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IL-28 supplants requirement for T(reg) cells in protein sigma1-mediated protection against murine experimental autoimmune encephalomyelitis (EAE).

Rynda A, Maddaloni M, Ochoa-Repáraz J, Callis G, Pascual DW - PLoS ONE (2010)

Bottom Line: Conventional methods to induce tolerance in humans have met with limited success.To test whether single-dose tolerance can be induced to treat EAE, proteolipid protein (PLP(130-151)) was genetically fused to OVA to psigma1 (PLP:OVA-psigma1) and shown to significantly ameliorate EAE, suppressing proinflammatory cytokines by IL-10(+) forkhead box P3 (FoxP3)(+) CD25(+)CD4(+) T(reg) and IL-4(+)CD25(-)CD4(+) Th2 cells.Thus, psigma1-based therapy can resolve EAE independently of or dependently upon CD25 and assigns IL-28 as an alternative therapy for autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Veterinary Molecular Biology, Montana State University, Bozeman, Montana, United States of America.

ABSTRACT
Conventional methods to induce tolerance in humans have met with limited success. Hence, efforts to redirect tolerogen uptake using reovirus adhesin, protein sigma 1 (psigma1), may circumvent these shortcomings based upon the recent finding that when reovirus psigma1 is engineered to deliver chicken ovalbumin (OVA) mucosally, tolerance is obtained, even with a single dose. To test whether single-dose tolerance can be induced to treat EAE, proteolipid protein (PLP(130-151)) was genetically fused to OVA to psigma1 (PLP:OVA-psigma1) and shown to significantly ameliorate EAE, suppressing proinflammatory cytokines by IL-10(+) forkhead box P3 (FoxP3)(+) CD25(+)CD4(+) T(reg) and IL-4(+)CD25(-)CD4(+) Th2 cells. IL-10R or IL-4 neutralization reversed protection to EAE conferred by PLP:OVA-psigma1, and adoptive transfer of Ag-specific T(reg) or Th2 cells restored protection against EAE in recipients. Upon assessment of each relative participant, functional inactivation of CD25 impaired PLP:OVA-psigma1's protective capacity, triggering TGF-beta-mediated inflammation; however, concomitant inactivation of TGF-beta and CD25 reestablished PLP:OVA-psigma1-mediated protection by IL-28-producing FoxP3(+)CD25(-)CD4(+) T cells. Thus, psigma1-based therapy can resolve EAE independently of or dependently upon CD25 and assigns IL-28 as an alternative therapy for autoimmunity.

