Characterization of murine CD34, a marker for hematopoietic progenitor and stem cells.
Characterization of murine CD34, a marker for hematopoietic progenitor and stem cells.
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DOI：
10.1182/blood.v84.3.691.691
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发表时间：
1994-08
期刊：
影响因子：
20.3
通讯作者：
D. Krause;T. Ito;M. Fackler;OM Smith;M. Collector;S. Sharkis;W. May
中科院分区：
文献类型：
--
作者：
D. Krause;T. Ito;M. Fackler;OM Smith;M. Collector;S. Sharkis;W. May
CD34在人的造血干/祖细胞上表达，其在干细胞纯化中的临床应用已得到证实。然而，小鼠CD34(MCD34)的类似表达模式尚未确定。为了研究mCD34蛋白在小鼠细胞系和造血祖细胞上的表达模式，我们研制了两种能识别内源性和重组小鼠CD34蛋白的多克隆抗体。荧光激活细胞分类分析表明，mCD34表达于NIH/3T3胚胎成纤维细胞、PA6基质细胞、胚胎干细胞、M1白血病细胞和正常骨髓细胞亚群。小鼠CD34是一种糖蛋白，在NIH/3T3和PA6细胞中以全长(约100kD)或截短(约90kD)的形式在细胞表面表达。重组全长CD34在CHO-K1细胞中表达时，分子量约为105kD。M1白血病细胞表达的全长CD34具有较高的表观分子量(110kD)。这些结果表明，不同细胞类型表达的CD34之间存在糖基化差异。全长形式，而不是截短形式，是一种对12-0-十四烷基佛波醇13-乙酸酯处理反应过度磷酸化的磷蛋白，这表明两种形式之间存在潜在的功能差异。选择3%表达最高的CD34+骨髓细胞作为造血祖细胞，形成集落形成单位--脾(CFU-S)、粒-巨噬细胞集落形成单位(CFU-GM)和爆裂形成单位--红系。用这些细胞移植致死照射的小鼠，显示出短期和长期的再繁殖能力，表明这个群体既有功能的造血祖细胞，也有假定的干细胞。这些抗体对小鼠造血干细胞的选择应该是有用的。
CD34 is expressed on human hematopoietic stem and progenitor cells, and its clinical usefulness for the purification of stem cells has been well established. However, a similar pattern of expression for murine CD34 (mCD34) has not yet been determined. Two polyclonal anti-mCD34 antibodies that specifically recognize both endogenous and recombinant murine CD34 were developed to characterize the mCD34 protein and to determine its pattern of expression on murine cell lines and hematopoietic progenitor cells. Fluorescence-activated cell sorter analysis showed that mCD34 is expressed on NIH/3T3 embryonic fibroblasts, PA6 stromal cells, embryonic stem cells, M1 leukemia cells, and a subpopulation of normal bone marrow cells. Murine CD34 was found to be a glycoprotein expressed on the cell surface as either a full-length (approximately 100 kD) or truncated (approximately 90 kD) protein in NIH/3T3 and PA6 cells. Recombinant full-length CD34, when expressed in the CHO-K1 cell line, had a molecular weight of approximately 105 kD. Full-length CD34 expressed on M1 leukemia cells, had a higher apparent molecular weight (110 kD). These results suggest that there are glycosylation differences between CD34 expressed by different cell types. The full-length form, but not the truncated form, is a phosphoprotein that is hyperphosphorylated in response to 12-0-Tetradecanoyl phorbol 13-acetate treatment, suggesting potential functional differences between the two forms. Selection of the 3% highest-expressing CD34+ bone marrow cells enriched for the hematopoietic precursors that form colony-forming unit-spleen (CFU-S), CFU-granulocyte-macrophage, and burst-forming unit-erythroid. Transplantation of lethally irradiated mice with these cells demonstrated both short- and long-term repopulating ability, indicating that this population contains both functional hematopoietic progenitors and the putative stem cell. These antibodies should be useful to select for murine hematopoietic stem cells.