二甲基亚砜联合羟乙基淀粉的非程控降温法低温冻存脐血来源不成熟树突状细胞的实验研究  

Non-programmed processing cryopreservation of immature dendritic cells derived from cord blood with dimethyl sulfoxide and hydroxyethyl starch

作　　者：王逸涛[1] 彭毅志[1] 王强[1] 王永权[1] 游波[1] 

机构地区：[1]第三军医大学附属西南医院全军烧伤研究所创伤烧伤与复合伤国家重点实验室,重庆400038

出　　处：《重庆医科大学学报》2006年第2期156-158,174,共4页Journal of Chongqing Medical University

基　　金：国家自然科学基金资助项目(3027134)

摘　　要：目的:联合使用二甲基亚砜(dimethyl sulfoxide,DMSO)和羟乙基淀粉(hydroxyethyl starch,HES)作为冷冻保护剂并使用非程控降温冷冻技术,研究脐血来源的不成熟树突状细胞低温冻存前后所表现的生物特性,为不成熟树突状细胞的临床应用提供保存方法。方法:GM-CSF(granulocyte-macrophage clonoy-stimulatingfactor)和IL-4(interleukin-4)体外诱导单个核细胞产生不成熟树突状细胞,5%DMSO+6%HES作为保护剂,-80℃冰箱降温,-196℃保存,37～40℃水浴复温,检测方法:台盼蓝拒染回收率,观察细胞形态,鉴定免疫表型,进行混合淋巴细胞反应,观察其诱导未致敏T淋巴细胞增殖情况。结果:冻存不成熟树突状细胞的台盼蓝拒染回收率为88.6%,其细胞形态,免疫表型及刺激未致敏T淋巴细胞的能力与新鲜DC相比无显著性差异。结论:冻存不成熟树突状细胞具有DC的显著特征,其细胞免疫表型和细胞功能实验上仍然保持了不成熟特征,联合使用二甲基亚砜和羟乙基淀粉作为冷冻保护剂对不成熟树突状细胞有良好的保护作用。Objective： This experiment investigated the biological properties of immature and cryopreserved DCs to develope a preservative method of immature dendritic cells for clinical use. Method： Immature dendritic cells were generated from human cord blood （CB） monocyte c μ ltured with rhGM-CSF and rhIL-4 and were cryopreservated with 5% DMSO and 6% HES in -80℃ refrigerator, then into -196℃ liquid nitrogen. We revived the cells into 37- 40℃ water bath after a month. Recovery rate was evaluated by TBBR. imDCs＇ morphological changes of DCs were analyzed by expression of surface antigens and allgenetic mixed lymphocyte reaction was performed to assess aUo-stim μ lating abilities of imDCs. Result： TBBR was 88.6%. The freeze-thawed immature dendritic cells＇ immunophenotype and function did not changed comparing with their fresh counterparts （P〉0.05）. Conclusion： Cryopreserved DCs exhibited typical characteristics of DCs, immature in cell phenotype and cell functions, suggesting that the non-programmed process using the combined two protectants is feasible.

关 键 词：脐血 树突状细胞 低温保存 二甲基亚砜 羟乙基淀粉. 

分 类 号：Q813.1[生物学—生物工程]