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Systemic Expression of Notch Ligand Delta-Like 4 during Mycobacterial Infection Alters the T Cell Immune Response

View Article: PubMed Central - PubMed

ABSTRACT

The Notch ligand delta-like 4 (DLL4) is known to fine-tune the CD4+ T cell cytokine response. DLL4 is expressed on the surface of antigen-presenting cells (APCs) in a MyD88-dependent manner. We found that DLL4 expression was upregulated on bone marrow progenitor cells and APCs in mice infected with BCG Mycobacterium. Transfer of DLL4+ progenitor cells from infected hosts resulted in an increase DLL4+ myeloid cells in the spleen, indicating that expression of the dll4 gene is propagated throughout hematopoiesis. We also found an increase in DLL4+ monocytes from individuals who were infected with Mycobacterium tuberculosis. In latent individuals, DLL4 expression correlated with increased cytokine production from T cells in response to PPD stimulation. Finally, antibody blockade of DLL4 reduced T cell cytokine production from naïve T cells stimulated with antigen. These results demonstrate that the Notch ligand DLL4 can influence T cell cytokine production in both humans and mice, and further reveal that expression of DLL4 is upregulated on early hematopoietic progenitors in response to chronic mycobacterial infection. These data suggest that widespread DLL4 expression may occur as a result of mycobacterial infection, and that this expression may alter CD4+ T cell responses to both previously encountered and novel antigens.

No MeSH data available.


Related in: MedlinePlus

Delta-like 4 expression is maintained during short-term hematopoiesis. (A) Flow cytometry plots depicting dll4 and MHCII staining in total splenocytes from recipient chimera mice 4 weeks post-engraftment. Donor cells were either dll4+ LSK or dll4− LSK cells isolated from mice infected with 5.0 × 105 cells at 5 weeks post-infection. Each mouse received 5.0 × 104 LSK cells. (B) QPCR analysis of whole splenocytes for dll4 expression at 4 weeks post-engraftment. N = 4 mice per group, and the experiment was repeated two times. (C) Quantification of dll4 expression on B220−CD11B+MHCII+ and B220−CD11B+MHCII− in the spleens of chimeric mice. N = 4 mice per group. (D) Flow plots depicting expression of dll4 on B220−CD11B+MHCII− cells as quantified in (C). Gates indicate percent of B220−CD11B+MHCII− cells expressing DLL4. For (B,C), Student’s t-test was performed (*p < 0.04). These experiments were repeated two times.
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Figure 2: Delta-like 4 expression is maintained during short-term hematopoiesis. (A) Flow cytometry plots depicting dll4 and MHCII staining in total splenocytes from recipient chimera mice 4 weeks post-engraftment. Donor cells were either dll4+ LSK or dll4− LSK cells isolated from mice infected with 5.0 × 105 cells at 5 weeks post-infection. Each mouse received 5.0 × 104 LSK cells. (B) QPCR analysis of whole splenocytes for dll4 expression at 4 weeks post-engraftment. N = 4 mice per group, and the experiment was repeated two times. (C) Quantification of dll4 expression on B220−CD11B+MHCII+ and B220−CD11B+MHCII− in the spleens of chimeric mice. N = 4 mice per group. (D) Flow plots depicting expression of dll4 on B220−CD11B+MHCII− cells as quantified in (C). Gates indicate percent of B220−CD11B+MHCII− cells expressing DLL4. For (B,C), Student’s t-test was performed (*p < 0.04). These experiments were repeated two times.

Mentions: To determine if the expression of DLL4 was propagated throughout the hematopoietic system as a result of hematopoiesis, we isolated DLL4+ and DLL4− Lineage− Sca1+ cKit+ (LSK) cells from the bone marrow of BCG-infected mice by FACS. Isolated cells were engrafted into lethally irradiated hosts and splenocytes were assessed for DLL4 expression at several time points post engraftment. We observed a significant increase in DLL4 expression in the spleen by flow cytometry on CD11b+ MHCII+ cells isolated from recipient mice at 4 weeks post-engraftment (Figures 2A,B). QPCR from whole spleen demonstrated a 1.8-fold increase in expression of dll4 at the RNA level (Figure 2B). We also observed an increase in DLL4 expression in CD11B+MHCII− cells in the spleen; however, the majority of DLL4 expression was found in the MHCII+ population (Figures 2C,D). We did not observe an increase in expression of DLL4 on lymphocytes in these mice (Figure S3 in Supplementary Material). We did not observe an increase in DLL4 expression in mice receiving DLL4+ LSK cells at 7 days post-engraftment or at 8 weeks post-engraftment, suggesting that the expression of DLL4 as a result of hematopoiesis is transient in the absence of BCG infection, and is dependent on donor cell expression of this ligand. Using congenic mice, we verified that the majority of DLL4 expression was detected on donor cells and not on radio-resistant recipient cells at 4 weeks post-engraftment (Figure S3 in Supplementary Material).


