Chronic infection with the protozoan Toxoplasma gondii prevents the development of experimental atopic dermatitis in mice

Background: Supporting the hypothesis thatT. gondii infection protects against allergy in humans we previously demonstrated that this infection can modulate not only the susceptibility to develop respiratory allergies in mice but also suppresses allergic responses at systemic level. This latter finding suggests that T. gondii infection could prevent the onset of other allergic diseases, such as atopic dermatitis. At present, few studies have investigated the modulation of atopic dermatitis by parasite infections. Objective: Here, we sought to investigate whether chronic infection with T. gondii is capable of modulating the development of atopic dermatitis. Methods: Chronically infected mice were sensitized by repeated epicutaneous ovalbumin administration. Skin histopathology, humoral response, cytokine production and innate type-II lymphoid cells (ILC2) were assessed. Results: A marked reduction in epidermal thickness and dermal inflammatory infiltrate along with a reduction in mast cell count was observed in infected mice compared to non-infected mice. These results correlated with a diminished TH2 and TH1 allergen specific response. Reduced type-II IL-4 and IL-5 cytokines were already detected during the first 24 h of allergen sensitization in splenocytes and draining lymph nodes from infected mice. Moreover, this reduced type-II profile in chronically infected animals correlated with diminished ILC2 number in draining lymph nodes. Conclusion: Chronic infection withT. gondii prevents the development of atopic dermatitis. The diminished susceptibility seems to result from changes in type-II innate immune response that may lead to the induction of a deficient TH2 response and consequently to a lower susceptibility to develop atopic dermatitis.Fil: Perrone Sibilia, Matías Damián. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; ArgentinaFil: Aldirico, María de Los Ángeles. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Soto, Ariadna Soledad. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Picchio, Mariano Sergio. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Sánchez, Vanesa Roxana. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Arcon, Nadia. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Moretta, Rosalia Ester. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Martín, Valentina. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Vanzulli, Silvia. Academia Nacional de Medicina de Buenos Aires. Instituto de Investigaciones Hematológicas "Mariano R. Castex"; ArgentinaFil: Fenoy, Ignacio Martín. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Goldman, Alejandra. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología. Centro de Estudios en Salud y Medio Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

rTgPI-1 treatment ameliorates lung histopathology.

<p>(A) BAL eosinophil count was performed on cytocentrifuge slides stained with a modified Wright-Giemsa stain. (B) Sections for microscopy were stained with Haematoxylin and PAS. Original magnification 100X. (C) A histological goblet cell score was obtained in Periodic acid- Schiff (PAS)-stained lung sections by examining 20 consecutive airways at 400x magnification. (D) An index of pathologic changes in H&E slides was obtained by scoring the inflammatory infiltrate around the airways and vessels for greatest severity and overall extent. Results for each group are expressed as the mean ± SEM.*<i>p</i> <0.05 and **<i>p</i> <0.01, α <i>p</i><0.01 vs N, ns: non-significant; ANOVA with Bonferroni's test <i>a posteriori</i>.</p

rTgPI-1 treatment attenuates airway hyperresponsiveness.

<p>Changes in pulmonary resistance in response to increasing doses of methacholine (0–30 mg/ml) were evaluated by invasive plethysmography. Results for each group are expressed as the mean ± SEM. ψ <i>p</i><0.01 OPI vs OO and O, Ѡ <i>p</i><0.001 OPI vs OO and O, φ <i>p</i><0.05 PI vs O, α <i>p</i><0.05 O and OO vs N; ANOVA with Bonferroni's test <i>a posteriori</i>.</p

Purification and activity of rTgPI-1.

<p>(A) SDS-PAGE of rTgPI-1 stained with Coomassie Blue. (B) Western blot of rTgPI-1 revealed with anti-His-Tag (lane 1) antibody or mouse anti-TgPI-1 serum (lane 2). (C) Analysis of inhibitory effect of rTgPI-1 on trypsin by gelatin substrate-SDS PAGE. All lanes contain 50 ng of trypsin. Lane 1, 1 μg of rTgPI-1; lane 2, control without rTgPI-1. Gelatinolytic activity was visualized by staining with Coomassie Brilliant Blue. The image was digitally inverted.</p

Schematic of asthma model and treatment protocol.

<p>Mice were sensitized by two injections of OVA/alum and challenged by exposure to aerosols of allergen on 3 consecutive days. Two days later mice were intranasally (in) treated with rTgPI-1 (PI) or rTgPI-1 plus OVA (OPI). Controls included non-sensitized mice treated with rTgPI-1 alone (N), and sensitized mice exposed intranasally to PBS (O) or OVA (OO). One week later, mice were re-challenged and analysis was performed 48 h after the last exposure.</p

Effect of rTgP-1 treatment on allergen-specific humoral response.

<p>Serum levels of OVA-specific IgE (dilution, 1:40), IgG1 (dilution, 1:64x10⁶) and IgG2a (dilution, 1:16x10³) antibodies were quantified in all experimental groups. Results are expressed as the mean ± SEM. *<i>p</i> <0.05, α <i>p</i><0.05 vs N, ns: non-significant; ANOVA with Bonferroni's test <i>a posteriori</i>.</p

Co-administration of rTgPI-1 plus allergen enhances generation of Tregs both local and systemically.

<p>Tregs were identified by flow cytometry. The percentage of FoxP3⁺CD4⁺ among total CD4⁺ cells from TLNC (A) or splenocytes (B) was calculated. Results for each group are expressed as the mean % CD4⁺FoxP3⁺/CD4⁺ ± SEM. *<i>p</i><0.05, **<i>p</i><0.01, ns: non-significant; ANOVA with Bonferroni's test <i>a posteriori</i>.</p

Local production of cytokines.

<p>Cytokine production by TLNC cells cultured <i>ex vivo</i> with OVA were analysed in all group of mice by ELISA. Results for each group are expressed as the mean ± SEM *<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001, α p<0.001 vs N, ns: non-significant; ANOVA with Bonferroni’s test <i>a posteriori</i>.</p

rTgPI-1 treatment suppresses cell proliferation.

<p>Proliferative responses of TLNC (A and B) or splenocytes (C and D) were determined by ³H-thymidine incorporation upon stimulation with OVA. Results for each group are expressed as the mean ± SEM. *<i>p</i><0.05, **<i>p</i><0.01, α <i>p</i><0.001 vs N, ns: non-significant; ANOVA with Bonferroni's test <i>a posteriori</i>.</p