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Negative feedback control of the autoimmune response through antigen-induced differentiation of IL-10-secreting Th1 cells.

Gabrysová L, Nicolson KS, Streeter HB, Verhagen J, Sabatos-Peyton CA, Morgan DJ, Wraith DC - J. Exp. Med. (2009)

Bottom Line: Loss of proliferative capacity correlated with a switch from the Th1-associated cytokines IL-2 and interferon (IFN)-gamma to the regulatory cytokine IL-10.IL-10 T reg cells suppressed dendritic cell maturation, prevented Th1 cell differentiation, and thereby created a negative feedback loop for Th1-driven immune pathology.These findings demonstrate that Th1 responses can be self-limiting in the context of peripheral tolerance to a self-antigen.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Medicine, University of Bristol School of Medical Sciences, Bristol BS8 1TD, England, UK. Leona.Gabrysova@bristol.ac.uk

ABSTRACT
Regulation of the immune response to self- and foreign antigens is vitally important for limiting immune pathology associated with both infections and hypersensitivity conditions. Control of autoimmune conditions can be reinforced by tolerance induction with peptide epitopes, but the mechanism is not currently understood. Repetitive intranasal administration of soluble peptide induces peripheral tolerance in myelin basic protein (MBP)-specific TCR transgenic mice. This is characterized by the presence of anergic, interleukin (IL)-10-secreting CD4(+) T cells with regulatory function (IL-10 T reg cells). The differentiation pathway of peptide-induced IL-10 T reg cells was investigated. CD4(+) T cells became anergic after their second encounter with a high-affinity MBP peptide analogue. Loss of proliferative capacity correlated with a switch from the Th1-associated cytokines IL-2 and interferon (IFN)-gamma to the regulatory cytokine IL-10. Nevertheless, IL-10 T reg cells retained the capacity to produce IFN-gamma and concomitantly expressed T-bet, demonstrating their Th1 origin. IL-10 T reg cells suppressed dendritic cell maturation, prevented Th1 cell differentiation, and thereby created a negative feedback loop for Th1-driven immune pathology. These findings demonstrate that Th1 responses can be self-limiting in the context of peripheral tolerance to a self-antigen.

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Related in: MedlinePlus

Anergy-associated down-regulation of Th1 cytokines and increase in IL-10 production by CD4+ T cells during tolerization. Splenic CD4+ T cells were positively selected from untreated or i.n. MBP Ac1-9[4Y]–treated Tg4 Rag1−/− mice at 2 h after each peptide administration. 5 × 104 CD4+ T cells per well were cultured with 100 µg/ml of MBP Ac1-9[4K] in the presence of 105 autologous irradiated CD4 depleted splenocytes as APCs and, where indicated, 20 U/ml of rhIL-2. (A and B) Proliferative responses were measured at 72 h by 3[H]thymidine incorporation in the absence or presence of rhIL-2. Results are depicted as the mean thymidine incorporation ± SEM. (C and D) Cytokine responses of CD4+ T cells from the cultures described in A and B were measured by ELISA at 24 h (IL-2) and 72 h (IFN-γ and IL-10) after in vitro restimulation in the absence or presence of rhIL-2. Results are depicted as the mean cytokine production in ng/ml of supernatant ± SEM. Data in each panel are representative of three individual experiments.
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fig2: Anergy-associated down-regulation of Th1 cytokines and increase in IL-10 production by CD4+ T cells during tolerization. Splenic CD4+ T cells were positively selected from untreated or i.n. MBP Ac1-9[4Y]–treated Tg4 Rag1−/− mice at 2 h after each peptide administration. 5 × 104 CD4+ T cells per well were cultured with 100 µg/ml of MBP Ac1-9[4K] in the presence of 105 autologous irradiated CD4 depleted splenocytes as APCs and, where indicated, 20 U/ml of rhIL-2. (A and B) Proliferative responses were measured at 72 h by 3[H]thymidine incorporation in the absence or presence of rhIL-2. Results are depicted as the mean thymidine incorporation ± SEM. (C and D) Cytokine responses of CD4+ T cells from the cultures described in A and B were measured by ELISA at 24 h (IL-2) and 72 h (IFN-γ and IL-10) after in vitro restimulation in the absence or presence of rhIL-2. Results are depicted as the mean cytokine production in ng/ml of supernatant ± SEM. Data in each panel are representative of three individual experiments.

