Contribution of STAT3 to Activation of Survivin by Granulocyte-Macrophage Colony-Stimulating Factor in CD34+ Cells.

Abstract Granulocyte-macrophage colony-stimulating factor (GM-CSF) has been shown to specifically stimulate proliferation and differentiation of CD34+ hematopoietic progenitor cells. Although STAT3 was thought to be essential for the transduction of GM-CSF-induced cell proliferation, the downstream signaling mediated by STAT3 to support cell proliferation and growth has not been completely understood. Because the inhibitor of apoptosis protein (IAP) survivin is believed to regulate cell proliferation and survival via its anti-apoptotic function, we chose to study the link between STAT3 signaling and survivin expression in CD34+ cells. We constructed plasmids containing the survivin promoter sequence and performed luciferase reporter assay in CD34+ KG-1 cells stimulated with GM-CSF. These experiments showed that GM-CSF stimulated survivin promoter activity. Chromatin immunoprecipitation (CHIP) and electrophoretic mobility shift assay (EMSA) revealed that STAT3 binds to the core survivin promoter containing a STAT response element (SRE) TT(N)5AA at sites −264 to −256. Mutation or deletion of this SRE completely abolished the effect of GM-CSF on survivin promoter activity. Furthermore, specific JAK inhibitor and STAT3 siRNA inhibited GM-CSF-induced survivin promoter activity and survivin expression. Inhibition of survivin by STAT3 siRNA or by withdrawal of GM-CSF in a GM-CSF-dependent CD34+ line TF-1 resulted in decreased cell growth and induction of apoptosis. These results suggest that the anti-apoptotic protein survivin is a transcriptional target of STAT3, and that GM-CSF stimulated-CD34+ cell proliferation is regulated by the JAK/STAT3/survivin signaling pathway.

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Granulocyte-macrophage colony-stimulating factor, phorbol ester, and sodium butyrate induce the CD11c integrin gene promoter activity during myeloid cell differentiation

Blood ◽

10.1182/blood.v86.10.3715.bloodjournal86103715 ◽

1995 ◽

Vol 86 (10) ◽

pp. 3715-3724 ◽

Cited By ~ 13

Author(s):

MA Rubio ◽

C Lopez-Rodriguez ◽

A Nueda ◽

P Aller ◽

AL Armesilla ◽

...

Keyword(s):

Phorbol Ester ◽

Promoter Activity ◽

Gene Promoter ◽

Proximal Region ◽

Myeloid Differentiation ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

To analyze the activity of the CD11c promoter during myeloid differentiation without the limitations of transient expression systems, we have stably transfected the myeloid U937 cell line with the pCD11C361-Luc plasmid, in which the expression of the firefly luciferase cDNA is driven by the CD11c promoter region -361/+43, previously shown to confer myeloid specificity to reporter genes. The stable transfectants (U937-C361) retained the ability to differentiate in response to phorbol-ester (PMA), sodium butyrate (SB), granulocyte- macrophage colony-stimulating factor (GM-CSF), and other differentiating agents. U937-C361 differentiation correlated with increased cellular luciferase levels, showing the inducibility of the CD11c promoter during myeloid differentiation and establishing the U937- C361 cells as a suitable system for studying the myeloid differentiation-inducing capacity of cytokines, growth, factors, and other biological response modifiers. Unexpectedly, the inducibility of the CD11c gene promoter showed distinct kinetics and magnitude on the PMA-, SB-, GM-CSF-triggered differentiation. Moreover, SB synergized with either PMA or GM-CSF in enhancing both the CD11c promoter activity and the cell surface expression of p150,95 on differentiating U937 cells. Furthermore, we showed the existence of a c-Myb-binding site at - 85, the importance of the -99/-61 region in the CD11c promoter inducibility during PMA- or SB-triggered differentiation, and the dependency of the GM-CSF and PMA responsiveness of the CD11c promoter on an intact AP-1-binding site located at -60. These results, together with the lack of functional effect of mutations disrupting the Sp1-and Myb-binding sites within the proximal region of the CD11c promoter, indicate that the myeloid differentiation pathways indicated by SB and phorbol esters (or GM-CSF) activate a distinct set of transcription factors and show that the myeloid differentiation-inducibility of the CD11c gene maps to the -99/-53 proximal region of the promoter.

