人脐血间充质干细胞对脐血CD_(34)^+细胞体外扩增作用的研究  

Effects of human umbilical cord blood mesenchymal stem cells on the expansion of CD_(34)^+ cells from umbilical cord blood

作　　者：周敦华[1] 黄绍良[1] 张绪超[1] 魏菁[1] 吴燕峰[1] 黄科[1] 黎阳[1] 方建培[1] 

机构地区：[1]中山大学附属第二医院儿科,广州510120

出　　处：《中华儿科杂志》2005年第7期494-498,共5页Chinese Journal of Pediatrics

基　　金：广东省自然科学基金资助项目(00137);广东省卫生厅基金资助项目(E200040);卫生部2004-2006年临床学科重点项目[2004(468)]

摘　　要：目的探讨含人脐血来源的间充质干细胞(MSCs)体系在体外对脐血造血干细胞(HSCs)扩增作用。方法(1)用含人脐血MSCs及不同造血生长因子(HGFs)组合的无血清扩增体系对人脐血CD+34细胞进行体外扩增。(2)于扩增前及扩增后第6、12天分别用双色流式细胞仪动态检测HSCs表面抗原标记:CD+34、CD+34CD-38、CD+34CD+3、CD+34CD+19、CD+34CD+33和CD+34CD+41a细胞的含量。(3)按本实验室方法行体外半固体培养,观察扩增前后脐血细胞粒单核细胞集落形成单位(CFUGM)、爆式红系集落形成单位(BFUE)、混合集落形成单位(CFUMix)及高增殖集落形成单位(CFUHPP)集落形成情况。结果(1)含人脐血MSCs体系对脐血CD+34CD38细胞的扩增倍数在第6天和第12天分别为159和437倍。该亚群百分比在单纯因子组扩增第12天时为1.98%,而在含脐血MSCs组为9.98%,明显高于扩增前。(2)集落培养表明,含脐血MSCs组扩增第12天与扩增第6天相比,其CFUMix和CFUHPP的扩增倍数增加,而单纯因子组这两种集落的扩增倍数下降。(3)随扩增天数的增加,两组扩增体系中CD+34CD+3和CD+34CD+41a细胞均明显增加,而CD+34CD+19和CD+34CD+3细胞均明显减少。两组相比,含脐血MSCs组差异更显著。结论(1)含脐血MSCs体系不仅能扩增更原始的造血干/祖细胞(HSPC),且具有在短期内(12d)保持HSCs不耗竭。(2)含脐血MSCs体系对脐血CD+34细胞向定向祖细胞的扩增,主要为髓系及巨核系祖细胞,而对其向淋巴系祖细胞的扩增具有抑制作用。Objective The previous studies indicated that mesenchymal stem cells (MSCs) either from umbilical cord blood (UCB) or from bone marrow (BM) had the same biological characteristics and the function of secreting hematopoietic growth factors (HGFs). The present study aimed to understand the effects of human UCB MSCs on the expansion of CD^+_(34) cells from UCB. Methods 1. Human UCB CD^+_(34) cells were incubated in the system containing UCB MSCs, HGFs and serum free medium. 2. The surface markers (CD^+_(34), CD^+_(34)CD^-_(38), CD^+_(34)CD^+_3, CD^+_(34)CD^+_(19), CD^+_(34)CD^+_(33), CD^+_(34)CD^+_(41a)) on expanded UCB cells were examined by flow cytometry on the 6^(th) and 12^(th) days. 3. The expanded and unexpanded cells were cultured in semi-solid culturing system and checked for colony forming units of granulocyte and macrophage (CFU-GM), erythroid burst-forming unit (BFU-E), colony forming units of granulocyte- erythrocyte-megakaryocyte-macrophage (CFU-Mix) and colony forming units of high-proliferative potential (CFU-HPP). Results 1.The expansion folds of CD^+_(34)CD^-_(38) cells from UCB MSCs+HGFs groups on the 6^(th) and 12^(th) days were 159.43 and 436.68, respectively. Interestingly, the percentage of CD^+_(34)CD^-_(38) cells declined in HGFs group after expanding for 12 days, but it rose to 9.98% in the UCB MSCs+HGFs group. 2. Colony forming capacity of expanded UCB cells showed that the folds of CFU-Mix and CFU-HPP of UCB MSCs+HGFs group increased from day 6 to day 12, but the folds decreased in the HGFs group. 3. From day 0 to day 12, CD^+_(34)CD^+_(33) cells and CD^+_(34)CD^+_(41a) cells were amplified gradually, but CD^+_(34)CD^+_(19) and CD^+_(34)CD^+_3 cells decreased gradually, and in UCB MSCs+HGFs group this phenomenon was more significant than that in HGFs group. Conclusion 1. UCB MSCs containing system not only has the ability to expand the primitive HSCs but also has the ability to sustain the proliferation of HSCs. 2. UCB MSCs containing system amplified mainly myeloid and megakaryocytoid pr

关 键 词：脐血CD34^+细胞 间充质干细胞 人脐血 体外扩增作用 CD34^+CD38^+细胞 红系集落形成单位 MSCs 脐血造血干细胞 双色流式细胞仪 CD3^+细胞 造血干/祖细胞 造血生长因子 粒-单核细胞 巨核系祖细胞 HSCs 半固体培养 实验室方法 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]