含人AGM区基质细胞对小鼠胚胎干细胞诱导体外分化为造血干/祖细胞的作用  

Effects of Human Aorta-gonad-mesonephros Region Stromal Cells on Inducing Differentiation of Murine Embryonic Stem Cells into Hematopoietic Stem /Progenitor Cells In Vitro

作　　者：蔡耘[1,2] 张绪超[2,3] 陈惠芹[1,2] 吴北燕[2] 黄绍良[2] 

机构地区：[1]中山大学附属第三医院儿科,广东广州510630 [2]中山大学附属第二医院干细胞研究中心,广东广州510120 [3]广东省人民医院医学研究中心,广东广州510080

出　　处：《中国实验血液学杂志》2011年第4期1023-1027,共5页Journal of Experimental Hematology

基　　金：国家自然科学基金资助项目(编号30300377);广东省科技计划项目(编号2008B030301046)

摘　　要：为探讨人主动脉一性腺一中肾(AGM)区基质细胞对小鼠胚胎干细胞(em bryonic stem cells,ESC)体外诱导分化为造血干细胞(HSC)的影响及其作用机制,将小鼠E14ESC诱导为拟胚体(EB),收获的EB细胞以Transw ell非接触共培养体系分别在人AGM区、胎肝(FL)或骨髓(BM)基质细胞饲养层上进一步诱导分化,分为EB对照、AGM诱导、FL诱导、BM诱导、AGM+FL诱导和AGM+BM诱导共6组。分别收获各组EB来源细胞,以流式细胞仪检测Sca-1+c-K it+细胞含量,并进行各系造血细胞集落形成单位(CFU)分析。结果显示:经不同基质细胞饲养层诱导后EB来源细胞中Sca-1+c-K it+细胞比例均较诱导前明显升高(p<0.01),AGM+FL诱导组为(21.96±2.54)%,显著优于其它诱导组(p<0.01),AGM+BM诱导组阳性细胞比例亦较单纯BM诱导组明显增加(p<0.01);EB对照组造血集落产率明显低于其它诱导组,AGM+FL、AGM+BM诱导组集落产率较高,尤以AGM+FL诱导组最佳(p<0.01)。结论:人AGM区基质细胞有助于维持一定的原始HSC数目及高增殖潜能,在AGM区基质细胞诱导基础上可明显提高FL或BM基质细胞对ESC源性原始HSC的进一步扩增作用。This study was aimed to investigate the effect of human aorta-gonad-mesonephros （AGM） region stromal cells on differentiation of murine embryonic stem cells （ESC） into hematopoietic stem cells （HSC） in vitro and to clarify their effect mechanism. El4 murine ESC were induced into embryo body （EB） firstly. Then the EB cells were further co-cultured with the stromal cells from human AGM region, fetal liver （FL） or bone marrow （BM） in Transwell non-contact system. According to the different culture methods, the EB cells were divided into 6 groups including EB control group, AGM group, FL group, BM group, AGM ＋ FL group and AGM ＋ BM group. The induced cells derived from EB were collected for Sca-1 ＋/c-Kit ＋ cells analysis by flow cytometry and colony forming unit （CFU） assay. The results showed that Sca-1 ＋/c-Kit ＋ cell proportion of EB cells significantly increased after being induced by different stromal cells （p 〈0.01 ）. The AGM ＋ FL group had most Sca-1 ＋/c-Kit＋ cells for the positive cell proportion reached （21.96 ＋ 2.54） % （p 〈0.01 ）. The Sca-1 ＋/c-Kit ＋ cell proportion of AGM ＋ BM group was much high than that of BM group too （p 〈 0.01 ）. The EB control group showed CFU amount less than any other groups, while the CFU amount of AGM ＋ FL, AGM ＋ BM groups were higher, especially in the AGM ＋ FL group （p 〈0.01 ）. It is concluded that the human AGM region stromal cells may help to maintain certain number of primitive HSC with high proliferation potential. Human AGM region, FL or BM stromal cells, applied in sequential orders, can significantly expand in vitro the primitive hematopoietic stem/progenitor cells derived from ESC.

关 键 词：主动脉-性腺-中肾区 基质细胞 胚胎干细胞 造血干细胞 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学] R331.2[医药卫生—基础医学]