细胞内外海藻糖对红细胞冰冻干燥保存的影响

目的研究细胞内海藻糖对红细胞冻干保存后血红蛋白回收率及ATP水平影响。在一定条件下负载红细胞,细胞内海藻糖浓度保持恒定,研究细胞外不同浓度的海藻糖对红细胞冰冻干燥保存的影响。方法将浓缩红细胞在37℃,浓度为800 mmol的海藻糖溶液中孵育7 h,制成海藻糖负载的浓缩红细胞,对照组为PBS负载浓缩红细胞,行冻干保存,测定Hb回收率及细胞内ATP水平。将PBS液、浓度为50 mmol、200 mmol、400 mmol的海藻糖溶液与海藻糖负载浓缩红细胞按1∶1比例混匀,行冻干保存及再水化,用氰化血红蛋白试剂盒测定Hb溶血率,计算Hb回收率。结果经海藻糖负载的红细胞冻干保存,Hb回收率(44.46±5.15)%,细胞内ATP水平(1.91±0.33)μmol/gHb,对照组Hb回收率(7.71±2.71)%,细胞内ATP水平(0.88±0.25)μmol/gHb。2组相比较,P0.05,差异有统计学意义。细胞外PBS液组Hb回收率为(10.36±0.97)%,50 mmol海藻糖组,Hb回收率为(33.57±2.89)%,200 mmol海藻糖组,Hb回收率为(38.64±0.54)%,400 mmol海藻糖组,Hb回收率为(18.10±1.9)%。对照组PBS液组与50 mmol组、200 mmol组、40 0mmol组分别比较,差异有统计学意义。400 mmol组与200 mmol组、5 0 mmol组分别比较,差异有统计学意义(P0.01)。200 mmol组与50 mmol组相比较,差异无统计学意义。结论细胞内的海藻糖大大提高冻干红细胞Hb回收率且可保持冻干红细胞正常ATP水平。细胞外的海藻糖对红细胞冻干保存有保护作用,随着细胞外液中海藻糖浓度增加,冻干红细胞Hb回收率减少。

Effect of intra/extra-cellular trehalose on the preservation of red blood cells by lyophilization

Objective To evaluate the effect of trehalose-loading of RBCs on the lyophilization and the effect of different extracellular trehalose concentration on the lyophilization of trehalose-loaded RBCs.Methods RBCs was incubated in 800mmol/L concentration of trehalose solution at 37℃ for7 hours,and RBCs incubated in phosphate-buffered saline were used as control.Trehalose loading red blood cells and solution of different extracellular trehalose concentration were mixed at ratio of 1∶1,Samples were lyophilizated,the recovery rate of hemoglobin was detected by using cyanohemeglobin kit.Intracellular ATP was determined by bioluminescence assay.Results The recovery rate of hemoglobin in trehalose-loading RBC was(44.46±5.15)%,the level of ATP(1.91±0.33)μmol/gHb,the recovery rate of hemoglobin in control was(7.71±2.71)%,the level of ATP(0.88±0.25)μmol/gHb.Comparision of recovery of hemoglobin and level ATP of lyphilizated RBC show significantly different for P<0.005.the recovery rate of hemoglobin in 50 mmol/L group was(33.57±2.89)%,in 200mmol/L group was(38.64±0.54)%,in 400mmol/L group was(18.10±1.95)%,while The recovery rate of hemoglobin in control group was(10.36±0.97)%.Comparision of recovery of hemoglobin of lyphilizateg RBC with solution of different extracellular trehalose cencontration,the data of recovery of hemoglobin of PBS(control group)show significantly different for P<0.001 with 50 mmol/L group,200mmol/L group,400mmol/L group.Comparision of recovery of hemoglobin of 400mmol/L group with 50mmol/L group,200mmol/L group,data show significantly different for P<0.001.50 mmol/L group,200mmol/L group,400mmol/L group were significantly higher than that of PBS group,50mmol/L group,200mmol/L group were significantly higher than that of 400mmol/L group.50mmol/L group was not significantly different with 200mmol/L group.Conclusion The recovery of hemoglobin and the level of ATP is improved in lyophilization of trehalose-loaded RBCs and the extracellular trehalose has protective effect,which decreased with increasing of extracellular trehalose concentrations beyond 200mmol/L.