Division of Molecular Genetics, Cancer Research Institute, Kanazawa University, Kanazawa, Japan.
Department of Hematology, Endocrinology and Metabolism, Faculty of Medicine, Niigata University, Niigata, Japan.
Sci Rep. 2017 Sep 12;7(1):11442. doi: 10.1038/s41598-017-11909-3.
Hematopoietic stem cells (HSCs) in a steady state can be efficiently purified by selecting for a combination of several cell surface markers; however, such markers do not consistently reflect HSC activity. In this study, we successfully enriched HSCs with a unique stemness-monitoring system using a transgenic mouse in which green florescence protein (GFP) is driven by the promoter/enhancer region of the nucleostemin (NS) gene. We found that the phenotypically defined long-term (LT)-HSC population exhibited the highest level of NS-GFP intensity, whereas NS-GFP intensity was strongly downregulated during differentiation in vitro and in vivo. Within the LT-HSC population, NS-GFP cells exhibited significantly higher repopulating capacity than NS-GFP cells. Gene expression analysis revealed that nine genes, including Vwf and Cdkn1c (p57), are highly expressed in NS-GFP cells and may represent a signature of HSCs, i.e., a stemness signature. When LT-HSCs suffered from remarkable stress, such as transplantation or irradiation, NS-GFP intensity was downregulated. Finally, we found that high levels of NS-GFP identified HSC-like cells even among CD34 cells, which have been considered progenitor cells without long-term reconstitution ability. Thus, high NS-GFP expression represents stem cell characteristics in hematopoietic cells, making this system useful for identifying previously uncharacterized HSCs.
造血干细胞（HSCs）在稳定状态下可以通过选择几种细胞表面标志物的组合来有效地进行纯化；然而，这些标志物并不始终反映 HSC 的活性。在这项研究中，我们使用一种转基因小鼠成功地利用独特的干细胞监测系统富集了 HSCs，在该系统中，绿色荧光蛋白（GFP）由核干细胞（NS）基因的启动子/增强子区域驱动。我们发现，表型定义的长期（LT）-HSC 群体表现出最高的 NS-GFP 强度，而 NS-GFP 强度在体外和体内分化过程中被强烈下调。在 LT-HSC 群体中，NS-GFP 细胞表现出明显更高的重编程能力比 NS-GFP 细胞。基因表达分析显示，包括 Vwf 和 Cdkn1c（p57）在内的九个基因在 NS-GFP 细胞中高度表达，这可能代表了 HSCs 的一个特征，即干细胞特征。当 LT-HSCs 遭受明显的压力时，如移植或辐射，NS-GFP 强度会下调。最后，我们发现，即使在 CD34 细胞中，高水平的 NS-GFP 也能识别出类 HSC 细胞，而 CD34 细胞被认为是没有长期重建能力的祖细胞。因此，高 NS-GFP 表达代表造血细胞中的干细胞特征，使该系统可用于鉴定以前未表征的 HSCs。
