Quan GuoBo, Zhang Liang, Guo Yong, Liu MinXia, Wang JieXi, Wang Yan, Dong Bo, Liu An, Zhang JinGang, Han Ying
Institute of Transfusion Medicine, Beijing, China.
Cryo Letters. 2007 Mar-Apr;28(2):95-108.
Cryopreservation with impermeable protectants has great significance on storage of human red blood cells. It has become feasible to use glycerol free cryopreservation for human red blood cells. This study focuses on the effect of intracellular trehalose or glucose on human red blood cells cryopreserved in the presence of polymer. Red blood cells were cryopreserved for 48 h-72 h at -80 degrees C. The data showed that the loading efficiency of glucose was significantly higher than that of trehalose, but trehalose loading process induced more hemolysis than glucose loading process. Compared with the other groups, the combination of intracellular glucose, PVP, and human serum albumin can significantly decrease the percent hemolysis after cryopreservation (P<0.01). However, the percent hemolysis induced by intracellular trehalose was less than that induced by extracellular trehalose, but the difference was not significant (P<0.05). The adenosine 5'-triphosphate (ATP) level and 2,3-diphosphoglycerate (2,3-DPG) level of cryopreserved red blood cells were significantly less than those of fresh red blood cells. However, sugars can provide certain protection for ATP and 2, 3-DPG compared with red blood cells cryopreserved in the absence of sugars. The protection of glucose on the metabolic function was more than that of trehalose. Cryopreservation can increase the percentage of cells with exposed phosphatidylserine (PS), but the ability of trehalose to maintain PS normal distribution is higher than that of glucose. Furthermore, intracellular sugars can protect membrane integrity of cryopreserved red blood cells, although a small portion of cells appeared spherocytic or echinocytic shape. Finally, most membrane proteins of cryopreserved red blood cells were similar to the membrane proteins of fresh red blood cells, but trehalose can result in loss of glyceraldehyde phosphate dehydrogenase (GAPD) and peroxiredoxin 2. In conclusion, it is feasible to cryopreserve red blood cells using polymer, human albumin and sugars as main protectants. The cryoprotective effect of glucose may be better than that of trehalose in the presence of PVP and human serum albumin, because sugar loading process causes more cell injuries in case of trehalose compared to glucose, and these injuries in turn manifest themselves during subsequent cryopreservation and thawing. In the future, finding an approach to decrease the injuries during trehalose loading process still is critical.
使用非渗透性保护剂进行低温保存对人类红细胞的储存具有重要意义。对人类红细胞使用无甘油低温保存已变得可行。本研究聚焦于细胞内海藻糖或葡萄糖对在聚合物存在下低温保存的人类红细胞的影响。红细胞在-80℃下低温保存48小时至72小时。数据显示，葡萄糖的加载效率显著高于海藻糖，但海藻糖加载过程比葡萄糖加载过程诱导更多溶血。与其他组相比，细胞内葡萄糖、聚乙烯吡咯烷酮（PVP）和人血清白蛋白的组合可显著降低低温保存后的溶血百分比（P<0.01）。然而，细胞内海藻糖诱导的溶血百分比低于细胞外海藻糖诱导的溶血百分比，但差异不显著（P<0.05）。低温保存红细胞的三磷酸腺苷（ATP）水平和2,3-二磷酸甘油酸（2,3-DPG）水平显著低于新鲜红细胞。然而，与在无糖情况下低温保存的红细胞相比，糖类可为ATP和2,3-DPG提供一定保护。葡萄糖对代谢功能的保护作用大于海藻糖。低温保存可增加磷脂酰丝氨酸（PS）暴露细胞的百分比，但海藻糖维持PS正常分布的能力高于葡萄糖。此外，细胞内糖类可保护低温保存红细胞的膜完整性，尽管一小部分细胞呈现球形或棘形。最后，低温保存红细胞的大多数膜蛋白与新鲜红细胞的膜蛋白相似，但海藻糖可导致甘油醛-3-磷酸脱氢酶（GAPD）和过氧化物酶2的丢失。总之，使用聚合物、人白蛋白和糖类作为主要保护剂低温保存红细胞是可行的。在PVP和人血清白蛋白存在的情况下，葡萄糖的低温保护作用可能优于海藻糖，因为与葡萄糖相比，海藻糖加载过程会导致更多细胞损伤，而这些损伤在随后的低温保存和解冻过程中会表现出来。未来，找到一种减少海藻糖加载过程中损伤的方法仍然至关重要。
