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The early progenitors of mouse dendritic cells and plasmacytoid predendritic cells are within the bone marrow hemopoietic precursors expressing Flt3.

D'Amico A, Wu L - J. Exp. Med. (2003)

Bottom Line: Most DC precursor activity, other than that given by multipotent hemopoietic stem cells, was within the downstream precursors expressing Flt3.The granulocyte and macrophage precursors and pro-B cells did not express Flt3 and had no DC or p-preDC precursor activity.These findings demonstrate that the early precursors for all DC subtypes are within the BM Flt3+ precursor populations, regardless of their lymphoid or myeloid lineage orientation.

View Article: PubMed Central - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, 1G, Royal Parade, Parkville, Victoria 3050, Australia.

ABSTRACT
Flt3 ligand (Flt3L) is a growth factor for hemopoietic progenitors and can promote the expansion of both conventional dendritic cells (DCs) and plasmacytoid predendritic cells (p-preDCs). The cells responding to Flt3L treatment and the precursors for the DCs and p-preDCs had not been fully characterized. We examined different mouse bone marrow (BM) hemopoietic precursor populations for the surface expression of Flt3 and tested them for early DC and p-preDC precursor activity. Most DC precursor activity, other than that given by multipotent hemopoietic stem cells, was within the downstream precursors expressing Flt3. The majority of mouse BM common lymphoid precursors expressed high levels of Flt3 and these were the most efficient precursors of both DCs and p-preDCs. In contrast, only a small proportion of the common myeloid precursors (CMPs) expressed Flt3, but the precursor activity for both DCs and p-preDCs was within this minor Flt3+ CMP fraction. The granulocyte and macrophage precursors and pro-B cells did not express Flt3 and had no DC or p-preDC precursor activity. These findings demonstrate that the early precursors for all DC subtypes are within the BM Flt3+ precursor populations, regardless of their lymphoid or myeloid lineage orientation.

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Surface expression of Flt3 by mouse BM cells and by different hemopoietic precursor populations. Total BM cells were stained with PE-conjugated anti-Flt3 antibody together with other fluorescence-conjugated antibodies against different surface molecules expressed by BM cells including CD19, B220, CD3, Mac-1, Gr-1, MHC class II, and c-kit. The expressions of these molecules on the gated Flt3+ BM cells are shown in A. The solid lines represent the staining of different surface molecules and the dashed lines represent the isotype controls. The precursor populations from mouse BM or thymus were first enriched for lineage marker–negative cells and then distinguished by four color fluorescence staining as (B) CLP (c-kit+ CD127+), (C) CMP (CD34+ CD16/32lo) and GMP (CD34+ CD16/32hi), (D) intrathymic CD4lo lymphoid precursors (c-kit+ Thy-1lo), and (E) BM pro-B precursors (c-kit+ CD19+). The surface expression of Flt3 by each precursor population was then examined. The contour plots display the gating for each precursor population (cells within the boxes). The histograms show the Flt3 expression on each gated precursor population. The solid lines represent the Flt3 staining and the dashed lines represent the isotype controls. Data presented in this figure is representative of at least three similar experiments on each precursor population.
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fig1: Surface expression of Flt3 by mouse BM cells and by different hemopoietic precursor populations. Total BM cells were stained with PE-conjugated anti-Flt3 antibody together with other fluorescence-conjugated antibodies against different surface molecules expressed by BM cells including CD19, B220, CD3, Mac-1, Gr-1, MHC class II, and c-kit. The expressions of these molecules on the gated Flt3+ BM cells are shown in A. The solid lines represent the staining of different surface molecules and the dashed lines represent the isotype controls. The precursor populations from mouse BM or thymus were first enriched for lineage marker–negative cells and then distinguished by four color fluorescence staining as (B) CLP (c-kit+ CD127+), (C) CMP (CD34+ CD16/32lo) and GMP (CD34+ CD16/32hi), (D) intrathymic CD4lo lymphoid precursors (c-kit+ Thy-1lo), and (E) BM pro-B precursors (c-kit+ CD19+). The surface expression of Flt3 by each precursor population was then examined. The contour plots display the gating for each precursor population (cells within the boxes). The histograms show the Flt3 expression on each gated precursor population. The solid lines represent the Flt3 staining and the dashed lines represent the isotype controls. Data presented in this figure is representative of at least three similar experiments on each precursor population.

