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CD4(+)CD25(-)Foxp3(-) Th1 cells are the source of IL-10-mediated immune suppression in chronic cutaneous leishmaniasis.

Anderson CF, Oukka M, Kuchroo VJ, Sacks D - J. Exp. Med. (2007)

Bottom Line: Nonhealing forms of leishmaniasis in humans are commonly associated with elevated levels of the deactivating cytokine IL-10, and in the mouse, normally chronic infections can be cleared in the absence of IL-10.IL-10 was produced by innate cells, as well as CD4(+)CD25(+)Foxp3(+) and CD4(+)CD25(-)Foxp3(-) T cells in the chronic lesion.Surprisingly, Rag(-/-) mice reconstituted with naive CD4(+) T cells depleted of natural T regulatory cells developed more severe infections, associated with elevated levels of IL-10 and, especially, Th2 cytokines in the site.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
Nonhealing forms of leishmaniasis in humans are commonly associated with elevated levels of the deactivating cytokine IL-10, and in the mouse, normally chronic infections can be cleared in the absence of IL-10. Using a Leishmania major strain that produces nonhealing dermal lesions in a T helper type 1 (Th1) cell-polarized setting, we have analyzed the cellular sources of IL-10 and their relative contribution to immune suppression. IL-10 was produced by innate cells, as well as CD4(+)CD25(+)Foxp3(+) and CD4(+)CD25(-)Foxp3(-) T cells in the chronic lesion. Nonetheless, only IL-10 production by antigen-specific CD4(+)CD25(-)Foxp3(-) T cells, the majority of which also produced IFN-gamma, was necessary for suppression of acquired immunity in Rag(-/-) reconstituted mice. Surprisingly, Rag(-/-) mice reconstituted with naive CD4(+) T cells depleted of natural T regulatory cells developed more severe infections, associated with elevated levels of IL-10 and, especially, Th2 cytokines in the site. The data demonstrate that IL-10-producing Th1 cells, activated early in a strong inflammatory setting as a mechanism of feedback control, are the principal mediators of T cell-derived IL-10-dependent immune suppression in a chronic intracellular infection.

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Natural T reg cells suppress Th2 cell development. 2 × 106 CD4+GFP− cells from naive Foxp3-GFP knock-in mice were adoptively transferred alone (shaded bars) or in combination with 2 × 105 CD4+GFP+ cells (open bars), followed by infection 1 d later. 6 wk after infection, mice were killed, and real-time PCR was performed on RNA isolated from ear lesions. Values shown are the mean ± SEM and are relative gene expression normalized to 18S rRNA. Data were generated from cells pooled from six mice in each group and are representative of three experiments. *, P < 0.05; **, P < 0.01.
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fig6: Natural T reg cells suppress Th2 cell development. 2 × 106 CD4+GFP− cells from naive Foxp3-GFP knock-in mice were adoptively transferred alone (shaded bars) or in combination with 2 × 105 CD4+GFP+ cells (open bars), followed by infection 1 d later. 6 wk after infection, mice were killed, and real-time PCR was performed on RNA isolated from ear lesions. Values shown are the mean ± SEM and are relative gene expression normalized to 18S rRNA. Data were generated from cells pooled from six mice in each group and are representative of three experiments. *, P < 0.05; **, P < 0.01.

