Scalable culture and cryopreservation of human embryonic stem cells on microcarriers.
Scalable culture and cryopreservation of human embryonic stem cells on microcarriers.
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DOI：
10.1002/btpr.110
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发表时间：
2009-01
影响因子：
2.9
通讯作者：
Palecek, Sean P.
中科院分区：
文献类型：
--
作者：
Nie, Ying;Bergendahl, Veit;Hei, Derek J.;Jones, Jeffrey M.;Palecek, Sean P.
由于其多能性和无限自我更新的潜力，人类胚胎干细胞（hESC）在再生医学领域具有巨大的前景。 hESCs广泛应用的一个重要先决条件是建立有效且高效的大规模细胞培养、储存和分配方案。在实验室规模中，hESC 贴壁培养；这些培养技术是劳动密集型的，并且不能扩展到高细胞数量。为了促进更大规模的 hESC 培养，我们研究了培养粘附于微载体的 hESC 的可行性。我们用 Matrigel 或小鼠胚胎成纤维细胞 (MEF) 修饰了 Cytodex 3 微载体的表面。 hESC 集落在基质胶包被的微载体和接种 MEF 单层的微载体上以多能、未分化状态有效扩增。虽然 MEF 微载体上的 hESC 扩增率低于 MEF 板上的 hESC 扩增率，但 Matrigel 微载体上 hESC 的倍增时间与 Matrigel 包被的组织培养板上扩增的 hESC 的倍增时间没有区别。标准 hESC 冷冻保存方法存在生存能力差和解冻后分化率高的问题。在这里，我们证明，与悬浮细胞冷冻保存相比，在冷冻管中冷冻保存粘附在微载体上的 hESC 可提供更高的未分化细胞回收率。总之，这些结果表明基于微载体的稳定和培养可以促进 hESC 的扩增和储存，以用于研究和治疗应用。
As a result of their pluripotency and potential for unlimited self-renewal, human embryonic stem cells (hESCs) hold tremendous promise in regenerative medicine. An essential prerequisite for the widespread application of hESCs is the establishment of effective and efficient protocols for large-scale cell culture, storage, and distribution. At laboratory scales hESCs are cultured adherent to tissue culture plates; these culture techniques are labor-intensive and do not scale to high cell numbers. In an effort to facilitate larger scale hESC cultivation, we investigated the feasibility of culturing hESCs adherent to microcarriers. We modified the surface of Cytodex 3 microcarriers with either Matrigel or mouse embryonic fibroblasts (MEFs). hESC colonies were effectively expanded in a pluripotent, undifferentiated state on both Matrigel-coated microcarriers and microcarriers seeded with a MEF monolayer. While the hESC expansion rate on MEF-microcarriers was less than that on MEF-plates, the doubling time of hESCs on Matrigel-microcarriers was indistinguishable from that of hESCs expanded on Matrigel-coated tissue culture plates. Standard hESC cryopreservation methodologies are plagued by poor viability and high differentiation rates upon thawing. Here, we demonstrate that cryopreservation of hESCs adherent to microcarriers in cryovials provides a higher recovery of undifferentiated cells than cryopreservation of cells in suspension. Together, these results suggest that microcarrier-based stabilization and culture may facilitate hESC expansion and storage for research and therapeutic applications.
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克隆衍生的人类胚胎干细胞系在长期培养过程中保持多能性和增殖潜力
影响因子：
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