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Recombinant human stem cell factor enhances the formation of colonies by CD34+ and CD34+lin- cells, and the generation of colony-forming cell progeny from CD34+lin- cells cultured with interleukin-3, granulocyte colony-stimulating factor, or granulocyte-macrophage colony-stimulating factor

Blood ◽

10.1182/blood.v77.11.2316.bloodjournal77112316 ◽

1991 ◽

Vol 77 (11) ◽

pp. 2316-2321 ◽

Cited By ~ 4

Author(s):

ID Bernstein ◽

RG Andrews ◽

KM Zsebo

Keyword(s):

Stem Cell Factor ◽

Liquid Culture ◽

Granulocyte Colony Stimulating Factor ◽

Human Stem Cell ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Cd34 Cells ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

We tested the ability of recombinant human stem cell factor (SCF) to stimulate isolated marrow precursor cells to form colonies in semisolid media and to generate colony-forming cells (CFC) in liquid culture. SCF, in combination with interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), or granulocyte colony-stimulating factor (G-CSF) caused CD34+ cells to form increased numbers of granulocyte-macrophage colonies (CFU-GM), and to form macroscopic erythroid burst-forming units (BFU-E) in the presence of IL-3, erythropoietin (Epo), and SCF. We tested isolated CD34+lin- cells, a minor subset of CD34+ cells that did not display antigens associated with lymphoid or myeloid lineages, and CD34+lin+ cells, which contain the vast majority of CFC, and found that the enhanced colony growth was most dramatic within the CD34+lin- population. CD34+lin- cells cultured in liquid medium containing SCF combined with IL-3, GM-CSF, or G-CSF gave rise to increased numbers of CFC. Maximal numbers of CFU-GM were generated from CD34+lin- cells after 7 to 21 days of culture, and required the presence of SCF from the initiation of liquid culture. The addition of SCF to IL-3 and/or G-CSF in cultures of single CD34+lin- cells resulted in increased numbers of CFC due to the proliferation of otherwise quiescent precursors and an increase in the numbers of CFC generated from individual precursors. These studies demonstrate the potent synergistic interaction between SCF and other hematopoietic growth factors on a highly immature population of CD34+lin- precursor cells.

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Harvesting and enrichment of hematopoietic progenitor cells mobilized into the peripheral blood of normal donors by granulocyte-macrophage colony-stimulating factor (GM-CSF) or G-CSF: potential role in allogeneic marrow transplantation

Blood ◽

10.1182/blood.v85.1.275.bloodjournal851275 ◽

1995 ◽

Vol 85 (1) ◽

pp. 275-282 ◽

Cited By ~ 207

Author(s):

TA Lane ◽

P Law ◽

M Maruyama ◽

D Young ◽

J Burgess ◽

...

Keyword(s):

Progenitor Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Allogeneic Transplantation ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Cd34 Cells ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