Mentions: Although IL-10 derived from natural T reg cells does not appear to contribute greatly to the suppression of immunity in C57BL/6 mice infected with L. major NIH/Sd, these cells might still play a critical role in suppression by production of other regulatory cytokines (e.g., TGF-β) or by cell contact–dependent mechanisms. To determine what effect, if any, the presence of natural T reg cells had on the development of the immune response in this infection, CD4+ T cells from the lymph nodes of naive C57BL/10 mice were depleted or not of CD25+ cells by sort purification and adoptively transferred into RAG−/− recipients. 10 wk after transfer and infection, the cell composition and the parasite burdens in the lesions were measured. Although Foxp3+ cells still accumulated in the ear lesions of mice reconstituted with CD25+-depleted cells (3.6% of CD4+ T cells), their frequency was significantly lower than in the lesions of mice receiving the total CD4+ cells (11.7% of CD4+ T cells; Fig. 5 A). Nonetheless, the mice receiving the CD25+-depleted cells had higher parasite burdens, suggesting that, if anything, the natural T reg cells appeared to play a role in promoting host resistance (Fig. 5 A). Because a percentage of the sort purified CD4+CD25− cells are Foxp3+ (Fig. 2 C) and because homeostatic proliferation of the transferred CD4+CD25− cells may result in an outgrowth of the Foxp3+ lineage (32), we used bicistronic mice with an enhanced GFP reporter inserted into the Foxp3 locus as a source of CD4+ T cells from which natural T reg cells might be more faithfully removed (33). 6 wk after adoptive transfer and infection, mice that had received CD4+GFP− cells had few Foxp3+ cells in the draining lymph nodes (2.3%) and at the site of infection (1.5%) compared with the group that received unselected CD4+ cells (8 and 23.8%, respectively; Fig. 5 B). In agreement with the experiment involving transfer of CD25+-depleted cells, the absence of Foxp3−GFP+ cells resulted in a significant increase in parasite burdens in the ear. The termination of the experiment at 6 wk after infection as opposed to a more chronic stage of infection was conditioned by the mice becoming moribund, demonstrating weight loss and signs of colitis that were confirmed on necropsy (unpublished data). The development of colitis in T cell–reconstituted RAG −/− mice is taken as additional evidence for the functional depletion of natural T reg cells in the these recipients (34, 35). The frequency of IL-10+Foxp3− cells, the majority of which also produced IFN-γ, was slightly greater than in the mice transferred with total CD4+ cells (unpublished data). These results were confirmed by the real-time PCR analysis of lesional cytokine mRNA, which revealed slight increases in IL-10, IFN-γ, and TNF mRNA levels in mice reconstituted with the Foxp3-depleted cells (Fig. 6). Interestingly, the lesional cells in these mice also showed an increase in IL-4, IL-5, and IL-13 mRNA expression, indicating that natural T reg cells functioned to suppress the development of a Th2 response. Thus, naive CD4+Foxp3− T cells alone were capable of being activated by the L. major NIH/Sd strain to produce IFN-γ and/or IL-10 and of being recruited to the site of infection in the skin, where they established the conditions necessary for the development of contained, but chronic, nonhealing lesions. The influence of natural T reg cells in this process appeared to be a global suppression of the effector response, including IL-10–producing Th1 cells and, especially, Th2 cells, so that on balance their activities may actually have promoted host immunity in the inflammatory site.


CD4(+)CD25(-)Foxp3(-) Th1 cells are the source of IL-10-mediated immune suppression in chronic cutaneous leishmaniasis.

Anderson CF, Oukka M, Kuchroo VJ, Sacks D - J. Exp. Med. (2007)

Natural T reg cells suppress Th2 cell development. 2 × 106 CD4+GFP− cells from naive Foxp3-GFP knock-in mice were adoptively transferred alone (shaded bars) or in combination with 2 × 105 CD4+GFP+ cells (open bars), followed by infection 1 d later. 6 wk after infection, mice were killed, and real-time PCR was performed on RNA isolated from ear lesions. Values shown are the mean ± SEM and are relative gene expression normalized to 18S rRNA. Data were generated from cells pooled from six mice in each group and are representative of three experiments. *, P < 0.05; **, P < 0.01.
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Related In: Results  -  Collection

