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Enforced granulocyte/macrophage colony-stimulating factor signals do not support lymphopoiesis, but instruct lymphoid to myelomonocytic lineage conversion.

Iwasaki-Arai J, Iwasaki H, Miyamoto T, Watanabe S, Akashi K - J. Exp. Med. (2003)

Bottom Line: We evaluated the effects of ectopic granulocyte/macrophage colony-stimulating factor (GM-CSF) signals on hematopoietic commitment and differentiation.Strikingly, >50% hGM-CSFR+ common lymphoid progenitors (CLPs) and >20% hGM-CSFR+ pro-T cells gave rise to granulocyte, monocyte, and/or myeloid dendritic cells, but not MegE lineage cells in the presence of hGM-CSF.Thus, hGM-CSF transduces permissive signals for myeloerythroid differentiation, whereas it transmits potent instructive signals for the GM differentiation to CLPs and early T cell progenitors.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA.

ABSTRACT
We evaluated the effects of ectopic granulocyte/macrophage colony-stimulating factor (GM-CSF) signals on hematopoietic commitment and differentiation. Lineage-restricted progenitors purified from mice with the ubiquitous transgenic human GM-CSF receptor (hGM-CSFR) were used for the analysis. In cultures with hGM-CSF alone, hGM-CSFR-expressing (hGM-CSFR+) granulocyte/monocyte progenitors (GMPs) and megakaryocyte/erythrocyte progenitors (MEPs) exclusively gave rise to granulocyte/monocyte (GM) and megakaryocyte/erythroid (MegE) colonies, respectively, providing formal proof that GM-CSF signals support the GM and MegE lineage differentiation without affecting the physiological myeloid fate. hGM-CSFR transgenic mice were crossed with mice deficient in interleukin (IL)-7, an essential cytokine for T and B cell development. Administration of hGM-CSF in these mice could not restore T or B lymphopoiesis, indicating that enforced GM-CSF signals cannot substitute for IL-7 to promote lymphopoiesis. Strikingly, >50% hGM-CSFR+ common lymphoid progenitors (CLPs) and >20% hGM-CSFR+ pro-T cells gave rise to granulocyte, monocyte, and/or myeloid dendritic cells, but not MegE lineage cells in the presence of hGM-CSF. Injection of hGM-CSF into mice transplanted with hGM-CSFR+ CLPs blocked their lymphoid differentiation, but induced development of GM cells in vivo. Thus, hGM-CSF transduces permissive signals for myeloerythroid differentiation, whereas it transmits potent instructive signals for the GM differentiation to CLPs and early T cell progenitors. These data suggest that a majority of CLPs and a fraction of pro-T cells possess plasticity for myelomonocytic differentiation that can be activated by ectopic GM-CSF signals, supporting the hypothesis that the down-regulation of GM-CSFR is a critical event in producing cells with a lymphoid-restricted lineage potential.

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Cytokine receptor expression in hematopoietic branch points. (A) Lineage relationships of prospectively purified lineage-restricted progenitors in steady-state hematopoiesis. LT-HSC, long-term HSCs (CD34− Lin− Sca-1+ c-Kit+); ST-HSC, short-term HSCs (CD34+ Lin− Sca-1+ c-Kit+). (B) Results of RT-PCR analyses of cytokine receptors targeted for 250 cells per progenitor species. The symbols under each lane depict relative amounts of mRNA in each population compared with control cDNA (2 × 105 cells, bands not shown) by the ratio pixel density units of target cDNA/pixel density units of control cDNA. −, <0.1; ±, 0.1–0.5; +, 0.5–1.5; ++, >1.5. (C) Expression of transgenic hGM-CSFRα in each purified population. Shaded and open lines show the staining of target populations by anti–hGM-CSFRα and control IgG antibodies, respectively. A majority of each stem and progenitor fraction express hGM-CSFRα.
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fig1: Cytokine receptor expression in hematopoietic branch points. (A) Lineage relationships of prospectively purified lineage-restricted progenitors in steady-state hematopoiesis. LT-HSC, long-term HSCs (CD34− Lin− Sca-1+ c-Kit+); ST-HSC, short-term HSCs (CD34+ Lin− Sca-1+ c-Kit+). (B) Results of RT-PCR analyses of cytokine receptors targeted for 250 cells per progenitor species. The symbols under each lane depict relative amounts of mRNA in each population compared with control cDNA (2 × 105 cells, bands not shown) by the ratio pixel density units of target cDNA/pixel density units of control cDNA. −, <0.1; ±, 0.1–0.5; +, 0.5–1.5; ++, >1.5. (C) Expression of transgenic hGM-CSFRα in each purified population. Shaded and open lines show the staining of target populations by anti–hGM-CSFRα and control IgG antibodies, respectively. A majority of each stem and progenitor fraction express hGM-CSFRα.

