不同造血微环境对诱导人胚胎干细胞向造血细胞分化的影响  

Effect of different hemopoietic microenvironment on the differentiation of hemopoietic cells from human embryonic stem cells

作　　者：赵惠萍[1] 赵海军[1] 林戈[1] 周菂[1] 刘天成[1] 卢光琇[1] 

机构地区：[1]中南大学生殖与干细胞工程研究所

出　　处：《中南大学学报（医学版）》2007年第6期992-996,共5页Journal of Central South University ：Medical Science

基　　金：国家自然科学基金(30300119);博士点基金(20030533002)~~

摘　　要：目的:通过观察人卵黄囊基质细胞、人胎肝基质细胞和人胎骨髓基质细胞对人胚胎干细胞向造血细胞分化诱导效率,探讨不同造血微环境对造血发生的影响。方法:本实验诱导人胚胎干细胞分化为造血细胞的方法采用两步法:先用细胞因子培养形成5d的拟胚体,然后再模拟体内造血发生的微环境,分别用人卵黄囊基质细胞、人胎肝基质细胞和人胎骨髓基质细胞诱导拟胚体10d,通过流式细胞术检测细胞的flk,CD34和CD45表面抗原表达情况,观察3种基质细胞对人胚胎干细胞向造血细胞分化的影响。结果:5d拟胚体与卵黄囊基质细胞接触培养10d生成的细胞表达flk,CD34,CD45百分率分别1.80%±0.56%,1.30%±0.14%,1.05%±0.63%;与胎肝基质细胞接触培养10d生成的细胞表达为flk,CD34,CD45百分率分别为34.0%±25.45%,38.4%±24.8%,72.6%±25.7%;与骨髓基质细胞接触培养10d生成的细胞表达flk,CD34,CD45百分率分别为2.50%±1.48%,3.20%±0.56%,1.65%±0.21%;5d拟胚体自发分化10d生成的细胞表达flk,CD34,CD45百分率分别为0.30%±0.07%,0.65%±0.07%,0.15%±0.07%。与自发分化10d比较,3种基质细胞与5d拟胚体接触培养,均能够促进拟胚体向造血细胞的分化,且胎肝基质细胞诱导的效率显著优于其他两种基质细胞(P(0.05)。结论:与卵黄囊基质细胞和骨髓基质细胞比较,胎肝基质细胞更有利于诱导人胚胎干细胞向造血细胞的分化。Objective To observe the inductive efficiency of deriving hematopoietic cells from human embryonic stem（hES）cells co-cultured with human yolk sac stromal cells,fetal liver stromal cells or fetal bone marrow stromal cells,in order to discuss the effect of the different hemopoietic microenvironment on hemopoietic cytogenesis.Methods We used two-step method to induce the hES cells into the hematopoietic cells.In the first step the hES cells were co-cultured with cytokines by formation of the day 5 embryoid bodies（5 d EBs）.In the second step the 5d EB cells were induced into the hematopoietic cells by co-culturing with human yolk sac stromal cells,fetal liver stromal cells or fetal bone marrow stromal cells for 10 days.The inductive efficiencies of deriving hematopoietic cells from hES cells co-cultured with the different hemopoietic microenvironment were reflected by the expression levels of flk,CD34 and CD45 antigen.Results Flow cytometry analysis demonstrated that the population of the cells co-cultured with human yolk sac stromal cells contained flk（1.80%±0.56%）,CD34（1.30%±0.14%）or CD45（1.05%±0.63%）positive cells;the population of the cells co-cultured with human fetal liver stromal cells contained flk（34.00%±25.45%）,CD34（38.40%±24.80%）or CD45（72.60%±25.70%）positive cells;the population of the cells co-cultured with human fetal bone marrow stromal cells contained flk（2.50%±1.48%）,CD34（3.20%±0.56%）or CD45（1.65%±0.21%）positive cells.Compared with spontaneous differentiation of EBs,all of the three stromal cells could induce EBs into the hematopoietic cells（P〈0.05）.Conclusion The inductive efficiency of deriving hematopoietic cells from EBs co-cultured with human fetal liver stromal cells was higher than EBs co-cultured with human yolk sac stromal cells and fetal bone marrow stromal cells.

关 键 词：胚胎干细胞 造血细胞 卵黄囊基质细胞 胎肝基质细胞 骨髓基质细胞 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]