3-D microwell culture of human embryonic stem cells
3-D microwell culture of human embryonic stem cells
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DOI：
10.1016/j.biomaterials.2006.07.012
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发表时间：
2006-12-01
期刊：
影响因子：
14
通讯作者：
Palecek, Sean P.
中科院分区：
文献类型：
--
作者：
Mohr, Jeffrey C.;de Pablo, Juan J.;Palecek, Sean P.
人类胚胎干细胞（human embryonic stem cells，hESC）具有无限增殖和分化成各种胚胎细胞谱系的能力。需要非常小心地维持未分化的hESC培养物，因为自发分化通常发生在培养物中，推测是由可溶性因子、细胞-细胞接触和/或细胞-基质信号传导引起的。hESC分化通常通过胚状体（EB）的产生来刺激，并且个体细胞的谱系定型取决于整个EB环境中的许多线索，包括EB形状和大小。然而，常见的EB形成方案产生非常不均匀的尺寸分布，可能降低定向分化的效率。我们已经开发了一种基于3-D微孔的方法来维持未分化的hESC培养数周，而不使用物理和细胞外基质图案化限制来限制集落生长。在微孔中培养2-3周的hESC超过90%是活的，并表达hESC转录标记Oct-4。在传代至基质胶包被的组织培养物处理的聚苯乙烯培养皿（TCPS）后，微孔培养的hESC维持未分化的增殖。微孔培养还允许形成具有限定尺寸的hESC集落，其然后可用于形成单分散EB。当在该系统中培养时，hESC保留多能性和自我更新，并且能够传代至标准无约束培养条件。(c)2006爱思唯尔有限公司保留所有权利。
Human embryonic stem cells (hESCs) have the ability to proliferate indefinitely and differentiate into each of the embryonic cell lineages. Great care is required to maintain undifferentiated hESC cultures since spontaneous differentiation often occurs in culture, presumably resulting from soluble factors, cell-cell contact, and/or cell-matrix signaling. hESC differentiation is typically stimulated via generation of embryoid bodies (EBs) and lineage commitment of individual cells depends upon numerous cues throughout the EB environment, including EB shape and size. Common EB formation protocols, however, produce a very heterogeneous size distribution, perhaps reducing efficiency of directed differentiation. We have developed a 3-D microwell-based method to maintain undifferentiated hESC cultures for weeks without passaging using physical and extracellular matrix patterning constraints to limit colony growth. Over 90% of hESCs cultured in microwells for 2-3 weeks were viable and expressed the hESC transcription marker Oct-4. Upon passaging to Matrigel-coated tissue culture-treated polystyrene dishes (TCPS), microwell cultured hESCs maintained undifferentiated proliferation. Microwell culture also permits formation of hESC colonies with a defined size, which can then be used to form monodisperse EBs. When cultured in this system, hESCs retained pluripotency and self-renewal, and were able to be passaged to standard unconstrained culture conditions. (c) 2006 Elsevier Ltd. All rights reserved.