脐血CD34^+细胞及红系祖细胞扩增的实验研究  

Expansion of Erythroid Progenitors and CD34+ Cells by Umbilical Cord Blood Mononuclear Cells

作　　者：张洁霞[1] 毛平[2] 

机构地区：[1]广州医学院第一附属医院肿瘤血液中心 [2]广州市第一人民医院血液内科,广州510180

出　　处：《中国实验血液学杂志》2005年第3期429-433,共5页Journal of Experimental Hematology

基　　金：广州市科学技术局科技攻关重点项目"脐血造血干细胞移植在血液病中的应用系列研究"基金资助项目;编号:2002zZ-E0241

摘　　要：脐血是造血祖细胞的丰富来源之一,选择合适的培养条件,体外诱导其定向扩增为红系祖细胞,输入体内产生成熟红细胞。本实验旨在探讨脐血单个核细胞(MNC)体外红系定向扩增的理想因子组合(Flt3配基FL联合TPO、SCF、EPO及FL、SCF、TPO)对CD34+细胞扩增的影响。将单个核细胞接种至stemspan无血清培养液中,共分3组:A组为对照组,B组为TPO+SCF+FL+EPO+IGF1组,C组为TPO+SCF+FL组,C组在第6天及以后换液加入EPO和IGF1。于培养0、6、10、14天进行细胞计数,细胞集落测定,流式细胞术测定细胞的CD34、CD34CD71、CD71GPA细胞的比例。结果表明:经10天培养后,B组总细胞数扩增6.89倍,而C组3.06倍;B组CD34+细胞增加4.83,而C组2.47倍;B组集落形成细胞数增加4.3倍,而C组增加2.5倍;B组红系祖细胞BFUE和CFUE数增加5.4倍,而C组3.1倍;B组CD34+CD71+细胞数增加8.72倍,而C组3.37倍;B组CD71+GPA+细胞数增加53.4倍,而C组30.29倍。结论:脐血MNC在无血清培养液中加入FL+SCF+TPO实现了CD34+细胞及集落形成细胞的扩增。脐血MNC在无血清培养液中加入FL+SCF+TPO+EPO+IGF1短期液体培养获得红系祖细胞的扩增,在第0天比6天加入EPO获得更多红祖细胞(P<0.05)。由于TPO+SCF+FL+EPO+IGF1组的集落形成细胞数、CFUE和BFUE数于第10天最多。Cord blood represents a large source of hematopoietic stem/progenitor cells. It can be induced to proliferate directly into erythroid progenitors in a appropriate ex vivo culture condition, then to generate mature red blood cells after injection into the body. The combination of Flt3 ligand,TPO,SCF and EPO is ideal for cord blood MNC to proliferate into erythroid progenitors. This study was aimed to evaluate the effect of FL, SCF and TPO on CD34+ expansion. and to investigate influence of the cytokine combination on the proliferation of the CD34+ cells . Mononuclear cells (MNC) were cultured in serum-free liquid culture system. Experiments were divided into 3 groups. In the group A as control no cytokines were added in the culture system;in the group B the cells cultured with SCF+FL+TPO+EPO+IGF-1; in the group C the cells were cultured with SCF+FL+TPO. EPO and IGF-1 were added at day 6. Part of the renewed MNCs and colony-forming units were counted, the proportion of CD34+, CD34+ CD71+, CD71+ GPA+ cells was detected by FACS. The result showed that after 10 days, the the total cord blood cells increased 6.89-folds in group B and 3.06-folds in group C (P<0.05). The CD34+ cells increased 4.83-folds in group B and 2.47-folds in group C (P<0.05). The colony-forming cells (CFCs) increased 4.3-folds in group B and 2.5-folds in group C (P<0.05) . Erythroid progenitors( BFU-E) and CFU-E increased 5.4-folds in group B and 3.1-folds in group C (P<0.05). The CD34+CD71+cells increased 8.72-folds in group B and 3.37-folds in group C (P<0.05). The CD71+GPA+ cells increased 53.4-folds in group B and 30.29-folds in group C.They were different at any time point (P<0.05). It is concluded that in the group with FL+SCF+TPO, CD34+ cells and CFC can greatly be expanded from cord blood MNC in the serum-free culture system. In the group with FL+SCF+TPO+EPO+IGF-1, erythroid progenitors can greatly be expanded in the serum-free culture system, supplying EPO at day 0 was better than supplying at day 6 . Since the largest number of colony-fo

关 键 词：脐血 单个核细胞 红系祖细胞 无血清培养 CD34^+细胞 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]