CD34(+) ENDOTHELIAL-CELL LINES DERIVED FROM MURINE YOLK-SAC INDUCE THE PROLIFERATION AND DIFFERENTIATION OF YOLK-SAC CD34(+) HEMATOPOIETIC PROGENITORS
CD34(+) ENDOTHELIAL-CELL LINES DERIVED FROM MURINE YOLK-SAC INDUCE THE PROLIFERATION AND DIFFERENTIATION OF YOLK-SAC CD34(+) HEMATOPOIETIC PROGENITORS
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DOI：
10.1182/blood.v86.12.4454.bloodjournal86124454
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发表时间：
1995-12-15
期刊：
影响因子：
20.3
通讯作者：
LASKY, LA
中科院分区：
文献类型：
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作者：
FENNIE, C;CHENG, J;LASKY, LA
胚胎造血部分起始于卵黄囊的血岛。以前的共聚焦显微镜分析表明，CD 34抗原，一种粘蛋白样细胞表面糖蛋白，表达的造血祖细胞和所有的内皮细胞的成人和胚胎，也被发现在一个子集的腔造血样细胞在卵黄囊血岛以及血管内皮内衬这些早期造血位置。我们发现，与迄今为止所研究的所有其他造血部位一样，免疫亲和纯化的来自小鼠卵黄囊的CD 34（+）非粘附细胞含有绝大多数红系和髓系祖细胞集落形成活性。为了检测这些卵黄囊的CD 34（+）造血祖细胞和CD 34（+）卵黄囊内皮之间的发育相互作用，我们使用CD 34抗血清从第10.5天的卵黄囊中免疫亲和纯化贴壁内皮细胞，并通过用表达多瘤中间T抗原的逆转录病毒转化产生细胞系。对这些细胞系的CD 34、血管性血友病因子、FLK 1和FLT 1表达以及基质胶中的毛细血管生长的分析表明，它们似乎是内皮细胞，与它们在体内的原始表型一致。在这些内皮细胞系上共培养卵黄囊CD 34（+）造血细胞，约8天后总造血细胞数增加60倍。对这些扩增的造血细胞的分析表明，大多数是单核细胞/巨噬细胞谱系。此外，对培养物的检查显示了许多鹅卵石区域的快速形成，这是一种先前描述的形态实体，被认为是早期多能干细胞的代表。对这些内皮细胞系扩增定向祖细胞的能力的检查显示，在培养3至6天后，红系和髓系集落形成细胞增加了6倍，这与这些胚胎内皮细胞介导这些前体细胞扩增的观点一致。聚合酶链反应分析显示，大多数细胞系产生FLK-2/FLT-3配体、干细胞因子、巨噬细胞集落刺激因子、白血病抑制因子和白细胞介素6（IL-6），而粒细胞集落刺激因子、粒细胞-巨噬细胞集落刺激因子、IL-1、IL-3、转化生长因子β-1、促红细胞生成素、或血小板生成素。可以通过添加外源性促红细胞生成素来修饰来自这些共培养物的成熟造血细胞的输出以包括红细胞群。这些数据表明，衍生自卵黄囊的内皮细胞系为卵黄囊祖细胞扩增和分化成至少髓系和红系谱系提供了适当的造血环境。(C)1995年，美国血液学会。
Embryonic hematopoiesis is initiated in part in the blood islands of the yolk sac. Previous confocal microscopic analysis has shown that the CD34 antigen, a mucin-like cell surface glycoprotein that is expressed by hematopoietic progenitors and all endothelial cells of the adult and embryo, is also found on a subset of luminal hematopoietic-like cells in the yolk sac blood islands as well as on the vascular endothelium lining these early hematopoietic locations. We show here that, as in all other hematopoietic sites thus far examined, immunoaffinity-purified CD34(+) nonadherent cells from murine yolk sacs contain the vast majority of erythroid and myeloid progenitor cell colony forming activity. To examine the developmental interactions between these CD34(+) hematopoietic progenitor cells of the yolk sac and the CD34(+) yolk sec endothelium, we have immunaffinity-purified adherent endothelial cells from day 10.5 yolk sacs using CD34 antiserum and produced cell lines by transformation with a retrovirus expressing the polyoma middle T antigen. Analysis of these cell lines for CD34, von Willebrand's factor, FLK 1 and FLT 1 expression, and capillary growth in Matrigel indicates that they appear to be endothelial cells, consistent with their original phenotype in vivo. Coculture of yolk sac CD34(+) hematopoietic cells on these endothelial cell lines results in up to a 60-fold increase in total hematopoietic cell number after approximately 8 days. Analysis of these expanded hematopoietic cells showed that the majority were of the monocyte/macrophage lineage. In addition, examination of the cultures showed the rapid formation of numerous cobblestone areas, a previously described morphologic entity thought to be representative of early pluripotential stem cells. Scrutiny of the ability of these endothelial cell lines to expand committed progenitor cells showed up to a sixfold increase in erythroid and myeloid colony-forming cells after 3 to 6 days in culture, consistent with the notion that these embryonic endothelial cells mediate the expansion of these precursor cells. Polymerase chain reaction analyses showed that most of the cell lines produce FLK-2/FLT-3 ligand, stem cell factor, macrophage colony-stimulating factor, leukemia-inhibitory factor, and interleukin 6 (IL-6), whereas there is a generally low or not measurable production of granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, IL-1, IL-3, transforming growth factor beta-1, erythropoietin, or thrombopoietin. The output of mature hematopoietic cells from these cocultures can be modified to include an erythroid population by the addition of exogenous erythropoietin. These data suggest that endothelial cell lines derived from the yolk sac provide an appropriate hematopoietic environment for the expansion and differentiation of yolk sac progenitor cells into at least the myeloid and erythroid lineages. (C) 1995 by The American Society of Hematology.