To explore the use of stem/progenitor cells from peripheral blood (PB) for allogeneic transplantation, we have studied the mobilization of progenitor cells in normal donors by growth factors. Normal subjects were administered either granulocyte-macrophage colony-stimulating factor (GM-CSF) at 10 micrograms/kg/d, or G-CSF at 10 micrograms/kg/d, or a combination of G- and GM-CSF at 5 micrograms/kg/d each, administered subcutaneously for 4 days, followed by leukapheresis on day 5. Mononuclear cells expressing CD34 (CD34+ cells) were selectively enriched by affinity labeling using Dynal paramagnetic microspheres (Baxter Isolex; Baxter Healthcare Corp, Santa Ana, CA). The baseline CD34+ cells in peripheral blood before mobilization was 0.07% +/- 0.05% (1.6 +/- 0.7/microL; n = 18). On the fifth day after stimulation (24 hours after the fourth dose), the CD34+ cells were 0.99% +/- 0.40% (61 +/- 14/microL) for the 8 subjects treated with G-CSF, 0.25% +/- 0.25% (3 +/- 3/microL, both P < .01 v G-CSF) for the 5 subjects administered GM-CSF, and for the 5 subjects treated with G- and GM-CSF, 0.65% +/- 0.28% (41 +/- 18/microL, P < .5 v GM-CSF). Parallel to this increase in CD34+ cells, clonogenic assays showed a corresponding increase in CFU- GM and BFU-E. The total number of CD34+ cells collected from the G-CSF group during a 3-hour apheresis was 119 +/- 65 x 10(6) and was not significantly different from that collected from the group treated with G- and GM-CSF (101 +/- 35 x 10(6) cells), but both were greater than that from the group treated with GM-CSF (12.6 +/- 6.1 x 10(6); P < .01 for both comparisons). Analysis of the CD34+ subsets showed that a significantly higher percentage of cells with the CD34+/CD38- phenotype is found after mobilization with G- and GM-CSF. In the G-CSF group, immunomagnetic selection of CD34+ cells permitted the enrichment of the CD34+ cells in the apheresis product to 81% +/- 11%, with a 48% +/- 12% yield and to a purity of 77% +/- 21% with a 51% +/- 15% recovery in the G- and GM-CSF group. T cells were depleted from a mean of 4.5 +/- 2.0 x 10(9) to 4.3 +/- 5.2 x 10(6) after selection, representing 99.9% depletion. We conclude that it is feasible to collect sufficient numbers of PB progenitor cells from normal donors with one to two leukapheresis procedures for allogeneic transplantation.(ABSTRACT TRUNCATED AT 250 WORDS)

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Positive and negative regulation of granulocyte-macrophage colony- stimulating factor promoter activity by AML1-related transcription factor, PEBP2

Blood ◽

10.1182/blood.v86.2.607.bloodjournal862607 ◽

1995 ◽

Vol 86 (2) ◽

pp. 607-616 ◽

Cited By ~ 119

Author(s):

A Takahashi ◽

M Satake ◽

Y Yamaguchi-Iwai ◽

SC Bae ◽

J Lu ◽

...

Keyword(s):

Transcription Factor ◽

Promoter Activity ◽

Binding Protein ◽

Consensus Sequence ◽

Jurkat Cells ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

The granulocyte-macrophage colony-stimulating factor (GM-CSF) gene promoter contains a consensus sequence for the polyomavirus enhancer binding-protein 2 (PEBP2) transcription factor, which consists of alpha and beta subunits. There are at least two genes, alpha A and alpha B, encoding the alpha subunit. alpha B is the mouse homologue of human AML1 gene detected at the breakpoints of t(8;21) and t(3;21) myeloid leukemias. We examined alpha A1 (an alpha A-gene product) and alpha B1 and alpha B2 (two alpha B-encoded isomers) for their effects on the GM- CSF promoter. PEBP2 alpha A1, alpha B1, and alpha B2 proteins bound the PEBP2 site within the mouse GM-CSF promoter. PEBP2 alpha A1 and alpha B1 enhanced the expression of the GM-CSF promoter-driven reporter plasmid in unstimulated and 12-O-tetradecanoylphorbol 13- acetate/phytohemagglutinin-stimulated human Jurkat T cells. In contrast, the promoter activity was suppressed by alpha B2. Coexpression of alpha B1 and alpha B2 showed that the promoter activity could be determined by the alpha B1/alpha B2 ratio. Jurkat cell extract contained PEBP2 site-binding protein(s) that cross-reacted with antimouse alpha A1 antibodies. Northern blot analysis indicated the expression of human PEBP2 alpha A, alpha B (AML1), and beta genes in Jurkat cells. Although further studies are required to determine the precise role of PEBP2 in the GM-CSF promoter activity, the present findings suggested the importance of the relative ratio of different PEBP2 isoforms in regulating the levels of the promoter activity.

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PU.1 (Spi-1) and C/EBP alpha regulate expression of the granulocyte-macrophage colony-stimulating factor receptor alpha gene.

Molecular and Cellular Biology ◽

10.1128/mcb.15.10.5830 ◽

1995 ◽

Vol 15 (10) ◽

pp. 5830-5845 ◽

Cited By ~ 209

Author(s):

S Hohaus ◽

M S Petrovick ◽

M T Voso ◽

Z Sun ◽

D E Zhang ◽

...

