Terstappen L W, Nguyen M, Lazarus H M, Medof M E
Becton Dickinson Immunocytometry Systems, San Jose, CA 95131.
J Leukoc Biol. 1992 Dec;52(6):652-60. doi: 10.1002/jlb.52.6.652.
Expression of decay-accelerating factor (DAF or CD55) and of CD59 during hematopoietic cell development in normal bone marrow and on peripheral blood leukocytes were characterized by three-color immunofluorescence experiments. With this technique cell subsets were identified by forward light scatter, orthogonal light scatter, and two cell-surface antigens. For each cell lineage, specific combinations of two monoclonal antibodies labeled with different fluorochromes were used. DAF or CD59 were then quantitated on the defined cell subsets from the fluorescence signal of the respective antibody conjugated with a third fluorochrome. Early uncommitted hematopoietic progenitor cells (CD34+, CD38-) all expressed both proteins homogeneously. Initial commitment to the erythroid (CD71+, CD45dim), myeloid (CD33+), or B lymphocyte (CD10+) lineages was not associated with changes in DAF or CD59 levels. With erythroid development, i.e., after loss of CD45 and decrease of CD71, expression of both proteins decreased. With myeloid maturation, expression of CD59 remained constant and expression of DAF varied. During neutrophil maturation, DAF decreased initially and then reemerged on maturing neutrophils concurrently with the appearance of CD16 (Fc gamma RIII), whereas during monocyte maturation, DAF increased concurrently with up-regulation of CD14. With B cell development, expression of DAF increased concurrently with down-regulation of CD10 and up-regulation of CD20, whereas expression of CD59 diminished slightly late in B cell maturation. Analysis of peripheral blood elements showed that monocytes, neutrophils, and B lymphocytes expressed both proteins homogeneously, but that in contrast to other cell subsets, which all expressed CD59, only a subset of (CD3+) T lymphocytes and (CD16+) Natural killer cells expressed DAF. The absence of DAF was not related to CD4 or CD8 expression or to the presence of activation markers (CD25+, CD38+), memory cell markers (CD58+, CD45RO+), or virgin T cell markers (CD45RA+), but was correlated with expression of CD11b (CR3) and CD11c (gp150/95). Although CD21+ (CR2) and CD35+ (CR1) cells all expressed DAF, CD11a (LFA-1) levels correlated inversely with those of DAF. Although the presence of CD55 and CD59 on early progenitor cells and throughout hematopoietic cell development is consistent with the requirements for both proteins in protection of host cells from complement-mediated injury, the physiological relevance of the unique patterns of variation for each cell lineage is unclear. Nevertheless, the availability of a detailed DAF and CD59 expression map in normal marrow will facilitate analyses of alterations during hematopoietic development that may occur in hematological disorders including paroxysmal nocturnal hemoglobinuria (PNH).
通过三色免疫荧光实验对正常骨髓造血细胞发育过程中以及外周血白细胞上衰变加速因子（DAF 或 CD55）和 CD59 的表达进行了表征。利用该技术，通过前向光散射、正交光散射和两种细胞表面抗原鉴定细胞亚群。对于每个细胞谱系，使用了两种用不同荧光染料标记的单克隆抗体的特定组合。然后根据与第三种荧光染料偶联的相应抗体的荧光信号，对定义的细胞亚群上的 DAF 或 CD59 进行定量。早期未定向造血祖细胞（CD34 +、CD38 -）均均匀表达这两种蛋白。向红系（CD71 +、CD45dim）、髓系（CD33 +）或 B 淋巴细胞（CD10 +）谱系的初始定向与 DAF 或 CD59 水平的变化无关。随着红系发育，即 CD45 丢失和 CD71 降低后，这两种蛋白的表达均降低。随着髓系成熟，CD59 的表达保持恒定，而 DAF 的表达有所变化。在中性粒细胞成熟过程中，DAF 最初降低，然后在成熟中性粒细胞上重新出现，同时出现 CD16（FcγRIII），而在单核细胞成熟过程中，DAF 随着 CD14 的上调而增加。随着 B 细胞发育，DAF 的表达随着 CD10 的下调和 CD20 的上调而增加，而 CD59 的表达在 B 细胞成熟后期略有降低。对外周血成分的分析表明，单核细胞、中性粒细胞和 B 淋巴细胞均均匀表达这两种蛋白，但与所有表达 CD59 的其他细胞亚群不同，只有一部分（CD3 +）T 淋巴细胞和（CD16 +）自然杀伤细胞表达 DAF。DAF 的缺失与 CD4 或 CD8 的表达无关，也与激活标志物（CD25 +、CD38 +）、记忆细胞标志物（CD58 +、CD45RO +）或原始 T 细胞标志物（CD45RA +）的存在无关，但与 CD11b（CR3）和 CD11c（gp150/95）的表达相关。尽管 CD21 +（CR2）和 CD35 +（CR1）细胞均表达 DAF，但 CD11a（LFA - 1）水平与 DAF 水平呈负相关。尽管早期祖细胞以及整个造血细胞发育过程中 CD55 和 CD59 的存在与这两种蛋白在保护宿主细胞免受补体介导的损伤方面的需求一致，但每个细胞谱系独特的变化模式的生理相关性尚不清楚。然而，正常骨髓中详细的 DAF 和 CD59 表达图谱的可得性将有助于分析造血发育过程中可能发生在包括阵发性夜间血红蛋白尿（PNH）在内的血液系统疾病中的改变。
