An effective serum- and xeno-free chemically defined freezing procedure for human embryonic and induced pluripotent stem cells.
An effective serum- and xeno-free chemically defined freezing procedure for human embryonic and induced pluripotent stem cells.
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DOI：
10.1093/humrep/deq040
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发表时间：
2010-05
期刊：
影响因子：
--
通讯作者：
Hovatta O
中科院分区：
文献类型：
--
作者：
Holm F;Ström S;Inzunza J;Baker D;Strömberg AM;Rozell B;Feki A;Bergström R;Hovatta O
人胚胎干细胞（human embryonic stem cells，hESC）和诱导多能干细胞（induced pluripotent stem cells，iPSCs）在再生医学中具有巨大的应用潜力。除了优化的临床级培养条件之外，还需要用于这些细胞的有效的临床级冷冻保存方法。解冻后获得良好的存活率一直是个问题。我们使用了一种新的化学定义的有效的无异种冷冻保存系统用于hESC和iPSC的冷冻储存和建库。早期建立的缓慢冷冻方案，即使在最近的改进，导致低活力和解冻的细胞有很高的分化倾向。该培养基是一种完全不含血清和动物物质的产品，含有二甲亚砜、无水葡萄糖和聚合物作为冷冻保护剂。细胞直接在-70 °C下冷冻，没有程序化冷冻机。HS 293（χ2 = 9.616，单自由度，双尾P = 0.0019）和HS 306（χ2 = 8.801，单自由度，双尾P = 0.0030）的冷冻集落数量与存活集落数量存在显著差异。解冻后，细胞具有高活力（90-96%），与活力低得多（49%）的标准冷冻程序相比，对增殖和分化没有任何影响。冻融后的hESC和iPSCs核型正常，保持了多能性细胞的特性，并具有相应的形态学特征，在培养10代后表达多能性标志物。它们形成含有三个胚层组织成分的畸胎瘤。这里描述的定义的冻融系统为hESC和iPSC的库提供了一个非常简单的选择。
Both human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) bear a great potential in regenerative medicine. In addition to optimized clinical grade culture conditions, efficient clinical grade cryopreservation methods for these cells are needed. Obtaining good survival after thawing has been problematic. We used a novel, chemically defined effective xeno-free cryopreservation system for cryostorage and banking of hESCs and iPSCs. The earlier established slow freezing protocols have, even after recent improvements, resulted in low viability and thawed cells had a high tendency to differentiate. The medium is a completely serum and animal substance free product containing dimethylsulfoxide, anhydrous dextrose and a polymer as cryoprotectants. The cells were directly frozen at −70°C, without a programmed freezer. The number of frozen colonies versus the number of surviving colonies differed significantly for both HS293 (χ2 = 9.616 with one degree of freedom and two-tailed P = 0.0019) and HS306 (χ2 = 8.801 with one degree of freedom and two-tailed P = 0.0030). After thawing, the cells had a high viability (90–96%) without any impact on proliferation and differentiation, compared with the standard freezing procedure where viability was much lower (49%). The frozen–thawed hESCs and iPSCs had normal karyotype and maintained properties of pluripotent cells with corresponding morphological characteristics, and expressed pluripotency markers after 10 passages in culture. They formed teratomas containing tissue components of the three germ layers. The defined freezing–thawing system described here offers an excellent simple option for banking of hESCs and iPSCs.
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影响因子：
5.2
作者：
Inzunza, J;Gertow, K;Hovatta, O
通讯作者：
Hovatta, O
人类胚胎干细胞的批量玻璃化冷冻。
影响因子：
6.1
作者：
Li, Tao;Zhou, Canquan;Zhuang, Guanglun
通讯作者：
Zhuang, Guanglun
影响因子：
56.9
作者：
Thomson, JA;Itskovitz-Eldor, J;Jones, JM
通讯作者：
Jones, JM
影响因子：
6.1
作者：
Reubinoff, BE;Pera, MF;Trounson, AO
通讯作者：
Trounson, AO
影响因子：
2.7
作者：
Wowk, B;Leitl, E;Fahy, GM
通讯作者：
Fahy, GM