TY - JOUR
T1 - Prevention of intestinal allergy in mice by rflaA:Ova is associated with enforced antigen processing and TLR5-dependent IL-10 secretion by mDC
AU - Schülke, Stefan
AU - Wolfheimer, Sonja
AU - Gadermaier, Gabriele
AU - Wangorsch, Andrea
AU - Siebeneicher, Susanne
AU - Briza, Peter
AU - Spreitzer, Ingo
AU - Schiller, Dirk
AU - Loeschner, Bettina
AU - Uematsu, Satoshi
AU - Ryffel, Bernard
AU - Akira, Shizuo
AU - Waibler, Zoe
AU - Vieths, Stefan
AU - Toda, Masako
AU - Scheurer, Stephan
N1 - Funding Information:
The following activities of SV could be perceived as a potential conflict of interest concerning this manuscript: In the past five years SV has received speakers honorarium from Phadia (now Thermofisher Scientific), Uppsala, a honorarium for organising and chairing an educational seminar on food allergens from Fresenius Academy, Dortmund, honoraria for acting as expert reviewer for the Allergen Online Database of the Food Allergy Resource and Research Program, Lincoln NE, and consultancy fees by the Institute for Product Quality, Berlin In the past five years the Division of Allergology has received research grants from the German Research Foundation, the EU, the German Federal Ministry for Research and Education, Monsanto Company, and Pioneer Hi-Bred International Inc with SV acting as principal investigator. There are no further patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials, as detailed online in the guide for authors.
PY - 2014/2/7
Y1 - 2014/2/7
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=84895730381&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84895730381&partnerID=8YFLogxK
U2 - 10.1371/journal.pone.0087822
DO - 10.1371/journal.pone.0087822
M3 - Article
C2 - 24516564
AN - SCOPUS:84895730381
SN - 1932-6203
VL - 9
JO - PLoS One
JF - PLoS One
IS - 2
M1 - e87822
ER -