Keyword(s):

B Cell ◽

Promoter Activity ◽

Colony Stimulating Factor ◽

Receptor Alpha ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Alpha Gene ◽

Myelomonocytic Cells ◽

Macrophage Colony ◽

Stimulating Factor

Growth factor receptors play an important role in hematopoiesis. In order to further understand the mechanisms directing the expression of these key regulators of hematopoiesis, we initiated a study investigating the transcription factors activating the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor alpha gene. Here, we demonstrate that the human GM-CSF receptor alpha promoter directs reporter gene activity in a tissue-specific fashion in myelomonocytic cells, which correlates with its expression pattern as analyzed by reverse transcription PCR. The GM-CSF receptor alpha promoter contains an important functional site between positions -53 and -41 as identified by deletion analysis of reporter constructs. We show that the myeloid and B cell transcription factor PU.1 binds specifically to this site. Furthermore, we demonstrate that a CCAAT site located upstream of the PU.1 site between positions -70 and -54 is involved in positive-negative regulation of the GM-CSF receptor alpha promoter activity. C/EBP alpha is the major CCAAT/enhancer-binding protein (C/EBP) form binding to this site in nuclear extracts of U937 cells. Point mutations of either the PU.1 site or the C/EBP site that abolish the binding of the respective factors result in a significant decrease of GM-CSF receptor alpha promoter activity in myelomonocytic cells only. Furthermore, we demonstrate that in myeloid and B cell extracts, PU.1 forms a novel, specific, more slowly migrating complex (PU-SF) when binding the GM-CSF receptor alpha promoter PU.1 site. This is the first demonstration of a specific interaction with PU.1 on a myeloid PU.1 binding site. The novel complex is distinct from that described previously as binding to B cell enhancer sites and can be formed by addition of PU.1 to extracts from certain nonmyeloid cell types which do not express PU.1, including T cells and epithelial cells, but not from erythroid cells. Furthermore, we demonstrate that the PU-SF complex binds to PU.1 sites found on a number of myeloid promoters, and its formation requires an intact PU.1 site adjacent to a single-stranded region. Expression of PU.1 in nonmyeloid cells can activate the GM-CSF receptor alpha promoter. Deletion of the amino-terminal region of PU.1 results in a failure to form the PU-SF complex and in a concomitant loss of transactivation, suggesting that formation of the PU-SF complex is of functional importance for the activity of the GM-CSF receptor alpha promoter. Finally, we demonstrate that C/EBP alpha can also active the GM-CSF receptor alpha promoter in nonmyeloid cells. These results suggest that PU.1 and C/EBP alpha direct the cell-type-specific expression of GM-CSF receptor alpha, further establish the role of PU.1 as a key regulator of hematopoiesis, and point to C/EBP alpha as an additional important factor in this process.

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The repeated sequence CATT(A/T) is required for granulocyte-macrophage colony-stimulating factor promoter activity

Molecular and Cellular Biology ◽

10.1128/mcb.10.11.6084-6088.1990 ◽

1990 ◽

Vol 10 (11) ◽

pp. 6084-6088

Author(s):

S Nimer ◽

J Fraser ◽

J Richards ◽

M Lynch ◽

J Gasson

Keyword(s):

T Lymphocytes ◽

Promoter Activity ◽

Repeated Sequence ◽

Inducible Promoter ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Flanking Sequences ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

The hematopoietic growth factor GM-CSF (granulocyte-macrophage colony-stimulating factor) is expressed by activated but not resting T lymphocytes. Previously, we localized GM-CSF-inducible promoter activity to a 90-bp region of GM-CSF 5'-flanking sequences extending from bp -53 to +37. To more precisely identify the GM-CSF DNA sequences required for inducible promoter activity in T lymphocytes, we have performed mutagenesis within a region of GM-CSF 5'-flanking sequences (bp -57 to -24) that contains the repeated sequence CATT(A/T). Mutations that do not alter the repeated CATT(A/T) sequence do not eliminate inducible promoter activity, whereas mutation or deletion of either of the CATT(A/T) repeats eliminates all inducible promoter activity in T-cell lines and in primary human T lymphocytes. Thus, both copies of the direct repeat CATT(A/T) are required for mitogen-inducible expression of GM-CSF in T cells.