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Abated protection against EAE in anti-CD25 mAb-treated PLP:OVA-pσ1-dosed mice is restored upon TGF-β co-neutralization.A. Experimental design for neutralization of TGF-β and CD25. B. PBS-dosed mice, independent of treatment, developed expected course of EAE. C. Treatment with anti-CD25 mAb abrogated PLP:OVA-pσ1-mediated protection against EAE, but concomitant treatment with anti-TGF-β and anti-CD25 mAbs restored PLP:OVA-pσ1-induced protection. Mean of 10 mice per group is shown. B and C. * P<0.05 vs. IgG-dosed mice. D. HNLN CD4+ T cells on day 10 post-EAE induction were cultured with PLP139–151 peptide for 72 h. PBS-dosed mice independent of Ab treatment produced elevated proinflammatory cytokines IFN-γ, IL-17, and IL-21, and little to no IL-4, IL-10, IL-13, and TGF-β. However, levels of IFN-γ and IL-17 produced by CD4+ T cells were diminished in PBS + anti-TGF-β-treated mice when compared to PBS + IgG- treated mice. PLP:OVA-pσ1-protected mice treated with IgG, anti-TGF-β mAb, or anti-TGF-β + anti-CD25 mAbs produced enhanced IL-4 and IL-28, but minimal proinflammatory cytokines. Additionally, mice dosed with PLP:OVA-pσ1 + anti-CD25 + anti-TGF-β mAbs produced significantly more IL-13 and IL-4 than diseased PLP:OVA-pσ1 + anti-CD25-dosed mice. Mean + SEM of 5 mice per group is shown * P<0.05 for the PLP:OVA-pσ1 + anti-TGF-β + anti-CD25 vs. PLP:OVA-pσ1 + IgG and PLP:OVA-pσ1 + anti-CD25.
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pone-0008720-g005: Abated protection against EAE in anti-CD25 mAb-treated PLP:OVA-pσ1-dosed mice is restored upon TGF-β co-neutralization.A. Experimental design for neutralization of TGF-β and CD25. B. PBS-dosed mice, independent of treatment, developed expected course of EAE. C. Treatment with anti-CD25 mAb abrogated PLP:OVA-pσ1-mediated protection against EAE, but concomitant treatment with anti-TGF-β and anti-CD25 mAbs restored PLP:OVA-pσ1-induced protection. Mean of 10 mice per group is shown. B and C. * P<0.05 vs. IgG-dosed mice. D. HNLN CD4+ T cells on day 10 post-EAE induction were cultured with PLP139–151 peptide for 72 h. PBS-dosed mice independent of Ab treatment produced elevated proinflammatory cytokines IFN-γ, IL-17, and IL-21, and little to no IL-4, IL-10, IL-13, and TGF-β. However, levels of IFN-γ and IL-17 produced by CD4+ T cells were diminished in PBS + anti-TGF-β-treated mice when compared to PBS + IgG- treated mice. PLP:OVA-pσ1-protected mice treated with IgG, anti-TGF-β mAb, or anti-TGF-β + anti-CD25 mAbs produced enhanced IL-4 and IL-28, but minimal proinflammatory cytokines. Additionally, mice dosed with PLP:OVA-pσ1 + anti-CD25 + anti-TGF-β mAbs produced significantly more IL-13 and IL-4 than diseased PLP:OVA-pσ1 + anti-CD25-dosed mice. Mean + SEM of 5 mice per group is shown * P<0.05 for the PLP:OVA-pσ1 + anti-TGF-β + anti-CD25 vs. PLP:OVA-pσ1 + IgG and PLP:OVA-pσ1 + anti-CD25.

Mentions: Treg cell neutralization exacerbated EAE, negating the protective capacity of PLP:OVA-pσ1, resulting in enhanced TGF-β production (Figure 4B) and implicating a proinflammatory role of this cytokine in EAE. To address TGF-β's participation in EAE development subsequent CD25 neutralization, PLP:OVA-pσ1- and PBS-dosed mice were treated in vivo with anti-TGF-β mAb, anti-CD25 mAb, both, or IgG (Figure 5A). No difference in onset or disease severity was observed between mice dosed with PBS and treated with the different combinations of mAbs, except those mice dosed with PBS + anti-TGF-β mAb recovered sooner from the acute disease (Figure 5B). In contrast to PLP:OVA-pσ1 + anti-CD25 mAb-treated mice, which developed a very aggressive disease, mice given PLP:OVA-pσ1 + any of the remaining Ab treatments developed only very mild disease with the average peak clinical score of ∼1.5; all of these mice recovered from acute EAE (Figure 5C). Thus, co-neutralization of CD25 and TGF-β in PLP:OVA-pσ1-treated mice resembled ameliorated disease, as seen in PLP:OVA-pσ1 + IgG-dosed mice. These results showed that mice functionally neutralized of their CD25+ Treg cells in the presence of tolerogen develop a more aggressive, TGF-β-dependent EAE. The suppressive activity of PLP:OVA-pσ1 could only be restored upon co-neutralization of TGF-β. These findings corroborate the results in Figure 3B in which minimal to no TGF-β was detected in PBS-treated mice, suggesting that TGF-β has a minimal role in PLP-mediated EAE.


