小鼠骨髓间充质干细胞对胚胎干细胞造血分化的影响  

Effect of Bone Marrow Mesenchymal Stem Cells on Hematopoietic Differentiation of Murine Embryonic Stem Cells

作　　者：王晓燕[1] 刘兵[1] 苑春慧[1] 要晖宇[1] 毛宁[1] 

机构地区：[1]军事医学科学院基础医学研究所细胞生物学研究室,北京100850

出　　处：《中国实验血液学杂志》2003年第4期329-334,共6页Journal of Experimental Hematology

基　　金：国家高技术研究发展计划"863"资助项目(编号2 0 0 1AA2 1713 1);国家重点基础研究发展规划"973"资助项目 (编号G19990 5 43 0 2 )

摘　　要：骨髓间充质干细胞作为骨髓基质细胞的前体细胞 ,在体外具有一定的造血支持作用 ,与造血干细胞共移植可促进其植入。本研究旨在初步探讨应用小鼠骨髓间充质干细胞与胚胎干细胞共培养作为一种新的分化体系的可行性。首先分离、培养并鉴定小鼠骨髓间充质干细胞 ,然后利用扩增培养的骨髓间充质干细胞与胚胎干细胞共培养 ,通过造血集落培养和RT PCR观察造血分化的特点。结果表明 ,分离、扩增培养至第四代之后的骨髓间充质干细胞形态均一 ,高表达Sca 1,CD2 9,CD4 4和CD10 5 ,而CD34和CD4 5等造血与内皮细胞特异性表面标志呈阴性 ;特异性诱导体系内传代后 (>4代 )的骨髓间充质干细胞可向脂肪细胞和成骨细胞分化。与悬浮分化体系相比 ,骨髓间充质干细胞共培养体系中初始分化的胚胎干细胞含有显著增加的拟胚体形成细胞而没有造血集落形成细胞。此外 ,RT PCR检测发现 :共培养细胞表达胚胎干细胞特异性转录因子Oct 4 ,而造血标记Flk 1,GATA 1和 β H1为阴性。结论 :间充质干细胞在一定程度上抑制了胚胎干细胞的初始分化 。Mesenchymal stem cells (MSCs), precursors of diverse stromal cells, can support hematopoiesis in vitro and can promote the implantation of hematopoietic stem cells in vivo when co transplanted with CD34 + cells. The aim of this study was to investigate the potential effect of MSCs on the hematopoietic development of embryonic stem cells (ES cells) and the feasibility of a novel system in which ES cells will be co cultured with MSCs. The murine bone marrow MSCs were isolated and cultured and then their phenotye and differentiation function were identified with FCM and histochemical technique. The CCE cells, murine ES cell line, were co cultured with the isolated MSCs and the hematopoietic differentiation of CCE cells was observed with hematopoietic clonogenic assay and RT PCR. The results showed that the morphology of MSCs became gradually homogeneous with the passage culture of cells. After passage 4, the marker of Sca 1, CD29, CD44 and CD105 were highly expressed, however, CD34 and CD45, the specific marker of hematopoietic and endothelial cells, could hardly be identified. The isolated MSCs differentiated into adipocytes and osteoblasts in specific induction culture system. After maintaining culture on mouse embryonic fibroblasts, CCE cells were plated in suspended culture system with only differentiation inductive agents and co culture system in which MSCs were added. Compared with CCE cell suspended culture, the cells differentiated into embryoid body were obviously enhanced and there were no colony forming cells in the co culture system of ES cells and MSCs. In addition, transcription factor Oct 4 in co cultured CCE cells was expressed and hematopoietic markers, Flk 1, GATA 1 and β H1, were negative. The ability of embryoid bodies derived from the co culture system to produce hematopoietic colonies was markedly higher than that from the suspended culture system. It is concluded that MSCs inhibit the initial differentiation of ESC and enhance hematopoietic differentiation ability of the co cu

关 键 词：胚胎干细胞 间充质干细胞 集落形成细胞 造血分化 

分 类 号：R331.2[医药卫生—人体生理学]