Mazur P, Rigopoulos N
Cryobiology. 1983 Jun;20(3):274-89. doi: 10.1016/0011-2240(83)90016-0.
The general belief is that slow freezing injury is either the result of exposure to high salt concentrations or the result of excessive cell shrinkage. Increased salt concentration arises as increasing amounts of pure ice precipitate out of solution during freezing and cause the liquid-filled channels in which the cells are sequestered to dwindle in size. Cell shrinkage is an osmotic response to the concentration of external solutes. The consensus has been that the injury is related to the composition of the solution in these channels and not to the amount of residual liquid. Ordinarily, salt concentration and the amount of liquid in the unfrozen channels are reciprocally related; but they can be separated within limits by varying the total concentration of solutes in the suspending medium while holding the mass ratio of additive to salt constant, and by then slowly freezing samples to various subzero temperatures, chosen to produce various molalities of salt, while holding the unfrozen fraction constant, or vice versa. We have recently reported (9) that when human red cells are frozen under these conditions and thawed rapidly, survival is more dependent on the unfrozen water fraction than it is on the salt concentration in that fraction. The present work compares these results with those obtained with slow thawing. While the general conclusion remains unaltered, slowly thawed cells were able to survive the freezing of a higher fraction of extracellular water than were rapidly thawed cells. Calculations were made of the changes in cell volume during the equilibration with glycerol and the subsequent freezing involved in these experiments. Cell size and cell solute concentration were found to be independent of the fraction of unfrozen extracellular water, but cell survival was strongly dependent on that fraction. If applicable to other than human red cells, this finding is likely to require major modifications in current views of slow-freezing injury and its prevention.
一般认为，慢速冷冻损伤要么是暴露于高盐浓度的结果，要么是细胞过度收缩的结果。冷冻过程中，随着越来越多的纯冰从溶液中析出，盐浓度会升高，这会导致细胞所处的充满液体的通道尺寸减小。细胞收缩是对外部溶质浓度的一种渗透反应。人们一直认为，损伤与这些通道中溶液的成分有关，而与残留液体的量无关。通常情况下，盐浓度与未冻结通道中的液体量呈反比关系；但通过在保持添加剂与盐的质量比不变的情况下改变悬浮介质中溶质的总浓度，然后将样品缓慢冷冻至不同的零下温度（选择这些温度以产生不同摩尔浓度的盐），同时保持未冻结部分恒定，或者反之，就可以在一定限度内将它们分开。我们最近报道（9），当人类红细胞在这些条件下冷冻并快速解冻时，细胞存活率更多地取决于未冻结水的比例，而不是该部分中的盐浓度。本研究将这些结果与慢速解冻的结果进行了比较。虽然总体结论不变，但与快速解冻的细胞相比，慢速解冻的细胞能够在更高比例的细胞外水被冻结的情况下存活。对这些实验中甘油平衡过程中以及随后冷冻过程中的细胞体积变化进行了计算。发现细胞大小和细胞溶质浓度与未冻结细胞外水的比例无关，但细胞存活率强烈依赖于该比例。如果这一发现适用于人类红细胞以外的其他细胞，那么可能需要对当前关于慢速冷冻损伤及其预防的观点进行重大修改。