Mentions: Based on the fact that both in vivo and in vitro treatment with Flt3L drastically increase the number of DCs and p-preDCs in mouse and human (10–15), we proposed that the precursors with the potential to generate DCs and p-preDCs express the receptor for Flt3L. Therefore, we examined the surface Flt3 expression on BM cells and on different defined hemopoietic precursor populations using multicolor flow cytometric analysis. We used PE-conjugated anti-Flt3 antibody together with a range of fluorochrome-conjugated antibodies to other cell surface markers for distinguishing different hemopoietic cell lineages and different precursor populations. The different hemopoietic lineage cells were distinguished as B cells (B220+ CD19+), myeloid cells, (Mac-1+ Gr-1+), and T cells (CD3+). For distinguishing the BM precursor populations, we used the procedures established in the Stanford laboratories (29, 30) and in this laboratory (28), which defined the precursor populations as: multipotent HSCs, Lin− Sca-1+ c-kithi; the CLPs, Lin− IL-7Rα+ Sca-1lo c-kitlo; the CMPs, Lin− IL-7Rα− Sca-1− c-kithi CD34+ CD16/32lo; and GMPs, Lin− IL-7Rα− Sca-1− c-kithi CD34+CD16/32hi. The surface expression of Flt3 on the more committed precursors for T and B lymphoid cells was also examined. These precursors were distinguished as: intrathymic lymphoid precursors (CD4lo precursors), Lin− CD3− 8− 25− 4lo Thy-1lo c-kit+; and BM pro-B cells, Lin− CD19+ B220+ c-kit+. The results are shown in Fig. 1 .


The early progenitors of mouse dendritic cells and plasmacytoid predendritic cells are within the bone marrow hemopoietic precursors expressing Flt3.

D'Amico A, Wu L - J. Exp. Med. (2003)

Surface expression of Flt3 by mouse BM cells and by different hemopoietic precursor populations. Total BM cells were stained with PE-conjugated anti-Flt3 antibody together with other fluorescence-conjugated antibodies against different surface molecules expressed by BM cells including CD19, B220, CD3, Mac-1, Gr-1, MHC class II, and c-kit. The expressions of these molecules on the gated Flt3+ BM cells are shown in A. The solid lines represent the staining of different surface molecules and the dashed lines represent the isotype controls. The precursor populations from mouse BM or thymus were first enriched for lineage marker–negative cells and then distinguished by four color fluorescence staining as (B) CLP (c-kit+ CD127+), (C) CMP (CD34+ CD16/32lo) and GMP (CD34+ CD16/32hi), (D) intrathymic CD4lo lymphoid precursors (c-kit+ Thy-1lo), and (E) BM pro-B precursors (c-kit+ CD19+). The surface expression of Flt3 by each precursor population was then examined. The contour plots display the gating for each precursor population (cells within the boxes). The histograms show the Flt3 expression on each gated precursor population. The solid lines represent the Flt3 staining and the dashed lines represent the isotype controls. Data presented in this figure is representative of at least three similar experiments on each precursor population.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2194069&req=5