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fig6: Natural T reg cells suppress Th2 cell development. 2 × 106 CD4+GFP− cells from naive Foxp3-GFP knock-in mice were adoptively transferred alone (shaded bars) or in combination with 2 × 105 CD4+GFP+ cells (open bars), followed by infection 1 d later. 6 wk after infection, mice were killed, and real-time PCR was performed on RNA isolated from ear lesions. Values shown are the mean ± SEM and are relative gene expression normalized to 18S rRNA. Data were generated from cells pooled from six mice in each group and are representative of three experiments. *, P < 0.05; **, P < 0.01.
Mentions: Although IL-10 derived from natural T reg cells does not appear to contribute greatly to the suppression of immunity in C57BL/6 mice infected with L. major NIH/Sd, these cells might still play a critical role in suppression by production of other regulatory cytokines (e.g., TGF-β) or by cell contact–dependent mechanisms. To determine what effect, if any, the presence of natural T reg cells had on the development of the immune response in this infection, CD4+ T cells from the lymph nodes of naive C57BL/10 mice were depleted or not of CD25+ cells by sort purification and adoptively transferred into RAG−/− recipients. 10 wk after transfer and infection, the cell composition and the parasite burdens in the lesions were measured. Although Foxp3+ cells still accumulated in the ear lesions of mice reconstituted with CD25+-depleted cells (3.6% of CD4+ T cells), their frequency was significantly lower than in the lesions of mice receiving the total CD4+ cells (11.7% of CD4+ T cells; Fig. 5 A). Nonetheless, the mice receiving the CD25+-depleted cells had higher parasite burdens, suggesting that, if anything, the natural T reg cells appeared to play a role in promoting host resistance (Fig. 5 A). Because a percentage of the sort purified CD4+CD25− cells are Foxp3+ (Fig. 2 C) and because homeostatic proliferation of the transferred CD4+CD25− cells may result in an outgrowth of the Foxp3+ lineage (32), we used bicistronic mice with an enhanced GFP reporter inserted into the Foxp3 locus as a source of CD4+ T cells from which natural T reg cells might be more faithfully removed (33). 6 wk after adoptive transfer and infection, mice that had received CD4+GFP− cells had few Foxp3+ cells in the draining lymph nodes (2.3%) and at the site of infection (1.5%) compared with the group that received unselected CD4+ cells (8 and 23.8%, respectively; Fig. 5 B). In agreement with the experiment involving transfer of CD25+-depleted cells, the absence of Foxp3−GFP+ cells resulted in a significant increase in parasite burdens in the ear. The termination of the experiment at 6 wk after infection as opposed to a more chronic stage of infection was conditioned by the mice becoming moribund, demonstrating weight loss and signs of colitis that were confirmed on necropsy (unpublished data). The development of colitis in T cell–reconstituted RAG −/− mice is taken as additional evidence for the functional depletion of natural T reg cells in the these recipients (34, 35). The frequency of IL-10+Foxp3− cells, the majority of which also produced IFN-γ, was slightly greater than in the mice transferred with total CD4+ cells (unpublished data). These results were confirmed by the real-time PCR analysis of lesional cytokine mRNA, which revealed slight increases in IL-10, IFN-γ, and TNF mRNA levels in mice reconstituted with the Foxp3-depleted cells (Fig. 6). Interestingly, the lesional cells in these mice also showed an increase in IL-4, IL-5, and IL-13 mRNA expression, indicating that natural T reg cells functioned to suppress the development of a Th2 response. Thus, naive CD4+Foxp3− T cells alone were capable of being activated by the L. major NIH/Sd strain to produce IFN-γ and/or IL-10 and of being recruited to the site of infection in the skin, where they established the conditions necessary for the development of contained, but chronic, nonhealing lesions. The influence of natural T reg cells in this process appeared to be a global suppression of the effector response, including IL-10–producing Th1 cells and, especially, Th2 cells, so that on balance their activities may actually have promoted host immunity in the inflammatory site.

Bottom Line: Nonhealing forms of leishmaniasis in humans are commonly associated with elevated levels of the deactivating cytokine IL-10, and in the mouse, normally chronic infections can be cleared in the absence of IL-10.IL-10 was produced by innate cells, as well as CD4(+)CD25(+)Foxp3(+) and CD4(+)CD25(-)Foxp3(-) T cells in the chronic lesion.Surprisingly, Rag(-/-) mice reconstituted with naive CD4(+) T cells depleted of natural T regulatory cells developed more severe infections, associated with elevated levels of IL-10 and, especially, Th2 cytokines in the site.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
Nonhealing forms of leishmaniasis in humans are commonly associated with elevated levels of the deactivating cytokine IL-10, and in the mouse, normally chronic infections can be cleared in the absence of IL-10. Using a Leishmania major strain that produces nonhealing dermal lesions in a T helper type 1 (Th1) cell-polarized setting, we have analyzed the cellular sources of IL-10 and their relative contribution to immune suppression. IL-10 was produced by innate cells, as well as CD4(+)CD25(+)Foxp3(+) and CD4(+)CD25(-)Foxp3(-) T cells in the chronic lesion. Nonetheless, only IL-10 production by antigen-specific CD4(+)CD25(-)Foxp3(-) T cells, the majority of which also produced IFN-gamma, was necessary for suppression of acquired immunity in Rag(-/-) reconstituted mice. Surprisingly, Rag(-/-) mice reconstituted with naive CD4(+) T cells depleted of natural T regulatory cells developed more severe infections, associated with elevated levels of IL-10 and, especially, Th2 cytokines in the site. The data demonstrate that IL-10-producing Th1 cells, activated early in a strong inflammatory setting as a mechanism of feedback control, are the principal mediators of T cell-derived IL-10-dependent immune suppression in a chronic intracellular infection.

Show MeSH
Related in: MedlinePlus