Vitrification of encapsulated hepatocytes with reduced cooling and warming rates.
Vitrification of encapsulated hepatocytes with reduced cooling and warming rates.
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以降低的冷却和升温速率对封装的肝细胞进行玻璃化冷冻。
DOI：
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发表时间：
2004
期刊：
影响因子：
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通讯作者：
H. Yu
中科院分区：
文献类型：
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作者：
L. Kuleshova;X. Wang;Y. Wu;Y. Zhou;H. Yu
我们已经使用微囊化肝细胞作为模型，以开发一种玻璃体冷冻保存大量含细胞构建体的方法。该方法包括预平衡程序，其中渗透性冷冻保护剂的量在每个步骤中逐渐增加15%。最佳玻璃化液组成为40%乙二醇和0.6M蔗糖。用于第一次稀释溶液的1 M蔗糖浓度，随后将蔗糖浓度降低至0.7 M蔗糖，并在每个后续步骤中降低0.2- 0.15 M。该蔗糖稀释程序对细胞功能没有不利影响。三种冷却速率（400摄氏度/分钟及以上）和三种升温速率（650摄氏度/分钟及以上）与所提出的玻璃化溶液组合，同样有效。程序和溶液的优化允许微囊化肝细胞被保存，几乎100%保留细胞功能，并且对脆弱的微囊没有可检测的损伤。冷却/升温速率与玻璃化的有效性的分离潜在地为复杂组织工程构建体的大规模冷冻保存铺平了道路。
We have used microencapsulated hepatocytes as model to develop a method of vitreous cryopreservation of large quantities of cell-containing constructs. The method included a pre-equilibration procedure in which the amount of penetrating cryoprotectant was gradually increased by 15% in each step. The optimal vitrification solution consists of 40% ethylene glycol and 0.6M sucrose. The concentration of 1M sucrose used for the first dilution solution with subsequent decrease of sucrose concentration to 0.7 M sucrose and by 0.2-0.15M for each subsequent step. This sucrose dilution procedure had no adverse effect on cell functions. Three cooling rates (400 degrees C/min and above) and three warming rates (650 degrees C/min and above), in combination with the proposed vitrification solution, were equally effective. The optimization of the procedure and solutions allow microencapsulated hepatocytes to be preserved with almost 100% retention of cell functions and no detectable damage to the fragile microcapsules. The de-linking of the cooling/warming rates with the effectiveness of vitrification potentially paves the way for large scale cryopreservation of complex tissue engineered constructs.