Han Ying, Quan Guo Bo, Liu Xiu Zhen, Ma En Pu, Liu An, Jin Peng, Cao Wei
Institute of Transfusion Medicine, Beijing 100850, China.
Cryobiology. 2005 Oct;51(2):152-64. doi: 10.1016/j.cryobiol.2005.06.002.
The lyophilization of human red blood cells has important implications for blood transfusion in clinical medicine. In this study, sugars, human serum albumin, polyvinylpyrrolidone, and dimethyl sulfoxide were used as protective reagents for the lyophilization of red blood cells. Freezing temperature, shelf temperature, and the rehydration conditions were optimized. The results showed that extracellular disaccharides, especially trehalose, did not increase the recovery of hemoglobin. However, when the concentration of human serum albumin was higher than 25%, it had a considerable protective effect on the recovery of lyophilized red blood cells; the cellular hemoglobin recovery was over 70%, which was significantly higher than that in the group without human serum albumin (P<0.01). As the concentration of polyvinylpyrrolidone was increased, the extent of vitrification also increased. But when the concentration of polyvinylpyrrolidone was over 40%, the resulting concentration of free hemoglobin was over 1g/L, which was significantly higher than that with 40% (P<0.01). When lyophilization was carried out after freezing at different temperatures, the recovery of cells and hemoglobin was 70-80% and there were no significant differences among the five groups. When the shelf temperature was higher than -30 degrees C, the samples were partly collapsed, but when the shelf temperature was lower than -30 degrees C, the recovery of cells in the -40 and -45 degrees C groups was significantly higher than in the -30 and -35 degrees C groups (P<0.05). The recovery of cells and hemoglobin after lyophilization and rehydration in solutions containing low concentrations of polymers was over 80%, which is significantly higher than the other groups (P<0.01). In addition, when the temperature was higher than 25 degrees C, the concentration of free hemoglobin was significantly lower than it was at 4 degrees C (P<0.01). In conclusion, our study showed the lyophilization of red blood cells is feasible. Disaccharides have no protective effect on lyophilized cells when they are only extracellular and extensive vitrification may be not beneficial. Although the recovery of cells after lyophilization and rehydration by our method was over 70%, the ultrastructure of the cells may be compromised and some hemolysis does still exist. Further research is required.
人红细胞的冻干对临床医学中的输血具有重要意义。在本研究中，糖、人血清白蛋白、聚乙烯吡咯烷酮和二甲亚砜被用作红细胞冻干的保护剂。对冷冻温度、保存温度和复水条件进行了优化。结果表明，细胞外二糖，尤其是海藻糖，并未提高血红蛋白的回收率。然而，当人血清白蛋白浓度高于25%时，它对冻干红细胞的回收率具有显著的保护作用；细胞血红蛋白回收率超过70%，显著高于无人血清白蛋白组（P<0.01）。随着聚乙烯吡咯烷酮浓度的增加，玻璃化程度也增加。但当聚乙烯吡咯烷酮浓度超过40%时，游离血红蛋白浓度超过1g/L，显著高于40%时的浓度（P<0.01）。在不同温度下冷冻后进行冻干时，细胞和血红蛋白的回收率为70 - 80%，五组之间无显著差异。当保存温度高于 - 30℃时，样品部分塌陷，但当保存温度低于 - 30℃时， - 40℃和 - 45℃组的细胞回收率显著高于 - 30℃和 - 35℃组（P<0.05）。在含有低浓度聚合物的溶液中冻干并复水后，细胞和血红蛋白的回收率超过80%，显著高于其他组（P<0.01）。此外，当温度高于25℃时，游离血红蛋白浓度显著低于4℃时的浓度（P<0.01）。总之，我们的研究表明红细胞冻干是可行的。仅细胞外存在的二糖对冻干细胞无保护作用，广泛的玻璃化可能并无益处。虽然通过我们的方法冻干并复水后的细胞回收率超过70%，但细胞的超微结构可能受损，仍存在一定程度的溶血。需要进一步研究。