Mentions: Although the serum cytokine profile provides an overall picture of the immune response to i.n. peptide administration, it does not allow discrimination of cytokine production by different cellular components of the immune system. We therefore investigated the contribution of CD4+ T cells to the aforementioned changes in serum cytokine concentrations. Tg4 CD4+ T cells were previously shown to respond to the first i.n. peptide treatment by initial proliferation and expansion accompanied by a cytokine burst (Burkhart et al., 1999). Indeed, splenocytes from Tg4 Rag1−/− mice, after successive i.n. MBP Ac1-9[4Y] treatments, showed a fourfold increase in their number at 2 h after the second treatment compared with untreated mice; the number of splenocytes decreased with subsequent i.n. peptide treatments (Fig. S1 A). The number of CD4+ T cells in the peptide-treated Tg4 Rag1−/− spleens increased with a similar pattern to that of whole splenocytes during the course of tolerance induction (Fig. S1 B). Given that CD4+ T cells from Tg4 mice treated repeatedly with i.n. peptide appear profoundly unresponsive (Sundstedt et al., 2003), we determined the kinetics of anergy induction as well as changes in the CD4+ T cell cytokine profile over the course of tolerization in Tg4 Rag1−/− mice. As shown in Fig. 2 A, CD4+ T cells from mice either left untreated or treated once with i.n. peptide proliferated vigorously in response to antigen restimulation in vitro. However, the level of CD4+ T cell proliferation declined dramatically upon subsequent i.n. peptide treatments (Fig. 2 A). The addition of recombinant human IL-2 (rhIL-2) to the unresponsive CD4+ T cells led to recovery of proliferation (Fig. 2 B), thereby demonstrating that the cells were in a state of clonal anergy. Supernatants from the CD4+ T cell proliferation assays were collected and analyzed for IL-2, IFN-γ, IL-4 and IL-10 levels by ELISA. Fig. 2 C shows changes in CD4+ T cell cytokine secretion after successive i.n. peptide treatments. CD4+ T cells from mice either left untreated or treated once with i.n. peptide secreted high levels of IL-2 and IFN-γ when restimulated with antigen in vitro. Levels of both cytokines decreased dramatically after just two i.n. peptide treatments (Fig. 2 C, top and middle, respectively). IL-10 was only detected in cultures of CD4+ T cells from mice treated three times or more, i.e., after the decrease in IL-2 and IFN-γ secretion (Fig. 2 C, bottom). A similar inverse relationship between IL-2 and IL-10 was also observed in the analysis of serum cytokines (Fig. 1). Although the addition of rhIL-2 restored proliferation (Fig. 2 A), it did not have an effect on endogenous IL-2 secretion and resulted in only a minor increase in IFN-γ production by CD4+ T cells isolated after the tenth i.n. peptide treatment (Fig. 2 D, top and middle, respectively). In contrast, the secretion of IL-10 was greatly augmented by the presence of rhIL-2 (Fig. 2 D, bottom). IL-4 remained below the detection level throughout the course of i.n. peptide treatment, irrespective of the addition of rhIL-2 (not depicted). These results support the findings of previous studies showing that IL-2 reverses the anergy of peptide-induced IL-10 T reg cells from mice treated with 10 doses of i.n. peptide without changing their cytokine secretion pattern (Anderson et al., 2005).


Negative feedback control of the autoimmune response through antigen-induced differentiation of IL-10-secreting Th1 cells.

Gabrysová L, Nicolson KS, Streeter HB, Verhagen J, Sabatos-Peyton CA, Morgan DJ, Wraith DC - J. Exp. Med. (2009)

Anergy-associated down-regulation of Th1 cytokines and increase in IL-10 production by CD4+ T cells during tolerization. Splenic CD4+ T cells were positively selected from untreated or i.n. MBP Ac1-9[4Y]–treated Tg4 Rag1−/− mice at 2 h after each peptide administration. 5 × 104 CD4+ T cells per well were cultured with 100 µg/ml of MBP Ac1-9[4K] in the presence of 105 autologous irradiated CD4 depleted splenocytes as APCs and, where indicated, 20 U/ml of rhIL-2. (A and B) Proliferative responses were measured at 72 h by 3[H]thymidine incorporation in the absence or presence of rhIL-2. Results are depicted as the mean thymidine incorporation ± SEM. (C and D) Cytokine responses of CD4+ T cells from the cultures described in A and B were measured by ELISA at 24 h (IL-2) and 72 h (IFN-γ and IL-10) after in vitro restimulation in the absence or presence of rhIL-2. Results are depicted as the mean cytokine production in ng/ml of supernatant ± SEM. Data in each panel are representative of three individual experiments.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2722173&req=5

