Takahashi T, Bross J B, Shaber R E, Williams R J
Cryobiology. 1985 Aug;22(4):336-50. doi: 10.1016/0011-2240(85)90181-6.
High concentrations of membrane permeable cryoprotectants are necessary to protect human polymorphonuclear leukocytes from osmotic stress injury during freezing, but there are reports that some cryoprotectants are chemically toxic. Cells were exposed to various concentrations of glycerol, dimethyl sulfoxide, or ethylene glycol for 5 min to 2 hr at 37, 22 or 0 degree C, adding or removing the cryoprotectant either slowly or rapidly. Assays included cell number recovery, membrane integrity, phagocytosis, microbicidal ability, and chemotaxis. We conclude that (1) 1 and 2 M concentrations generally are not toxic if they are added and removed slowly at 22 degrees C; (2) addition and removal of glycerol at 0 degree C was injurious even at 1 M; (3) slow addition and removal allowed better recovery than rapid addition or removal; (4) salt concentration in cryoprotectant solutions should be adjusted to isotonic on the basis of moles per liter of solution, rather than moles per kilogram of water; (5) the toxicity reported by other investigators can be largely explained by osmotic stress or dilution shock rather than chemical toxicity; and (6) ethylene glycol is the easiest cryoprotectant to add to and remove from these cells.
为了在冷冻过程中保护人类多形核白细胞免受渗透应激损伤，需要高浓度的可透过细胞膜的冷冻保护剂，但有报道称某些冷冻保护剂具有化学毒性。将细胞在37、22或0摄氏度下暴露于不同浓度的甘油、二甲基亚砜或乙二醇中5分钟至2小时，缓慢或快速添加或去除冷冻保护剂。检测项目包括细胞数量恢复、膜完整性、吞噬作用、杀菌能力和趋化性。我们得出以下结论：（1）如果在22摄氏度下缓慢添加和去除，1M和2M的浓度通常无毒；（2）在0摄氏度下添加和去除甘油即使在1M时也有损伤；（3）缓慢添加和去除比快速添加或去除能实现更好的恢复；（4）冷冻保护剂溶液中的盐浓度应根据每升溶液的摩尔数而非每千克水的摩尔数调整为等渗；（5）其他研究者报道的毒性很大程度上可由渗透应激或稀释冲击而非化学毒性来解释；（6）乙二醇是向这些细胞添加和从这些细胞中去除最容易的冷冻保护剂。