IL-28 supplants requirement for T(reg) cells in protein sigma1-mediated protection against murine experimental autoimmune encephalomyelitis (EAE).

Rynda A, Maddaloni M, Ochoa-Repáraz J, Callis G, Pascual DW - PLoS ONE (2010)

Abated protection against EAE in anti-CD25 mAb-treated PLP:OVA-pσ1-dosed mice is restored upon TGF-β co-neutralization.A. Experimental design for neutralization of TGF-β and CD25. B. PBS-dosed mice, independent of treatment, developed expected course of EAE. C. Treatment with anti-CD25 mAb abrogated PLP:OVA-pσ1-mediated protection against EAE, but concomitant treatment with anti-TGF-β and anti-CD25 mAbs restored PLP:OVA-pσ1-induced protection. Mean of 10 mice per group is shown. B and C. * P<0.05 vs. IgG-dosed mice. D. HNLN CD4+ T cells on day 10 post-EAE induction were cultured with PLP139–151 peptide for 72 h. PBS-dosed mice independent of Ab treatment produced elevated proinflammatory cytokines IFN-γ, IL-17, and IL-21, and little to no IL-4, IL-10, IL-13, and TGF-β. However, levels of IFN-γ and IL-17 produced by CD4+ T cells were diminished in PBS + anti-TGF-β-treated mice when compared to PBS + IgG- treated mice. PLP:OVA-pσ1-protected mice treated with IgG, anti-TGF-β mAb, or anti-TGF-β + anti-CD25 mAbs produced enhanced IL-4 and IL-28, but minimal proinflammatory cytokines. Additionally, mice dosed with PLP:OVA-pσ1 + anti-CD25 + anti-TGF-β mAbs produced significantly more IL-13 and IL-4 than diseased PLP:OVA-pσ1 + anti-CD25-dosed mice. Mean + SEM of 5 mice per group is shown * P<0.05 for the PLP:OVA-pσ1 + anti-TGF-β + anti-CD25 vs. PLP:OVA-pσ1 + IgG and PLP:OVA-pσ1 + anti-CD25.
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pone-0008720-g005: Abated protection against EAE in anti-CD25 mAb-treated PLP:OVA-pσ1-dosed mice is restored upon TGF-β co-neutralization.A. Experimental design for neutralization of TGF-β and CD25. B. PBS-dosed mice, independent of treatment, developed expected course of EAE. C. Treatment with anti-CD25 mAb abrogated PLP:OVA-pσ1-mediated protection against EAE, but concomitant treatment with anti-TGF-β and anti-CD25 mAbs restored PLP:OVA-pσ1-induced protection. Mean of 10 mice per group is shown. B and C. * P<0.05 vs. IgG-dosed mice. D. HNLN CD4+ T cells on day 10 post-EAE induction were cultured with PLP139–151 peptide for 72 h. PBS-dosed mice independent of Ab treatment produced elevated proinflammatory cytokines IFN-γ, IL-17, and IL-21, and little to no IL-4, IL-10, IL-13, and TGF-β. However, levels of IFN-γ and IL-17 produced by CD4+ T cells were diminished in PBS + anti-TGF-β-treated mice when compared to PBS + IgG- treated mice. PLP:OVA-pσ1-protected mice treated with IgG, anti-TGF-β mAb, or anti-TGF-β + anti-CD25 mAbs produced enhanced IL-4 and IL-28, but minimal proinflammatory cytokines. Additionally, mice dosed with PLP:OVA-pσ1 + anti-CD25 + anti-TGF-β mAbs produced significantly more IL-13 and IL-4 than diseased PLP:OVA-pσ1 + anti-CD25-dosed mice. Mean + SEM of 5 mice per group is shown * P<0.05 for the PLP:OVA-pσ1 + anti-TGF-β + anti-CD25 vs. PLP:OVA-pσ1 + IgG and PLP:OVA-pσ1 + anti-CD25.
Mentions: Treg cell neutralization exacerbated EAE, negating the protective capacity of PLP:OVA-pσ1, resulting in enhanced TGF-β production (Figure 4B) and implicating a proinflammatory role of this cytokine in EAE. To address TGF-β's participation in EAE development subsequent CD25 neutralization, PLP:OVA-pσ1- and PBS-dosed mice were treated in vivo with anti-TGF-β mAb, anti-CD25 mAb, both, or IgG (Figure 5A). No difference in onset or disease severity was observed between mice dosed with PBS and treated with the different combinations of mAbs, except those mice dosed with PBS + anti-TGF-β mAb recovered sooner from the acute disease (Figure 5B). In contrast to PLP:OVA-pσ1 + anti-CD25 mAb-treated mice, which developed a very aggressive disease, mice given PLP:OVA-pσ1 + any of the remaining Ab treatments developed only very mild disease with the average peak clinical score of ∼1.5; all of these mice recovered from acute EAE (Figure 5C). Thus, co-neutralization of CD25 and TGF-β in PLP:OVA-pσ1-treated mice resembled ameliorated disease, as seen in PLP:OVA-pσ1 + IgG-dosed mice. These results showed that mice functionally neutralized of their CD25+ Treg cells in the presence of tolerogen develop a more aggressive, TGF-β-dependent EAE. The suppressive activity of PLP:OVA-pσ1 could only be restored upon co-neutralization of TGF-β. These findings corroborate the results in Figure 3B in which minimal to no TGF-β was detected in PBS-treated mice, suggesting that TGF-β has a minimal role in PLP-mediated EAE.

