Cryopreservation of Human Midbrain Dopaminergic Neural Progenitor Cells Poised for Neuronal Differentiation.
Cryopreservation of Human Midbrain Dopaminergic Neural Progenitor Cells Poised for Neuronal Differentiation.
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DOI：
10.3389/fcell.2020.578907
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发表时间：
2020
影响因子：
5.5
通讯作者：
Kunath T
中科院分区：
文献类型：
--
作者：
Drummond NJ;Singh Dolt K;Canham MA;Kilbride P;Morris GJ;Kunath T
人多能干细胞可通过底板祖细胞期向中脑多巴胺能（mDA）神经元分化。利用底板方案产生的mDA神经元的发育特性与黑质神经元相似，这提高了在培养皿中模拟帕金森病（PD）的能力。结合多能干细胞的无限生长潜力，mDA神经祖细胞的生产可以为人类多巴胺能（DA）神经元的各种应用提供可扩展的来源。然而，由于方案的复杂性和长度以及细胞系之间的固有差异，经常观察到最终神经元群体的相当大的变异性。解决这一问题的一种方法是将mDA神经祖细胞冷冻保存为可随时使用的形式。建立一个冷冻保存的mDA神经祖细胞库，用于神经元分化，可以显著提高可重复性，促进合作。在这里，我们比较了六种不同的商业冷冻保存介质和不同的冷冻条件对人胚胎干细胞（hESC）分化的mDA神经祖细胞的影响。解冻后24 h细胞恢复有显著差异，但解冻后立即没有差异。ROCK抑制剂的存在改善了所有低温保存培养基24小时的细胞恢复。在所有测试条件下，更快的冷却速度（1-2°C/min）明显优于0.5°C/min，而在37°C下快速解冻并不总是优于在4°C下缓慢解冻。重要的是，mDA神经祖细胞的低温保存不会改变它们向mDA神经元恢复分化的潜力。低温保存的mDA神经祖细胞库提供了一种产生人类DA神经元的方法，减少了批次间的可变性，并建立了一种共享谱系启动细胞的机制，用于合作研究。
Human pluripotent stem cells can be differentiated into midbrain dopaminergic (mDA) neurons by directing cells through a floor plate progenitor stage. The developmental identity of mDA neurons produced using floor plate protocols is similar to substantia nigra neurons, and this has improved the ability to model Parkinson’s disease (PD) in a dish. Combined with the unlimited growth potential of pluripotent stem cells, mDA neural progenitor cell production can provide a scalable source of human dopaminergic (DA) neurons for diverse applications. However, due to the complexity and length of the protocols and inherent differences between cell lines, considerable variability of the final population of neurons is often observed. One solution to this problem is to cryopreserve committed mDA neural progenitor cells in a ready-to-use format. Creating a bank of cryopreserved mDA neural progenitor cells poised for neuronal differentiation could significantly improve reproducibility and facilitate collaborations. Here we have compared six (6) different commercial cryopreservation media and different freezing conditions for mDA neural progenitor cells differentiated from human embryonic stem cell (hESC) lines. Significant differences in cell recovery were observed at 24 h post-thawing, but no differences were observed immediately upon thawing. The presence of ROCK inhibitors improved cell recovery at 24 h for all cryopreservation media tested. A faster cooling rate of 1–2°C/min was significantly better than 0.5°C/min for all conditions tested, while rapid thawing at 37°C was not always superior to slow thawing at 4°C. Importantly, cryopreservation of mDA neural progenitor cells did not alter their potential to resume differentiation into mDA neurons. Banks of cryopreserved committed mDA neural progenitor cells provide a method to generate human DA neurons with reduced batch-to-batch variability, and establish a mechanism to share lineage-primed cells for collaborative research.
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影响因子：
64.8
作者：
Kriks, Sonja;Shim, Jae-Won;Piao, Jinghua;Ganat, Yosif M.;Wakeman, Dustin R.;Xie, Zhong;Carrillo-Reid, Luis;Auyeung, Gordon;Antonacci, Chris;Buch, Amanda;Yang, Lichuan;Beal, M. Flint;Surmeier, D. James;Kordower, Jeffrey H.;Tabar, Viviane;Studer, Lorenz
通讯作者：
Studer, Lorenz
影响因子：
2.9
作者：
Kim, Yoon Young;Ku, Seung-Yup;Choi, Young Min
通讯作者：
Choi, Young Min
预测标记指导分化以改善基于hESC的帕金森氏病疗法的临床翻译中的移植结果。
影响因子：
23.9
作者：
Kirkeby A;Nolbrant S;Tiklova K;Heuer A;Kee N;Cardoso T;Ottosson DR;Lelos MJ;Rifes P;Dunnett SB;Grealish S;Perlmann T;Parmar M
通讯作者：
Parmar M
影响因子：
6.2
作者：
Bonilla, Sonia;Hall, Anita C.;Arenas, Ernest
通讯作者：
Arenas, Ernest
影响因子：
23.9
作者：
Fasano CA;Chambers SM;Lee G;Tomishima MJ;Studer L
通讯作者：
Studer L