Characterization of murine CD34, a marker for hematopoietic progenitor and stem cells

Abstract

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.

Low oxygen tension favored expansion and hematopoietic reconstitution of CD34+ CD38− cells expanded from human cord blood‐derived CD34+ Cells

“…Human HSPCs are enriched in CD34 + cells. In fact, most colony-forming cells (CFCs) are found in the CD34 + cell fraction [18][19][20] and the most primitive self-renewing HSCs with long-term reconstituting ability reside in CD34 + CD38cells [21]. Therefore, in this study, the effects of oxygen tension on the ex vivo expansion characteristics of CD34 + cells derived from human UCB, including the ex vivo expansion of total cells, CD34 + cells, colony-forming cells (CFCs) and CD34 + CD38cells were analyzed.…”

ABC Transporters as Phenotypic Markers and Functional Regulators of Stem Cells

“…It was first demonstrated 10 years ago by Chaudhary and Roninson, that the P-gp is highly expressed on CD34 + hematopoietic cells [15], suggesting that efflux pump activity could be responsible for the low retention of Rho123 in primitive subsets of cells. The CD34 gene codes for a transmembrane cell surface phosphoprotein that has been generally accepted as being a stem cell marker based on engraftment following bone marrow transplantation into baboons [16], humans [17], and mice [18]. However, in recent years, this has become a point of controversy in the field.…”

Generation of a conditionally immortalized myeloid progenitor cell line requiring the presence of both interleukin‐3 and stem cell factor to survive and proliferate

“…The H-2K b -tsA58 cells showed the expression of ER-MP 58, ER-MP 12 and ER-MP 20, consistent with a primitive myeloid phenotype (Fig 3). Subsets of the population expressed the Sca-1 antigen (Ly-6C), a phosphatidylinositol-anchored protein that is expressed on stem and progenitor cells (Van de Rijn et al, 1989;Miles et al, 1997) and CD34, a marker for primitive haemopoietic progenitor cells (Krause et al, 1994).…”

“…Flow cytometric analysis of cells after culture in IL-3 and SCF was used to determine the phenotype of the MIST-D cell line and changes stimulated by the addition of M-CSF or GM-CSF. MIST-D cells grown in the presence of low or high concentrations of IL-3 plus SCF and at the permissive temperature for the A58 oncogene expressed the primitive markers Sca-1 ( Van de Rijn et al, 1989;Miles et al, 1997), CD34 (Krause et al, 1994) and ER-MP 12 and were predominantly ER-MP 58 + . ER-MP 58 is an early marker for cells committed to the myeloid lineage (de-Bruijn et al, Results are shown for 1-4 d in culture and are expressed as a percentage of the response of the cells to optimal concentrations of mIL-3 + SCF on each of these days at 32°C.…”