Section of Hematology, Department of Internal Medicine and Yale Cancer Center, Yale University School of Medicine, New Haven, CT 06520-8073, USA.
Exp Hematol. 2010 Feb;38(2):90-103. doi: 10.1016/j.exphem.2009.11.004. Epub 2009 Dec 5.
Mutations in the CCAAT enhancer binding protein epsilon (C/EBPepsilon) gene have been identified in the cells of patients with neutrophil specific granule deficiency, a rare congenital disorder marked by recurrent bacterial infections. Their neutrophils, in addition to lacking specific granules required for normal respiratory burst activity, also lack normal phagocytosis and chemotaxis. Although the specific granule deficiency phenotype has been replicated in C/EBPepsilon(-/-) (knockout [KO]) mice, the mechanisms by which C/EBPepsilon mutations act to decrease neutrophil function are not entirely clear.
In order to determine the role of C/EBPepsilon in neutrophil differentiation and migration, we generated immortalized progenitor cell lines from C/EBPepsilon KO and wild-type mice and performed expression and flow cytometric analysis and functional studies.
Expression of lineage-specific cell surface antigens on our in vitro differentiated cell lines revealed persistent expression of monocytic markers on KO granulocytes. We verified this in primary murine peripheral blood and bone marrow cells. In addition, KO bone marrow had an increase in immature myeloid precursors at the common myeloid progenitor and granulocyte/monocyte progenitor levels, suggesting a critical role for C/EBPepsilon not only in granulocyte maturation beyond the promyelocyte stage, but also in the monocyte/granulocyte lineage decision. We found that restoration of Hlx (H2.0-like homeo box 1) expression, which was decreased in C/EBPepsilon KO cells, rescued chemotaxis, but not the other defects of C/EBPepsilon KO neutrophils.
We show two new regulatory functions of C/EBPepsilon in myelopoiesis: in the absence of C/EBPepsilon, there is not only incomplete differentiation of granulocytes, but myelopoiesis is disrupted with the appearance of an intermediate cell type with monocyte and granulocyte features, and the neutrophils have abnormal chemotaxis. Restoration of expression of Hlx provides partial recovery of function; it has no effect on neutrophil maturation, but can completely ameliorate the chemotaxis defect in C/EBPepsilon KO cells.
在中性粒细胞特异性颗粒缺乏症患者的细胞中已经鉴定出 CCAAT 增强子结合蛋白 epsilon（C/EBPepsilon）基因突变，这是一种罕见的先天性疾病，其特征是反复发生细菌感染。他们的中性粒细胞除了缺乏正常呼吸爆发活动所需的特异性颗粒外，还缺乏正常的吞噬作用和趋化性。尽管 C/EBPepsilon(-/-)（敲除[KO]）小鼠已经复制了特异性颗粒缺乏表型，但 C/EBPepsilon 突变作用于降低中性粒细胞功能的机制尚不完全清楚。
为了确定 C/EBPepsilon 在中性粒细胞分化和迁移中的作用，我们从 C/EBPepsilon KO 和野生型小鼠中生成了永生化祖细胞系，并进行了表达和流式细胞术分析以及功能研究。
我们体外分化细胞系上谱系特异性细胞表面抗原的表达显示 KO 粒细胞上持续表达单核细胞标志物。我们在原代鼠外周血和骨髓细胞中验证了这一点。此外，KO 骨髓在共同髓系祖细胞和粒细胞/单核细胞祖细胞水平上具有增加的未成熟髓系前体，这表明 C/EBPepsilon 的作用不仅在嗜中性粒细胞成熟超出早幼粒细胞阶段，而且在单核细胞/粒细胞谱系决定中起关键作用。我们发现，恢复在 C/EBPepsilon KO 细胞中表达降低的 Hlx（H2.0 样同源盒 1）表达可挽救趋化性，但不能挽救 C/EBPepsilon KO 中性粒细胞的其他缺陷。
我们在髓样发生中展示了 C/EBPepsilon 的两个新的调节功能：在缺乏 C/EBPepsilon 的情况下，不仅粒细胞的分化不完全，而且髓样发生被破坏，出现具有单核细胞和粒细胞特征的中间细胞类型，并且中性粒细胞的趋化性异常。Hlx 表达的恢复提供了功能的部分恢复；它对中性粒细胞成熟没有影响，但可以完全改善 C/EBPepsilon KO 细胞的趋化性缺陷。
