Searching for the factors concerning hematopoiesis

这项研究的目的之一是克隆在早期造血中起一定作用的因子的基因。为此，我们将小鼠胚胎干细胞体外诱导造血细胞分化的方法与信号序列捕捉法相结合，筛选出含有信号肽的分泌蛋白。从第5天诱导的含有血管母细胞和造血祖细胞的中胚层细胞克隆中构建cDNA文库。本研究克隆了ESOP-1基因。推导的氨基酸序列显示160个氨基酸的蛋白质。人ESOP-1基因也被克隆出来，与的同源性为%。ESOP-1基因在妊娠7.5天的胚胎中高表达。该基因在成体和胚胎造血系统中均有表达。免疫组织化学染色显示该蛋白在卵黄囊、发育中的神经系统和成体生殖器中均有表达。另一项结果是…分析BclX基因在红细胞生成中的更多作用。我们利用了BclX缺失的小鼠胚胎干细胞(ES细胞)，并证明了BclX对于胚胎原始红细胞(EryP)和成体最终红细胞(EryD)的形成都是不可或缺的。在体内，bcl-x缺失的ES细胞对成年嵌合小鼠的循环EryD没有贡献，而这是通过囊胚显微注射bcl-x缺失的ES细胞产生的。将ES细胞在巨噬细胞集落刺激因子缺陷型基质细胞系OP9上进行体外诱导分化，产生bclx基因缺失的EryP和EryD，并进行进一步分析。Bclx基因缺失的ES细胞出现未成熟的EryP和EryD与正常ES细胞相似。而bclx基因缺失的EryP和EryD在细胞成熟后出现明显的细胞死亡。促红细胞生成素被认为可以防止红系祖细胞的凋亡性死亡，而bclx的抗凋亡作用发生在成熟末期。较少

One of the purposes of this study is cloning the genes of the factors that have some roles in early hematopoiesis. For this purpose, we combined the in vitro hematopoietic differentiation induction method from mouse embryonic stem cells and the signal sequence trap method that is a method to select secretory proteins containing signal peptide. cDNA library was constructed from the day 5 induced mesodermal cell colonies in which hemangioblasts and hematopoietic progenitors existed. As a result, cDNA named ESOP-1 was cloned. The deduced amino acid sequence shows 160 amino acids protein. Human ESOP-1 was also cloned and shows 64% homology. ESOP-1 cDNA was highly expressed in embryos at day 7.5 of gestation. This gene was also expressed in both adult and embryonic hematopoietic system. Antibody was produced against this protein and immuno staining revealed that the protein was expressed in yolk sac, developing nervous system and adult genital organs.The other outcome is the analysis of the … More fuction of Bcl-X gene on erythropoiesis. We utilized Bcl-X null mouse embryonic stem cells (ES cells), and showed that Bcl-X is indispensable for the production of both embryonic primitive erythrocytes (EryP) and adult definitive erythrocytes (EryD) at the end of their maturation. In vivo, bcl-x null ES cells did not contribute to circulating EryD in adult chimeric mice that were produced by blastocyst microinjection of the bcl-x null ES cells. bcl-x null EryP and EryD were produced by in vitro differentiation induction of ES cells on a macrophage colony-stimulating factor deficient stromal cell line OP9, and further analysis was carried out. The emergence of immature EryP and EryD from bcl-x null ES cells was similar to that from normal ES cells. However, prominent cell death of bcl-x null EryP and EryD occurred when the cells matured. Erythropoietin is thought to prevent the apoptotic cell death of erythroid progenitor cells, while the anti-apoptotic function of bcl-x acts at the very end of maturation. Less