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Cloning and Characterization of the Human Interleukin-3 (IL-3)/IL-5/ Granulocyte-Macrophage Colony-Stimulating Factor Receptor βc Gene: Regulation by Ets Family Members

Blood ◽

10.1182/blood.v92.10.3636.422k45_3636_3646 ◽

1998 ◽

Vol 92 (10) ◽

pp. 3636-3646 ◽

Cited By ~ 1

Author(s):

Thamar B. van Dijk ◽

Belinda Baltus ◽

Eric Caldenhoven ◽

Hiroshi Handa ◽

Jan A.M. Raaijmakers ◽

...

Keyword(s):

Gene Regulation ◽

Promoter Activity ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Ets Family ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Β Chain ◽

Stimulating Factor

High-affinity receptors for interleukin-3 (IL-3), IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) are composed of two distinct subunits, a ligand-specific  chain and a common β chain (βc). Whereas the mouse has two homologous β subunits (βc and βIL-3), in humans, only a single β chain is identified. We describe here the isolation and characterization of the gene encoding the human IL-3/IL-5/GM-CSF receptor β subunit. The gene spans about 25 kb and is divided into 14 exons, a structure very similar to that of the murine βc/βIL-3 genes. Surprisingly, we also found the remnants of a second βc chain gene directly downstream of βc. We identified a functional promoter that is active in the myeloid cell lines U937 and HL-60, but not in HeLa cells. The proximal promoter region, located from −103 to +33 bp, contains two GGAA consensus binding sites for members of the Ets family. Single mutation of those sites reduces promoter activity by 70% to 90%. The 5′ element specifically binds PU.1, whereas the 3′ element binds a yet-unidentified protein. These findings, together with the observation that cotransfection of PU.1 and other Ets family members enhances βc promoter activity in fibroblasts, reinforce the notion that GGAA elements play an important role in myeloid-specific gene regulation.

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A comparative study of the phenotype and proliferative capacity of peripheral blood (PB) CD34+ cells mobilized by four different protocols and those of steady-phase PB and bone marrow CD34+ cells

Blood ◽

10.1182/blood.v84.9.2930.2930 ◽

1994 ◽

Vol 84 (9) ◽

pp. 2930-2939 ◽

Cited By ~ 135

Author(s):

LB To ◽

DN Haylock ◽

T Dowse ◽

PJ Simmons ◽

S Trimboli ◽

...

Keyword(s):

Bone Marrow ◽

Steady State ◽

Peripheral Blood ◽

Proliferative Capacity ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Cd34 Cells ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

Abstract Peripheral blood (PB) CD34+ cells from four commonly used mobilization protocols were studied to compare their phenotype and proliferative capacity with steady-state PB or bone marrow (BM) CD34+ cells. Mobilized PB CD34+ cells were collected during hematopoietic recovery after myelosuppressive chemotherapy with or without granulocyte- macrophage colony-stimulating factor (GM-CSF) or granulocyte colony- stimulating factor (G-CSF) or during G-CSF administration alone. The expression of activation and lineage-associated markers and c-kit gene product were studied by flow cytometry. Proliferative capacity was measured by generation of nascent myeloid progenitor cells (granulocyte- macrophage colony-stimulating factor; CFU-GM) and nucleated cells in a stroma-free liquid culture stimulated by a combination of six hematopoietic growth factors (interleukin-1 (IL-1), IL-3, IL-6, GM-CSF, G-CSF, and stem cell factor). G-CSF-mobilized CD34+ cells have the highest percentage of CD38- cells (P < .0081), but otherwise, CD34+ cells from different mobilization protocols were similar to one another in their phenotype and proliferative capacity. The spectrum of primitive and mature myeloid progenitors in mobilized PB CD34+ cells was similar to their steady-state counterparts, but the percentages of CD34+ cells expressing CD10 or CD19 were lower (P < .0028). Although steady-state PB and chemotherapy-mobilized CD34+ cells generated fewer CFU-GM at day 21 than G-CSF-mobilized and steady-state BM CD34+ cells (P < .0449), the generation of nucleated cells and CFU-GM were otherwise comparable. The presence of increased or comparable numbers of hematopoietic progenitors within PB collections with equivalent proliferative capacity to BM CD34+ cells is not unexpected given the rapid and complete hematopoietic reconstitution observed with mobilized PB. However, all four types of mobilized PB CD34+ cells are different from steady-state BM CD34+ cells in that they express less c-kit (P < .0002) and CD71 (P < .04) and retain less rhodamine 123 (P < .0001). These observations are novel and suggest that different mobilization protocols may act via similar pathways involving the down-regulation of c-kit and may be independent of cell-cycle status.