Mentions: Fig. 1 A depicts the hematopoietic developmental tree and the progenitors used in this study. We analyzed the distribution of mGM-CSFRα, IL-3Rα, EpoR, and IL-7Rα by RT-PCR in murine stem and progenitor populations (23, 24). In this analysis, we separated Lin− Sca-1+ c-Kit+ HSC population (35) into CD34− Sca-1+ c-Kit+ HSCs that possess long-term reconstituting activity at the single cell level (28), and their descendants, CD34+ Sca-1+ c-Kit+ short-term HSCs (28, 29). As shown in Fig. 1 B, the expression of mGM-CSFRα and IL-3Rα is initiated at the short-term HSC stage, and gradually up-regulated along the myelomonocytic pathway including CMPs and GMPs, whereas they are down-regulated in CLPs and MEPs. FACS® analysis showed that murine βc is expressed in all hematopoietic cells (not depicted). Thus, the expression of βc-related cytokine receptors such as GM-CSFR and IL-3R are progressively up-regulated along the myelomonocytic differentiation pathway, but down-regulated if cells commit to the lymphoid or the MegE lineages. EpoR is up-regulated during the development from long-term HSCs to MEPs, but down-regulated in GMPs or CLPs. In contrast, lymphoid-related IL-7Rα is exclusively expressed in CLPs, but not in HSCs or myeloid progenitors. In H-2Ld-hGM-CSFRα/βc double transgenic mice (8), hGM-CSFRα/βc transgenes are driven by the MHC class I promoter, and most hematopoietic cells express hGM-CSFR. As shown in Fig. 1 C, 85–96% of cells in each progenitor population were stained with monoclonal anti–hGM-CSFRα antibodies. Because hGM-CSFR does not react with endogenous mGM-CSF, cells in H-2Ld-hGM-CSFRα/βc double transgenic mice never receive hGM-CSF signals in vivo in the absence of hGM-CSF (8). Thus, FACS® analysis showed that the frequency of each myeloid and lymphoid progenitor population in hGM-CSFR transgenic mice was identical to that of wild-type mice (unpublished data).


Enforced granulocyte/macrophage colony-stimulating factor signals do not support lymphopoiesis, but instruct lymphoid to myelomonocytic lineage conversion.

Iwasaki-Arai J, Iwasaki H, Miyamoto T, Watanabe S, Akashi K - J. Exp. Med. (2003)

Cytokine receptor expression in hematopoietic branch points. (A) Lineage relationships of prospectively purified lineage-restricted progenitors in steady-state hematopoiesis. LT-HSC, long-term HSCs (CD34− Lin− Sca-1+ c-Kit+); ST-HSC, short-term HSCs (CD34+ Lin− Sca-1+ c-Kit+). (B) Results of RT-PCR analyses of cytokine receptors targeted for 250 cells per progenitor species. The symbols under each lane depict relative amounts of mRNA in each population compared with control cDNA (2 × 105 cells, bands not shown) by the ratio pixel density units of target cDNA/pixel density units of control cDNA. −, <0.1; ±, 0.1–0.5; +, 0.5–1.5; ++, >1.5. (C) Expression of transgenic hGM-CSFRα in each purified population. Shaded and open lines show the staining of target populations by anti–hGM-CSFRα and control IgG antibodies, respectively. A majority of each stem and progenitor fraction express hGM-CSFRα.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2193786&req=5