fig1: Surface expression of Flt3 by mouse BM cells and by different hemopoietic precursor populations. Total BM cells were stained with PE-conjugated anti-Flt3 antibody together with other fluorescence-conjugated antibodies against different surface molecules expressed by BM cells including CD19, B220, CD3, Mac-1, Gr-1, MHC class II, and c-kit. The expressions of these molecules on the gated Flt3+ BM cells are shown in A. The solid lines represent the staining of different surface molecules and the dashed lines represent the isotype controls. The precursor populations from mouse BM or thymus were first enriched for lineage marker–negative cells and then distinguished by four color fluorescence staining as (B) CLP (c-kit+ CD127+), (C) CMP (CD34+ CD16/32lo) and GMP (CD34+ CD16/32hi), (D) intrathymic CD4lo lymphoid precursors (c-kit+ Thy-1lo), and (E) BM pro-B precursors (c-kit+ CD19+). The surface expression of Flt3 by each precursor population was then examined. The contour plots display the gating for each precursor population (cells within the boxes). The histograms show the Flt3 expression on each gated precursor population. The solid lines represent the Flt3 staining and the dashed lines represent the isotype controls. Data presented in this figure is representative of at least three similar experiments on each precursor population.
Mentions: Based on the fact that both in vivo and in vitro treatment with Flt3L drastically increase the number of DCs and p-preDCs in mouse and human (10–15), we proposed that the precursors with the potential to generate DCs and p-preDCs express the receptor for Flt3L. Therefore, we examined the surface Flt3 expression on BM cells and on different defined hemopoietic precursor populations using multicolor flow cytometric analysis. We used PE-conjugated anti-Flt3 antibody together with a range of fluorochrome-conjugated antibodies to other cell surface markers for distinguishing different hemopoietic cell lineages and different precursor populations. The different hemopoietic lineage cells were distinguished as B cells (B220+ CD19+), myeloid cells, (Mac-1+ Gr-1+), and T cells (CD3+). For distinguishing the BM precursor populations, we used the procedures established in the Stanford laboratories (29, 30) and in this laboratory (28), which defined the precursor populations as: multipotent HSCs, Lin− Sca-1+ c-kithi; the CLPs, Lin− IL-7Rα+ Sca-1lo c-kitlo; the CMPs, Lin− IL-7Rα− Sca-1− c-kithi CD34+ CD16/32lo; and GMPs, Lin− IL-7Rα− Sca-1− c-kithi CD34+CD16/32hi. The surface expression of Flt3 on the more committed precursors for T and B lymphoid cells was also examined. These precursors were distinguished as: intrathymic lymphoid precursors (CD4lo precursors), Lin− CD3− 8− 25− 4lo Thy-1lo c-kit+; and BM pro-B cells, Lin− CD19+ B220+ c-kit+. The results are shown in Fig. 1 .

Bottom Line: Most DC precursor activity, other than that given by multipotent hemopoietic stem cells, was within the downstream precursors expressing Flt3.The granulocyte and macrophage precursors and pro-B cells did not express Flt3 and had no DC or p-preDC precursor activity.These findings demonstrate that the early precursors for all DC subtypes are within the BM Flt3+ precursor populations, regardless of their lymphoid or myeloid lineage orientation.

View Article: PubMed Central - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, 1G, Royal Parade, Parkville, Victoria 3050, Australia.

ABSTRACT
Flt3 ligand (Flt3L) is a growth factor for hemopoietic progenitors and can promote the expansion of both conventional dendritic cells (DCs) and plasmacytoid predendritic cells (p-preDCs). The cells responding to Flt3L treatment and the precursors for the DCs and p-preDCs had not been fully characterized. We examined different mouse bone marrow (BM) hemopoietic precursor populations for the surface expression of Flt3 and tested them for early DC and p-preDC precursor activity. Most DC precursor activity, other than that given by multipotent hemopoietic stem cells, was within the downstream precursors expressing Flt3. The majority of mouse BM common lymphoid precursors expressed high levels of Flt3 and these were the most efficient precursors of both DCs and p-preDCs. In contrast, only a small proportion of the common myeloid precursors (CMPs) expressed Flt3, but the precursor activity for both DCs and p-preDCs was within this minor Flt3+ CMP fraction. The granulocyte and macrophage precursors and pro-B cells did not express Flt3 and had no DC or p-preDC precursor activity. These findings demonstrate that the early precursors for all DC subtypes are within the BM Flt3+ precursor populations, regardless of their lymphoid or myeloid lineage orientation.

Show MeSH
Related in: MedlinePlus