Systemic Expression of Notch Ligand Delta-Like 4 during Mycobacterial Infection Alters the T Cell Immune Response
Delta-like 4 expression is maintained during short-term hematopoiesis. (A) Flow cytometry plots depicting dll4 and MHCII staining in total splenocytes from recipient chimera mice 4 weeks post-engraftment. Donor cells were either dll4+ LSK or dll4− LSK cells isolated from mice infected with 5.0 × 105 cells at 5 weeks post-infection. Each mouse received 5.0 × 104 LSK cells. (B) QPCR analysis of whole splenocytes for dll4 expression at 4 weeks post-engraftment. N = 4 mice per group, and the experiment was repeated two times. (C) Quantification of dll4 expression on B220−CD11B+MHCII+ and B220−CD11B+MHCII− in the spleens of chimeric mice. N = 4 mice per group. (D) Flow plots depicting expression of dll4 on B220−CD11B+MHCII− cells as quantified in (C). Gates indicate percent of B220−CD11B+MHCII− cells expressing DLL4. For (B,C), Student’s t-test was performed (*p < 0.04). These experiments were repeated two times.
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Related In: Results  -  Collection

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Figure 2: Delta-like 4 expression is maintained during short-term hematopoiesis. (A) Flow cytometry plots depicting dll4 and MHCII staining in total splenocytes from recipient chimera mice 4 weeks post-engraftment. Donor cells were either dll4+ LSK or dll4− LSK cells isolated from mice infected with 5.0 × 105 cells at 5 weeks post-infection. Each mouse received 5.0 × 104 LSK cells. (B) QPCR analysis of whole splenocytes for dll4 expression at 4 weeks post-engraftment. N = 4 mice per group, and the experiment was repeated two times. (C) Quantification of dll4 expression on B220−CD11B+MHCII+ and B220−CD11B+MHCII− in the spleens of chimeric mice. N = 4 mice per group. (D) Flow plots depicting expression of dll4 on B220−CD11B+MHCII− cells as quantified in (C). Gates indicate percent of B220−CD11B+MHCII− cells expressing DLL4. For (B,C), Student’s t-test was performed (*p < 0.04). These experiments were repeated two times.
Mentions: To determine if the expression of DLL4 was propagated throughout the hematopoietic system as a result of hematopoiesis, we isolated DLL4+ and DLL4− Lineage− Sca1+ cKit+ (LSK) cells from the bone marrow of BCG-infected mice by FACS. Isolated cells were engrafted into lethally irradiated hosts and splenocytes were assessed for DLL4 expression at several time points post engraftment. We observed a significant increase in DLL4 expression in the spleen by flow cytometry on CD11b+ MHCII+ cells isolated from recipient mice at 4 weeks post-engraftment (Figures 2A,B). QPCR from whole spleen demonstrated a 1.8-fold increase in expression of dll4 at the RNA level (Figure 2B). We also observed an increase in DLL4 expression in CD11B+MHCII− cells in the spleen; however, the majority of DLL4 expression was found in the MHCII+ population (Figures 2C,D). We did not observe an increase in expression of DLL4 on lymphocytes in these mice (Figure S3 in Supplementary Material). We did not observe an increase in DLL4 expression in mice receiving DLL4+ LSK cells at 7 days post-engraftment or at 8 weeks post-engraftment, suggesting that the expression of DLL4 as a result of hematopoiesis is transient in the absence of BCG infection, and is dependent on donor cell expression of this ligand. Using congenic mice, we verified that the majority of DLL4 expression was detected on donor cells and not on radio-resistant recipient cells at 4 weeks post-engraftment (Figure S3 in Supplementary Material).

View Article: PubMed Central - PubMed

ABSTRACT

The Notch ligand delta-like 4 (DLL4) is known to fine-tune the CD4+ T cell cytokine response. DLL4 is expressed on the surface of antigen-presenting cells (APCs) in a MyD88-dependent manner. We found that DLL4 expression was upregulated on bone marrow progenitor cells and APCs in mice infected with BCG Mycobacterium. Transfer of DLL4+ progenitor cells from infected hosts resulted in an increase DLL4+ myeloid cells in the spleen, indicating that expression of the dll4 gene is propagated throughout hematopoiesis. We also found an increase in DLL4+ monocytes from individuals who were infected with Mycobacterium tuberculosis. In latent individuals, DLL4 expression correlated with increased cytokine production from T cells in response to PPD stimulation. Finally, antibody blockade of DLL4 reduced T cell cytokine production from na&iuml;ve T cells stimulated with antigen. These results demonstrate that the Notch ligand DLL4 can influence T cell cytokine production in both humans and mice, and further reveal that expression of DLL4 is upregulated on early hematopoietic progenitors in response to chronic mycobacterial infection. These data suggest that widespread DLL4 expression may occur as a result of mycobacterial infection, and that this expression may alter CD4+ T cell responses to both previously encountered and novel antigens.

No MeSH data available.


Related in: MedlinePlus