Lessons from Nature: Antifreeze Glycolipids for Cryopreservation of Mammalian Cel

描述(申请人提供)：对于许多研究人员来说，细胞的存储只是在悬浮的细胞中添加10%的DMSO或类似的低温保护剂，将它们放在冷冻箱中，缓慢冷却到零下温度，然后储存在机械或氮气冷却的冰箱或杜瓦瓶中。只要解冻后存在活细胞，由于细胞的高增殖潜力，细胞产量可能是次要的考虑因素。然而，有一些重要的细胞类型很难用现有的冷冻保存技术来保存，包括许多原代细胞，如心肌细胞、肝细胞，以及胚胎干细胞和可诱导的多能干细胞。其他的，如角质形成细胞，很容易在悬浮液中超低温保存，但在附着在基质上时很难保存。对生物如何在极端温度下生存的研究表明，为了促进生存，可能会同时发生多种现象，包括产生低温保护剂、防冻剂(多肽和脂类)、脱水和成核剂。通过模仿某些昆虫在零度以下生存的策略，哺乳动物细胞的冷冻保存可能是可能的。这些昆虫在寒冷的环境条件下，结合低温保护剂(如甘油)含量的增加和防冻剂化合物的产生，能够在低至-80摄氏度的温度下生存。这些化合物的存在降低了溶液的冰点，也改变了冰的形状和形成。通过这种方式，潜在较少的冷冻保护剂，如DMSO或甘油，可以用于保存细胞，减少潜在的冷冻保护剂引起的细胞毒性问题，并为细胞和组织提供替代的保存方法，可用于临床应用和研究应用的细胞和组织筛选分析。

DESCRIPTION (provided by applicant): Storage of cells for many researchers simply involves addition of 10% DMSO or similar cryoprotectants to cells in suspension, putting them in cryovials and slow cooling to subzero temperaturesf and storage in a mechanical or nitrogen cooled freezer or dewar. As long as viable cells are present upon thawing, cell yield may be a secondary consideration due to the high proliferative potential of the cells. However, there are significant cell types that are difficult to preserve with existing cryopreservation techniques including many primary cells, such as cardiac myocytes, and hepatocytes, plus embryonic stem cells and inducible pluripotent stem cells. Others, such as keratinocytes, are easy to cryopreserve in suspension but difficult when attached to a substrate. The study of how organisms survive extreme temperatures has revealed that multiple phenomena may occur in synchrony to promote survival including production of cryoprotectants, antifreeze compounds (peptides and lipids), dehydration and nucleators. Mammalian cell cryopreservation may be possible by mimicking strategies employed by certain insects to survive sub-zero temperatures. These insects combine increases in cryoprotectant content (such as glycerol) with production of antifreeze compounds, at the onset of cold environmental conditions, to survive temperatures as low as -80oC. The presence of these compounds lowers the freezing point of the solution and also changes the shape and formation of ice. In this way, potentially less cryoprotectants, such as DMSO or glycerol, may be used to preserve cells reducing potential cryoprotectant induced cytotoxicity concerns and offering alternative preservation methods for cells and tissues that can be used in clinical applications and cell and tissue-based screening assays for research applications.