Immunosuppression in early postnatal days induces persistent and allergen-specific immune tolerance to asthma in adult mice

Abstract

Bronchial asthma is a chronic airway inflammatory condition with high morbidity, and effective treatments for asthma are limited. Allergen-specific immunotherapy can only induce peripheral immune tolerance and is not sustainable. Exploring new therapeutic strategies is of great clinical importance. Recombinant adenovirus (rAdV) was used as a vector to make cells expressing cytotoxic T lymphocyte-associated antigen-4-immunoglobulin (CTLA4Ig) a soluble CTLA4 immunoglobulin fusion protein. Dendritic cells (DCs) were modified using the rAdVs together with allergens. Then these modified DCs were transplanted to mice before allergen sensitization. The persistence and specificity of immune tolerance were evaluated in mice challenged with asthma allergens at 3 and 7 months. DCs modified by CTLA4Ig showed increased IL-10 secretion, decreased IL-12 secretion, and T cell stimulation in vitro. Mice treated with these DCs in the early neonatal period developed tolerance against the allergens that were used to induce asthma in the adult stage. Asthma symptoms, lung damage, airway reactivity, and inflammatory response all improved. Humoral immunity indices showed that this therapeutic strategy strongly suppressed mice immune responses and was maintained for as long as 7 months. Furthermore, allergen cross-sensitization and challenge experiments demonstrated that this immune tolerance was allergen-specific. Treatment with CTLA4Ig modified DCs in the early neonatal period, inducing persistent and allergen-specific immune tolerance to asthma in adult mice. Our results suggest that it may be possible to develop a vaccine for asthma.

Fig 1. In vitro evaluation of the function of DCs modified by rAdV-CTLA4Ig.

A, IL-10 (left Y-axis) and IL-12 (right Y-axis) secretion of modified DCs. B, characteristics of T cell stimulation of modified DCs evaluated by MLR. MLR, mixed lymphocyte reaction.

Fig 2. Persistence of immune tolerance to asthma antigens in the adult stage.

Saline and OVA represented mice treated, sensitized and challenged by saline and OVA respectively. OVA+rAdV-CTLA4Ig and OVA+rAdV-GFP represented mice treated with OVA+rAdV-CTLA4Ig and OVA+rAdV-GFP in the neonatal period sensitized and challenged by OVA at 3 and 7 months old. Each group contains 10 mice. A, regimen of sensitization, challenge, and treatment. B, representative histopathological characteristics of mice lung: upper, lungs of 3-month-old mice; lower, lungs of 7-month-old mice. C, airway reactivity testing in 3-month-old mice. * indicates significantly low airway reactivity of OVA+rAdV-CTLA4Ig treated mice versus the asthma mice (OVA group). D. airway reactivity testing in 7-month-old mice. * indicates significantly low airway reactivity of OVA+rAdV-CTLA4Ig treated mice versus the asthma mice (OVA group).

Fig 3. Specificity assessment.

To evaluate whether the persistent immune tolerance was allergen-specific, the neonatal mice were treated with OVA+rAdV-CTLA4Ig-modified DCs or Derp+rAdV-CTLA4Ig-modified DCs, 3 months later, the specificity of the therapeutic strategy was assessed by allergen cross challenge. Each group contains 10 mice. Saline, OVA and Derp represented mice treated, sensitized and challenged by saline, OVA and Derp respectively. OVA+rAdV-CTLA4Ig+OVA represented mice treated with rAdV-CTLA4Ig, sensitized by OVA and challenged by OVA. Derp+rAdV-CTLA4Ig+OVA represented mice treated with rAdV-CTLA4Ig, sensitized by Derp and challenged by OVA. Derp+rAdV-CTLA4Ig+Derp represented mice treated with rAdV-CTLA4Ig, sensitized by Derp and challenged by Derp. OVA+rAdV-CTLA4Ig+Derp represented mice treated with rAdV-CTLA4Ig, sensitized by OVA and challenged by Derp. A, the regimen of sensitization, challenge, and treatment in specificity assessment. B, airway reactivity testing of OVA-sensitized mice challenged by OVA or Derp. C, airway reactivity testing of Derp-sensitized mice challenged with Derp or OVA.

Fig 4. Humoral immunological changes in the specificity assessment.

For easy comparisons, the rates of change in IL-4, IL-10, IFN-γ, and IgE in both BALF and sera were adjusted by the values of the control (sensitized and challenged by saline). * indicates statistical significance of each comparison. A, the increasing rates of IL-4 in the regimens of sensitization, challenge, and treatment. B, IFN-γ decreasing rates in the regimens of sensitization, challenge, and treatment. C, IL-10 changes in the regimens of sensitization, challenge, and treatment. D, the increasing rates of IgE in the regimens of sensitization, challenge, and treatment.