Background: The ability to preserve living cells or stem cells is critical for their use in cell therapy, especially for regenerative, reproductive, and transfusion medicine. This article addresses the low survival rates of cells and their loss of function during traditional freezing and banking (cells in a liquid medium with cryoprotectants). Aim: In this article, we developed multiple emulsions (water-in-oil-in-water type) for the effective encapsulation and cryopreservation of cells. In multiple emulsions, the oil drops, acting as a protective membrane, contain even smaller water droplets with encapsulated living cells, dispersed in the continuous water phase. Materials and Methods: The multiple emulsions with HEK293 cells encapsulated in the internal alginate droplets were successfully prepared in a Couette-Taylor flow biocontactor. The cryoprotectants (sucrose/dimethyl sulfoxide-DMSO) were located within the internal or external or both water phases of the emulsions. Encapsulated and non-encapsulated cells were frozen to -80°C (cooling rate: -1°C/min) and then transferred to liquid nitrogen (-196°C) for 24 hours. The standard rapid warming procedure was applied to thaw samples. Cell proliferation and viability were measured by using the AlamarBlue™ assay after recovery of cells. Results: The results showed that the viability of cells encapsulated in the internal droplets of multiple emulsions, and then cryopreserved, was significantly higher, up to 27.9%, than that observed for cells conventionally cryopreserved (non-encapsulated cells in water). Moreover, the effective cell-loaded multiple emulsions-based banking method allowed DMSO-toxic cryoprotectant-to be eliminated from the cryopreservation system. Conclusion: The proposed approach of the cryoprotection of cells encapsulated in multiple emulsions could minimize cell damage, degradation, and their loss during freezing-thawing processes.

中文翻译：

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有效冷冻保存和回收包裹在多种乳液中的活细胞。

背景：保存活细胞或干细胞的能力对于其在细胞治疗中的应用至关重要，特别是在再生，生殖和输血医学中。本文讨论了在传统的冷冻和储存过程中（具有冷冻保护剂的液体培养基中的细胞）细胞存活率低以及功能丧失的问题。目的：在本文中，我们开发了多种乳剂（水包油型水）有效地封装和冷冻细胞。在多种乳液中，油滴起着保护膜的作用，甚至包含了更小的水滴，这些水滴具有包裹的活细胞，分散在连续的水相中。材料和方法：在Couette-Taylor流式生物接触器中成功制备了具有包裹在内部藻酸盐液滴中的HEK293细胞的多重乳剂。防冻剂（蔗糖/二甲基亚砜-DMSO）位于乳液的内部或外部或两个水相内。将封装和未封装的细胞冷冻至-80°C（冷却速率：-1°C / min），然后转移至液氮（-196°C）中24小时。将标准快速加热程序应用于解冻样品。细胞恢复后，通过使用AlamarBlue TM测定法测量细胞增殖和活力。结果：结果表明，包裹在多种乳剂内部液滴中，然后冷冻保存的细胞的活力显着高于常规冷冻保存的细胞（水中未包裹的细胞），最高可达27.9％。此外，有效的基于细胞的多重乳液基银行方法使DMSO有毒的冷冻保护剂从冷冻保存系统中消除。结论：所提出的冷冻保护多种乳液中的细胞的方法可以最大程度地减少细胞在冻融过程中的损伤，降解及其损失。