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The repeated sequence CATT(A/T) is required for granulocyte-macrophage colony-stimulating factor promoter activity.

Molecular and Cellular Biology ◽

10.1128/mcb.10.11.6084 ◽

1990 ◽

Vol 10 (11) ◽

pp. 6084-6088 ◽

Cited By ~ 22

Author(s):

S Nimer ◽

J Fraser ◽

J Richards ◽

M Lynch ◽

J Gasson

Keyword(s):

T Lymphocytes ◽

Promoter Activity ◽

Repeated Sequence ◽

Inducible Promoter ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Flanking Sequences ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

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Distribution of receptors for granulocyte-macrophage colony-stimulating factor on immature CD34+ bone marrow cells, differentiating monomyeloid progenitors, and mature blood cell subsets

Blood ◽

10.1182/blood.v84.3.764.bloodjournal843764 ◽

1994 ◽

Vol 84 (3) ◽

pp. 764-774 ◽

Cited By ~ 42

Author(s):

AW Wognum ◽

Y Westerman ◽

TP Visser ◽

G Wagemaker

Keyword(s):

Bone Marrow ◽

Receptor Expression ◽

Colony Stimulating Factor ◽

Short Term ◽

Gm Csf ◽

Cd34 Cells ◽

Macrophage Colony Stimulating Factor ◽

Low Levels ◽

Macrophage Colony ◽

Stimulating Factor

Biotin-labeled granulocyte-macrophage colony-stimulating factor (GM- CSF), in combination with phycoerythrin-conjugated streptavidin, enabled flow cytometric analysis of specific cell-surface GM-CSF receptors on rhesus monkey bone marrow (BM) and peripheral blood (PB) cells. GM-CSF receptors were readily detected on PB monocytes and neutrophils, but not on lymphocytes. In BM, GM-CSF receptors were identified on monocyte and neutrophil precursors and on subsets of cells that expressed the CD34 antigen. CD34+ cells with high GM-CSF- receptor expression coexpressed high levels of the class II major histocompatibility antigen RhLA-DR, whereas CD34+/RhLA-DRlow cells, which represent developmentally earlier cells, were either GM-CSF- receptor negative or expressed GM-CSF receptors at very low levels. The fluorescence histogram of CD34bright/RhLA-DRdull cells stained with biotin-GM-CSF showed that at least a fraction of these cells expressed low levels of GM-CSF receptors. CD34+ cells with high GM-CSF-receptor expression, purified by cell sorting, did not form colonies in culture or proliferate in response to GM-CSF. Instead, GM-CSF stimulation resulted in terminal differentiation into adherent cells, showing that these cells represented monocyte precursors. A distinct subset of CD34+ cells expressed GM-CSF receptors at low-to-intermediate levels and proliferated strongly in the presence of GM-CSF during short-term culture, but produced very few erythroid or monomyeloid colonies after longer culture periods. Most colony-forming cells, also those responsive to GM-CSF alone, were recovered in the subset of CD34+ cells on which GM-CSF receptors were virtually undetectable. These cells showed weaker proliferation in short-term proliferation assays than the CD34+/GM-CSF-receptor-intermediate cells, consistent with an immature phenotype. The results show that GM-CSF-receptor expression is initiated in a subset of immature, CD34bright/RhLA-DRdull cells and is progressively increased during differentiation into mature granulocytes and monocytes. The method used provides a new way to deplete developmentally early CD34+ cell of differentiating granulocyte and monocyte precursor cells.

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