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目的通过在红细胞深低温冰冻的冻存液中添加渗透压调节物-四氢嘧啶(ectoine),检测红细胞冰冻前后的质量,探讨ectoine用于深低温冰冻红细胞的可能性,为ectoine可用于红细胞深低温冰冻的研究提供实验依据。方法取悬浮红细胞样品8袋,每袋分为对照组(复方甘油组、甘油组)和实验组(1.5%ectoine甘油组、3%ectoine甘油组和4.5%ectoine甘油组)共5组。实验组:ectoine起始浓度为1.5%、3%与4.5%(W/V)(加红细胞后终浓度为1.05%、2.10%与3.15%)。将5组(2组对照组与3组实验组)冻存液分别加入红细胞,红细胞冰冻后解冻并洗涤。测定冰冻前(未加冻存液)及解冻洗涤后红细胞数、血红蛋白量、红细胞变形性、渗透脆性,并用电镜扫描,观察红细胞显微形态。结果与2组对照组相比,3%ectoine甘油组红细胞回收率最高(89.26±2.60)%(P<0.05);3%ectoine甘油组解冻洗涤的红细胞与冰冻前红细胞,在50~(-1)、100~(-1)、200~(-1)和1 000~(-1)的剪切率值上相比P>0.05,差异无统计学意义;1.5%ectoine甘油组解冻洗涤的红细胞,50%溶血率时氯化钠浓度为(4.57±0.08)g/L,3%ectoine甘油组(4.80±0.17)g/L,2组分别与复方甘油组(4.98±0.26)g/L相比P<0.05,差异具有统计学意义;电镜扫描显示3组实验组红细胞与复方甘油组红细胞解冻洗涤后,均无棘状和球形红细胞,甘油组可见破膜、皱缩的红细胞。结论3%Ectoine作为渗透压调节物用于冰冻红细胞,可用于高浓度甘油冰冻红细胞时作为渗透压调节物,相对于3%乳酸钠的复方甘油(商品化复方甘油),可提高解冻洗涤的红细胞回收率,改善解冻洗涤的红细胞质量,具有在深低温冰冻红细胞的潜力。
OBJECTIVE: To investigate the potential of ectoine for cryogenic freezing of erythrocytes by adding ectoine, an osmoregulation regulator, to the cryopreservation of erythrocytes. The ectoine can be used in deep red blood cells Low temperature freezing provides experimental evidence. Methods Eight bags of suspended erythrocytes were collected and divided into 5 groups: control group (glycerin group) and experimental group (1.5% ectoine glycerol group, 3% ectoine glycerol group and 4.5% ectoine glycerol group). In the experimental group, the starting concentrations of ectoine were 1.5%, 3% and 4.5% (W / V) (final concentrations 1.05%, 2.10% and 3.15%, respectively). Five groups (two control groups and three experimental groups) were added into the red blood cells frozen, red blood cells were thawed and washed after freezing. The number of erythrocytes, hemoglobin, erythrocyte deformability and osmotic fragility were measured before freezing (without frozen solution) and after thaw washing, and the microscopic morphology of erythrocytes was observed by scanning electron microscope. Results Compared with the two control groups, the recovery rate of erythrocytes in 3% ectoine glycerol group was the highest (89.26 ± 2.60)% (P <0.05). In the 3% ectoine glycerol group, the washed erythrocytes and pre- ), 100 ~ (-1), 200 ~ (-1) and 1 000 ~ (-1), P> 0.05, the difference was not statistically significant; 1.5% ectoine glycerol group thawed washed red blood cells (4.57 ± 0.08) g / L and 3% ectoine glycerol group (4.80 ± 0.17) g / L at 50% hemolysis rate, respectively. Compared with the compound glycerol group (4.98 ± 0.26) g / L Compared with P <0.05, the difference was statistically significant. Electron microscopy showed that erythrocytes and glycated glycerol group did not have spiny and spherical erythrocytes after being thawed in the erythrocytes and glycated glycerol groups, and ruptured and shrunken erythrocytes were seen in the glycerin group. Conclusions 3% Ectoine is used as an osmoregulator for frozen red blood cells. It can be used as a regulator of osmotic pressure when high concentration of glycerol is used to freeze erythrocytes. Compared with 3% sodium lactate compound glycerin (commercial compound glycerin), Ectoine can improve the recovery of thawed red blood cells Rate, improve the quality of thawed washed red blood cells, with the potential to freeze red blood cells at cryogenic temperatures.