fig2: Anergy-associated down-regulation of Th1 cytokines and increase in IL-10 production by CD4+ T cells during tolerization. Splenic CD4+ T cells were positively selected from untreated or i.n. MBP Ac1-9[4Y]–treated Tg4 Rag1−/− mice at 2 h after each peptide administration. 5 × 104 CD4+ T cells per well were cultured with 100 µg/ml of MBP Ac1-9[4K] in the presence of 105 autologous irradiated CD4 depleted splenocytes as APCs and, where indicated, 20 U/ml of rhIL-2. (A and B) Proliferative responses were measured at 72 h by 3[H]thymidine incorporation in the absence or presence of rhIL-2. Results are depicted as the mean thymidine incorporation ± SEM. (C and D) Cytokine responses of CD4+ T cells from the cultures described in A and B were measured by ELISA at 24 h (IL-2) and 72 h (IFN-γ and IL-10) after in vitro restimulation in the absence or presence of rhIL-2. Results are depicted as the mean cytokine production in ng/ml of supernatant ± SEM. Data in each panel are representative of three individual experiments.
Mentions: Although the serum cytokine profile provides an overall picture of the immune response to i.n. peptide administration, it does not allow discrimination of cytokine production by different cellular components of the immune system. We therefore investigated the contribution of CD4+ T cells to the aforementioned changes in serum cytokine concentrations. Tg4 CD4+ T cells were previously shown to respond to the first i.n. peptide treatment by initial proliferation and expansion accompanied by a cytokine burst (Burkhart et al., 1999). Indeed, splenocytes from Tg4 Rag1−/− mice, after successive i.n. MBP Ac1-9[4Y] treatments, showed a fourfold increase in their number at 2 h after the second treatment compared with untreated mice; the number of splenocytes decreased with subsequent i.n. peptide treatments (Fig. S1 A). The number of CD4+ T cells in the peptide-treated Tg4 Rag1−/− spleens increased with a similar pattern to that of whole splenocytes during the course of tolerance induction (Fig. S1 B). Given that CD4+ T cells from Tg4 mice treated repeatedly with i.n. peptide appear profoundly unresponsive (Sundstedt et al., 2003), we determined the kinetics of anergy induction as well as changes in the CD4+ T cell cytokine profile over the course of tolerization in Tg4 Rag1−/− mice. As shown in Fig. 2 A, CD4+ T cells from mice either left untreated or treated once with i.n. peptide proliferated vigorously in response to antigen restimulation in vitro. However, the level of CD4+ T cell proliferation declined dramatically upon subsequent i.n. peptide treatments (Fig. 2 A). The addition of recombinant human IL-2 (rhIL-2) to the unresponsive CD4+ T cells led to recovery of proliferation (Fig. 2 B), thereby demonstrating that the cells were in a state of clonal anergy. Supernatants from the CD4+ T cell proliferation assays were collected and analyzed for IL-2, IFN-γ, IL-4 and IL-10 levels by ELISA. Fig. 2 C shows changes in CD4+ T cell cytokine secretion after successive i.n. peptide treatments. CD4+ T cells from mice either left untreated or treated once with i.n. peptide secreted high levels of IL-2 and IFN-γ when restimulated with antigen in vitro. Levels of both cytokines decreased dramatically after just two i.n. peptide treatments (Fig. 2 C, top and middle, respectively). IL-10 was only detected in cultures of CD4+ T cells from mice treated three times or more, i.e., after the decrease in IL-2 and IFN-γ secretion (Fig. 2 C, bottom). A similar inverse relationship between IL-2 and IL-10 was also observed in the analysis of serum cytokines (Fig. 1). Although the addition of rhIL-2 restored proliferation (Fig. 2 A), it did not have an effect on endogenous IL-2 secretion and resulted in only a minor increase in IFN-γ production by CD4+ T cells isolated after the tenth i.n. peptide treatment (Fig. 2 D, top and middle, respectively). In contrast, the secretion of IL-10 was greatly augmented by the presence of rhIL-2 (Fig. 2 D, bottom). IL-4 remained below the detection level throughout the course of i.n. peptide treatment, irrespective of the addition of rhIL-2 (not depicted). These results support the findings of previous studies showing that IL-2 reverses the anergy of peptide-induced IL-10 T reg cells from mice treated with 10 doses of i.n. peptide without changing their cytokine secretion pattern (Anderson et al., 2005).

Bottom Line: Loss of proliferative capacity correlated with a switch from the Th1-associated cytokines IL-2 and interferon (IFN)-gamma to the regulatory cytokine IL-10.IL-10 T reg cells suppressed dendritic cell maturation, prevented Th1 cell differentiation, and thereby created a negative feedback loop for Th1-driven immune pathology.These findings demonstrate that Th1 responses can be self-limiting in the context of peripheral tolerance to a self-antigen.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Medicine, University of Bristol School of Medical Sciences, Bristol BS8 1TD, England, UK. Leona.Gabrysova@bristol.ac.uk

ABSTRACT
Regulation of the immune response to self- and foreign antigens is vitally important for limiting immune pathology associated with both infections and hypersensitivity conditions. Control of autoimmune conditions can be reinforced by tolerance induction with peptide epitopes, but the mechanism is not currently understood. Repetitive intranasal administration of soluble peptide induces peripheral tolerance in myelin basic protein (MBP)-specific TCR transgenic mice. This is characterized by the presence of anergic, interleukin (IL)-10-secreting CD4(+) T cells with regulatory function (IL-10 T reg cells). The differentiation pathway of peptide-induced IL-10 T reg cells was investigated. CD4(+) T cells became anergic after their second encounter with a high-affinity MBP peptide analogue. Loss of proliferative capacity correlated with a switch from the Th1-associated cytokines IL-2 and interferon (IFN)-gamma to the regulatory cytokine IL-10. Nevertheless, IL-10 T reg cells retained the capacity to produce IFN-gamma and concomitantly expressed T-bet, demonstrating their Th1 origin. IL-10 T reg cells suppressed dendritic cell maturation, prevented Th1 cell differentiation, and thereby created a negative feedback loop for Th1-driven immune pathology. These findings demonstrate that Th1 responses can be self-limiting in the context of peripheral tolerance to a self-antigen.

Show MeSH
Related in: MedlinePlus