学领域展现出了令人兴奋的前景，但对这些细胞进行高效的定向分化仍然具有挑战性。神经诱导方案通常包括悬浮培养或与其他细胞类型共培养，这会引入异质性并使分析变得复杂。此外，在耗时数周才能完成的过程中，常常会使用昂贵的重组因子，这使得此类实验在经济上难以开展。我们开发了一种完全贴壁且无饲养层的神经分化方案，使用诸如多斯莫尔芬等小分子和常见的培养基添加剂。通过此方案，我们获得了超过90%的细胞发育为神经前体细胞，这是通过巢蛋白染色来测定的。源自这些前体细胞的神经元具有电生理活性。经过三周的终末分化，我们获得了功能性神经元，这些神经元在去极化时会产生高幅度动作电位。一部分神经元还会产生重复放电序列。该方案为涉及培养中神经前体细胞和神经元分化的研究提供了一种更简单且成本更低的方法。