Bottom Line: Conventional methods to induce tolerance in humans have met with limited success.To test whether single-dose tolerance can be induced to treat EAE, proteolipid protein (PLP(130-151)) was genetically fused to OVA to psigma1 (PLP:OVA-psigma1) and shown to significantly ameliorate EAE, suppressing proinflammatory cytokines by IL-10(+) forkhead box P3 (FoxP3)(+) CD25(+)CD4(+) T(reg) and IL-4(+)CD25(-)CD4(+) Th2 cells.Thus, psigma1-based therapy can resolve EAE independently of or dependently upon CD25 and assigns IL-28 as an alternative therapy for autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Veterinary Molecular Biology, Montana State University, Bozeman, Montana, United States of America.

ABSTRACT
Conventional methods to induce tolerance in humans have met with limited success. Hence, efforts to redirect tolerogen uptake using reovirus adhesin, protein sigma 1 (psigma1), may circumvent these shortcomings based upon the recent finding that when reovirus psigma1 is engineered to deliver chicken ovalbumin (OVA) mucosally, tolerance is obtained, even with a single dose. To test whether single-dose tolerance can be induced to treat EAE, proteolipid protein (PLP(130-151)) was genetically fused to OVA to psigma1 (PLP:OVA-psigma1) and shown to significantly ameliorate EAE, suppressing proinflammatory cytokines by IL-10(+) forkhead box P3 (FoxP3)(+) CD25(+)CD4(+) T(reg) and IL-4(+)CD25(-)CD4(+) Th2 cells. IL-10R or IL-4 neutralization reversed protection to EAE conferred by PLP:OVA-psigma1, and adoptive transfer of Ag-specific T(reg) or Th2 cells restored protection against EAE in recipients. Upon assessment of each relative participant, functional inactivation of CD25 impaired PLP:OVA-psigma1's protective capacity, triggering TGF-beta-mediated inflammation; however, concomitant inactivation of TGF-beta and CD25 reestablished PLP:OVA-psigma1-mediated protection by IL-28-producing FoxP3(+)CD25(-)CD4(+) T cells. Thus, psigma1-based therapy can resolve EAE independently of or dependently upon CD25 and assigns IL-28 as an alternative therapy for autoimmunity.

Show MeSH
Related in: MedlinePlus