Mazur P, Cole K W
Cryobiology. 1985 Dec;22(6):509-36. doi: 10.1016/0011-2240(85)90029-x.
We have shown previously that the survival of human red blood cells during slow freezing at 2% hematocrit is dependent more on the magnitude of the unfrozen fraction than on the salt concentration in that unfrozen fraction. In parallel, first Nei and more recently Pegg and colleagues have shown that survival is affected by the hematocrit of the suspension. Freezing at hematocrits above 30% becomes increasingly damaging. The present studies were designed to see whether there is a link between the two phenomena. Cells were suspended at nominal hematocrits of 0.4, 2, 8, 40, or 60% in five test solutions of glycerol-NaCl. The test solutions were of such composition that when frozen to a specified temperature, the magnitude of the unfrozen fraction differed but the NaCl concentration (ms) remained constant. At low hematocrits (0.4 to 8%), red cell survival was dependent predominantly on the unfrozen fraction and was relatively independent of the salt concentration in that fraction. This we term the "rheological" effect because injury appears to be related to interaction with the ice walls and perhaps is due to shearing forces or cell deformation. But at high hematocrits (40 or 60%), cell survival became dependent on both the unfrozen fraction and the salt concentration in that fraction. When freezing occurs at high hematocrits, increasing numbers of cells are presumably brought into contact with their neighbors. Furthermore, they are increasingly shrunken cells, for the progressive removal of liquid water, which is responsible for the crowding, also causes a rise in ms and the consequent osmotic shrinkage of cells. Our data suggest that at unfrozen fractions above those producing injurious rheological forces, the tight packing of less shrunken cells (i.e., high hematocrit, low ms) and the extensive shrinking of loosely packed cells (high ms, low hematocrit) are both quite innocuous. Injury becomes substantial only when extensively shrunken cells are brought into close contact (i.e., high ms, high hematocrit). At high hematocrit the cells occupy a substantial fraction of the unfrozen space, and the water that they lose during slow freezing adds substantially to the volume of extracellular ice. Accordingly, we defined other measures of unfrozen fraction that include these perturbations. However, we found that the conclusions on the relation between survival, unfrozen fraction, and hematocrit were not affected by the method of expressing the unfrozen fraction. Freezing at high hematocrit to high ms and low values of unfrozen fraction is one way to produce contact between shrunken cells at low temperatures.(ABSTRACT TRUNCATED AT 400 WORDS)
我们之前已经表明，在2%血细胞比容下缓慢冷冻时，人类红细胞的存活更多地取决于未冻部分的大小，而非该未冻部分中的盐浓度。与此同时，内井首先，以及最近佩格及其同事都表明，存活受到悬浮液血细胞比容的影响。血细胞比容高于30%时冷冻造成的损伤越来越大。本研究旨在探究这两种现象之间是否存在联系。将细胞悬浮于甘油 - 氯化钠的五种测试溶液中，标称血细胞比容分别为0.4%、2%、8%、40%或60%。测试溶液的组成使得当冷冻至特定温度时，未冻部分的大小不同，但氯化钠浓度（ms）保持恒定。在低血细胞比容（0.4%至8%）时，红细胞存活主要取决于未冻部分，且相对独立于该部分中的盐浓度。我们将此称为“流变学”效应，因为损伤似乎与与冰壁的相互作用有关，可能是由于剪切力或细胞变形。但在高血细胞比容（40%或60%）时，细胞存活变得既取决于未冻部分，也取决于该部分中的盐浓度。当在高血细胞比容下冷冻时，越来越多的细胞可能会与相邻细胞接触。此外，它们是越来越萎缩的细胞，因为导致拥挤的液态水的逐渐去除也会导致ms升高，从而使细胞发生渗透性萎缩。我们的数据表明，在高于产生有害流变学力的未冻部分时，较少萎缩细胞的紧密堆积（即高血细胞比容，低ms）和松散堆积细胞的广泛萎缩（高ms，低血细胞比容）都是相当无害的。只有当广泛萎缩的细胞紧密接触时（即高ms，高血细胞比容），损伤才会变得严重。在高血细胞比容下，细胞占据了相当一部分未冻空间，并且它们在缓慢冷冻过程中失去的水分会显著增加细胞外冰的体积。因此，我们定义了其他未冻部分的测量方法，其中包括这些扰动因素。然而，我们发现关于存活、未冻部分和血细胞比容之间关系的结论不受表示未冻部分方法的影响。在高血细胞比容下冷冻至高ms和低未冻部分值是在低温下使萎缩细胞接触的一种方法。（摘要截断于400字）
