Prevention of intestinal allergy in mice by rflaA:Ova is associated with enforced antigen processing and TLR5-dependent IL-10 secretion by mDC

Stefan Schülke; Sonja Wolfheimer; Gabriele Gadermaier; Andrea Wangorsch; Susanne Siebeneicher; Peter Briza; Ingo Spreitzer; Dirk Schiller; Bettina Loeschner; Satoshi Uematsu; Bernard Ryffel; Shizuo Akira; Zoe Waibler; Stefan Vieths; Masako Toda; Stephan Scheurer

doi:10.1371/journal.pone.0087822

Prevention of intestinal allergy in mice by rflaA:Ova is associated with enforced antigen processing and TLR5-dependent IL-10 secretion by mDC

PLoS One. 2014 Feb 7;9(2):e87822. doi: 10.1371/journal.pone.0087822. eCollection 2014.

Authors

Stefan Schülke¹, Sonja Wolfheimer¹, Gabriele Gadermaier², Andrea Wangorsch¹, Susanne Siebeneicher³, Peter Briza², Ingo Spreitzer⁴, Dirk Schiller¹, Bettina Loeschner⁴, Satoshi Uematsu⁵, Bernard Ryffel⁶, Shizuo Akira⁵, Zoe Waibler⁷, Stefan Vieths¹, Masako Toda³, Stephan Scheurer¹

Affiliations

¹ Division of Allergology, Paul-Ehrlich-Institut, Langen, Hessen, Germany.
² Christian Doppler Laboratory for Allergy Diagnosis and Therapy, Department of Molecular Biology, University of Salzburg, Salzburg, Austria.
³ Junior Research Group "Experimental Allergy Models", Paul-Ehrlich-Institut, Langen, Hessen, Germany.
⁴ Division of Microbiology, Paul-Ehrlich-Institut, Langen, Hessen, Germany.
⁵ Department of Host Defense, Research Institute for Microbial Diseases, Osaka University, Osaka, Osaka Prefecture, Japan.
⁶ CNRS Department of Molecular and Experimental Immunology and Neurogenetics, Allergy and Lung Inflammation, University of Orléans, Orléans, Loiret département, France.
⁷ Junior Research Group "Novel vaccination strategies and early immune responses", Paul-Ehrlich-Institut, Langen, Hessen, Germany.

Abstract

Conjugated vaccines consisting of flagellin and antigen activate TLR5 and induce strong innate and adaptive immune responses. Objective of the present study was to gain further insight into the mechanisms by which flagellin fusion proteins mediate their immune modulating effects. In a mouse model of Ova-induced intestinal allergy a fusion protein of flagellin and Ova (rflaA:Ova) was used for intranasal and intraperitoneal vaccination. Aggregation status of flaA, Ova and flaA:Ova were compared by light scattering, uptake of fluorescence labeled proteins into mDC was analyzed, processing was investigated by microsomal digestion experiments. Mechanism of DC-activation was investigated using proteasome and inflammasome inhibitors. Immune responses of wildtype, IL-10(-/-), TLR5(-/-) mDCs and Ova-transgenic T cells were investigated. Mucosal and i.p.-application of rflaA:Ova were able to prevent allergic sensitization, suppress disease-related symptoms, prevent body weight loss and reduction in food uptake. Intranasal vaccination resulted in strongest suppression of Ova-specific IgE production. These protective effects were associated with increased aggregation of rflaA:Ova and accompanied by tenfold higher uptake rates into mDC compared to the mixture of both proteins. Microsomal digestion showed that stimulation with rflaA:Ova resulted in faster degradation and the generation of different peptides compared to rOva. rflaA:Ova-mediated activation of mDC could be suppressed in a dose-dependent manner by the application of both inflammasome and proteasome inhibitors. Using TLR5(-/-) mDC the rflaA:Ova induced IL-10 secretion was shown to be TLR5 dependent. In co-cultures of IL-10(-/-) mDC with DO11.10 T cells the lack of rflaA:Ova-mediated IL-10 secretion resulted in enhanced levels of both TH2 (IL-4, IL-5) and TH1 (IL-2 and IFN-y) cytokines. In summary, mucosal vaccination with flaA:Ova showed strongest preventive effect. Stimulation with rflaA:Ova results in strong immune modulation mediated by enhanced uptake of the aggregated fusion protein, likely resulting in a different processing by DC as well as stronger TLR5 mediated cell activation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adaptive Immunity / immunology
Animals
Antigen Presentation / immunology*
Disease Models, Animal
Hypersensitivity / prevention & control*
Interleukin-10 / immunology*
Intestines / immunology*
Mice
Toll-Like Receptor 5 / immunology*
Vaccination / methods*

Substances

Toll-Like Receptor 5
Interleukin-10

Grants and funding

The study was funded by Deutsche Forschungsgemeinschaft (German Research Foundation) DFG SCHE637/3. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.