Alginate Hydrogel Microencapsulation Inhibits Devitrification and Enables Large-Volume Low-CPA Cell Vitrification.
Alginate Hydrogel Microencapsulation Inhibits Devitrification and Enables Large-Volume Low-CPA Cell Vitrification.
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DOI：
10.1002/adfm.201503047
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发表时间：
2015-11-25
影响因子：
19
通讯作者：
He X
中科院分区：
文献类型：
--
作者：
Huang H;Choi JK;Rao W;Zhao S;Agarwal P;Zhao G;He X
干细胞的冷冻保存对于满足蓬勃发展的细胞医学日益增长的需求具有重要意义。传统的干细胞慢速冷冻方法存在着与结冰相关的不可避免的细胞损伤，而玻璃化(即看不见的冰形成)方法正在成为一种新的细胞冷冻保存策略。细胞玻璃化的一个主要挑战是在将玻璃化细胞在低温下加热到超低温时，由于去玻璃化(即在先前玻璃化的溶液中形成可见的冰)而导致的细胞内冰形成(IIF，对细胞的致命事件)。因此，在玻璃化过程中，使用了高浓度和有毒的穿透性冷冻保护剂(即高CPA，高达~8M)和/或有限的样品体积(高达~2.5CPA L)来最大限度地减少IIF。我们发现海藻酸水凝胶微胶囊化可以有效地抑制升温过程中的玻璃化。我们的数据表明，如果在冷却过程中最大限度地减少冰的形成，在玻璃化的整个冷却和升温过程中，藻酸盐水凝胶微胶囊细胞中的IIF可以忽略不计。这使得多能和多能干细胞玻璃化冷冻时，穿透性CPA浓度最低可达~4倍(高达2M，低CPA)，样品体积最大可达~100倍(高达~250μL，大体积)。
Cryopreservation of stem cells is important to meet their ever-increasing demand by the burgeoning cell-based medicine. The conventional slow freezing for stem cell cryopreservation suffers from inevitable cell injury associated with ice formation and the vitrification (i.e., no visible ice formation) approach is emerging as a new strategy for cell cryopreservation. A major challenge to cell vitrification is intracellular ice formation (IIF, a lethal event to cells) induced by devitrification (i.e., formation of visible ice in previously vitrified solution) during warming the vitrified cells at cryogenic temperature back to super-zero temperatures. Consequently, high and toxic concentrations of penetrating cryoprotectants (i.e., high CPAs, up to ~8 M) and/or limited sample volumes (up to ~2.5 μl) have been used to minimize IIF during vitrification. We reveal that alginate hydrogel microencapsulation can effectively inhibit devitrification during warming. Our data show that if ice formation were minimized during cooling, IIF is negligible in alginate hydrogel-microencapsulated cells during the entire cooling and warming procedure of vitrification. This enables vitrification of pluripotent and multipotent stem cells with up to ~4 times lower concentration of penetrating CPAs (up to 2 M, low CPA) in up to ~100 times larger sample volume (up to ~250 μl, large volume).