fig1: Cytokine receptor expression in hematopoietic branch points. (A) Lineage relationships of prospectively purified lineage-restricted progenitors in steady-state hematopoiesis. LT-HSC, long-term HSCs (CD34− Lin− Sca-1+ c-Kit+); ST-HSC, short-term HSCs (CD34+ Lin− Sca-1+ c-Kit+). (B) Results of RT-PCR analyses of cytokine receptors targeted for 250 cells per progenitor species. The symbols under each lane depict relative amounts of mRNA in each population compared with control cDNA (2 × 105 cells, bands not shown) by the ratio pixel density units of target cDNA/pixel density units of control cDNA. −, <0.1; ±, 0.1–0.5; +, 0.5–1.5; ++, >1.5. (C) Expression of transgenic hGM-CSFRα in each purified population. Shaded and open lines show the staining of target populations by anti–hGM-CSFRα and control IgG antibodies, respectively. A majority of each stem and progenitor fraction express hGM-CSFRα.
Mentions: Fig. 1 A depicts the hematopoietic developmental tree and the progenitors used in this study. We analyzed the distribution of mGM-CSFRα, IL-3Rα, EpoR, and IL-7Rα by RT-PCR in murine stem and progenitor populations (23, 24). In this analysis, we separated Lin− Sca-1+ c-Kit+ HSC population (35) into CD34− Sca-1+ c-Kit+ HSCs that possess long-term reconstituting activity at the single cell level (28), and their descendants, CD34+ Sca-1+ c-Kit+ short-term HSCs (28, 29). As shown in Fig. 1 B, the expression of mGM-CSFRα and IL-3Rα is initiated at the short-term HSC stage, and gradually up-regulated along the myelomonocytic pathway including CMPs and GMPs, whereas they are down-regulated in CLPs and MEPs. FACS® analysis showed that murine βc is expressed in all hematopoietic cells (not depicted). Thus, the expression of βc-related cytokine receptors such as GM-CSFR and IL-3R are progressively up-regulated along the myelomonocytic differentiation pathway, but down-regulated if cells commit to the lymphoid or the MegE lineages. EpoR is up-regulated during the development from long-term HSCs to MEPs, but down-regulated in GMPs or CLPs. In contrast, lymphoid-related IL-7Rα is exclusively expressed in CLPs, but not in HSCs or myeloid progenitors. In H-2Ld-hGM-CSFRα/βc double transgenic mice (8), hGM-CSFRα/βc transgenes are driven by the MHC class I promoter, and most hematopoietic cells express hGM-CSFR. As shown in Fig. 1 C, 85–96% of cells in each progenitor population were stained with monoclonal anti–hGM-CSFRα antibodies. Because hGM-CSFR does not react with endogenous mGM-CSF, cells in H-2Ld-hGM-CSFRα/βc double transgenic mice never receive hGM-CSF signals in vivo in the absence of hGM-CSF (8). Thus, FACS® analysis showed that the frequency of each myeloid and lymphoid progenitor population in hGM-CSFR transgenic mice was identical to that of wild-type mice (unpublished data).

Bottom Line: We evaluated the effects of ectopic granulocyte/macrophage colony-stimulating factor (GM-CSF) signals on hematopoietic commitment and differentiation.Strikingly, >50% hGM-CSFR+ common lymphoid progenitors (CLPs) and >20% hGM-CSFR+ pro-T cells gave rise to granulocyte, monocyte, and/or myeloid dendritic cells, but not MegE lineage cells in the presence of hGM-CSF.Thus, hGM-CSF transduces permissive signals for myeloerythroid differentiation, whereas it transmits potent instructive signals for the GM differentiation to CLPs and early T cell progenitors.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA.

ABSTRACT
We evaluated the effects of ectopic granulocyte/macrophage colony-stimulating factor (GM-CSF) signals on hematopoietic commitment and differentiation. Lineage-restricted progenitors purified from mice with the ubiquitous transgenic human GM-CSF receptor (hGM-CSFR) were used for the analysis. In cultures with hGM-CSF alone, hGM-CSFR-expressing (hGM-CSFR+) granulocyte/monocyte progenitors (GMPs) and megakaryocyte/erythrocyte progenitors (MEPs) exclusively gave rise to granulocyte/monocyte (GM) and megakaryocyte/erythroid (MegE) colonies, respectively, providing formal proof that GM-CSF signals support the GM and MegE lineage differentiation without affecting the physiological myeloid fate. hGM-CSFR transgenic mice were crossed with mice deficient in interleukin (IL)-7, an essential cytokine for T and B cell development. Administration of hGM-CSF in these mice could not restore T or B lymphopoiesis, indicating that enforced GM-CSF signals cannot substitute for IL-7 to promote lymphopoiesis. Strikingly, >50% hGM-CSFR+ common lymphoid progenitors (CLPs) and >20% hGM-CSFR+ pro-T cells gave rise to granulocyte, monocyte, and/or myeloid dendritic cells, but not MegE lineage cells in the presence of hGM-CSF. Injection of hGM-CSF into mice transplanted with hGM-CSFR+ CLPs blocked their lymphoid differentiation, but induced development of GM cells in vivo. Thus, hGM-CSF transduces permissive signals for myeloerythroid differentiation, whereas it transmits potent instructive signals for the GM differentiation to CLPs and early T cell progenitors. These data suggest that a majority of CLPs and a fraction of pro-T cells possess plasticity for myelomonocytic differentiation that can be activated by ectopic GM-CSF signals, supporting the hypothesis that the down-regulation of GM-CSFR is a critical event in producing cells with a lymphoid-restricted lineage potential.

Show MeSH
Related in: MedlinePlus