Identification of the causative gene for inherited sideroblastic anemia and analysis of sideroblastis induced in vitro

遗传性铁粒幼细胞贫血致病基因的鉴定及体外诱导铁粒幼细胞的分析

基本信息

批准号：
11670977
负责人：
HARIGAE Hideo
金额：
$ 2.3万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

alas2 encodes the erythroid specific 8-aniinoleyuIinate synthase (ALAS2), the first enzyme in heme biosynthesis in erythroid cells. In vivo in mice, alas2-null primitive erythroblasts showed a maturation arrest as well as massive cytoplasmic iron accumulation. However, the effect of ALAS2 deficiency, on definitive erythropoiesis still remains unclear, because alas2-null mice died in utero by embryonic day 11.5 before definitive erythropoiesis developed. To clarity the effect of heme deficiency on differentiation of definitive erythroid cells, ES cells lacking the alas2 gene were induced to differentiate specifically into erythroid lineage in vitro, and phenotypes of definitive erythroblasts were examined. In comparison to red cell pellets of Wild-type erythroblasts, alas2-null definitive erythroblasts were totally white due to a marked deficiency of heme, although the morphology was similar to wild-type erythroblasts. Consistent with their similar morphology, levels of erythroid-specific gene expression such as GATA-1, NF-E2 andTER119 in alas2-null definitive erythroblasts were also similar to those of wild-type cells, indicating that both alas2-null and wild-type erythroblasts developed to the stage of mature definitive erythroblasts. In contrast to their similar morphology, however, iron content in alas2-null definitive erythroblasts was twice more than that of wild-type, cells. Consistent with the aberrant increase in iron content, alas2-null definitive erythroblasts were more oxidized compared with wild-type erythroblasts as judged by increased formation of peroxidized metabolites. These findings suggest that alas2 deficiency does not influence erythroid differentiation per se, but induces aberrant iron accumulation and oxidized condition in definitive erythroblasts.

alas2 编码红细胞特异性 8-氨基油酰酸合酶 (ALAS2)，这是红细胞中血红素生物合成的第一个酶。在小鼠体内，alas2 缺失的原始成红细胞表现出成熟停滞以及大量细胞质铁积累。然而，ALAS2 缺陷对确定性红细胞生成的影响仍不清楚，因为 alas2 缺失小鼠在确定性红细胞生成发育之前的胚胎第 11.5 天就在子宫内死亡。为了阐明血红素缺乏对定形红细胞分化的影响，在体外诱导缺乏alas2基因的ES细胞特异性分化为红细胞谱系，并检查了定形红细胞的表型。与野生型成红细胞的红细胞沉淀相比，由于血红素明显缺乏，alas2 缺失的定型成红细胞是全白色的，尽管形态与野生型成红细胞相似。与它们相似的形态相一致，alas2 缺失定型成红细胞中的红系特异性基因表达水平（如 GATA-1、NF-E2 和 TER119）也与野生型细胞相似，表明 alas2 缺失和野生型细胞成红细胞发育至成熟定型成红细胞阶段。然而，与它们相似的形态相反，alas2 缺失的定形成红细胞中的铁含量是野生型细胞的两倍。与铁含量的异常增加一致，根据过氧化代谢物形成的增加判断，与野生型成红细胞相比，alas2 缺失的定型成红细胞氧化程度更高。这些发现表明，alas2 缺乏本身并不影响红细胞分化，但会诱导终形红细胞中异常的铁积累和氧化状态。