Staphylococcus aureus fibronectin-binding protein specifically binds IgE from patients with atopic dermatitis and requires antigen presentation for cellular immune responses - 11/08/11

Doi : 10.1016/j.jaci.2011.02.034

Kavita Reginald, PhD ^a, Kerstin Westritschnig, MD ^a, Birgit Linhart, PhD ^a, Margarete Focke-Tejkl, PhD ^b, Beatrice Jahn-Schmid, PhD ^a, Julia Eckl-Dorna, MD, PhD ^c, Annice Heratizadeh, MD ^f, Angelika Stöcklinger, PhD ^g, Nadja Balic ^d, Susanne Spitzauer, MD ^d, Verena Niederberger, MD ^c, Thomas Werfel, MD ^f, Josef Thalhamer, PhD ^g, Stephan Weidinger, MD ⁱ, Natalija Novak, MD ^j, Markus Ollert, MD ^h,ⁱ, Alexander M. Hirschl, MD ^e, Rudolf Valenta, MD ^a,^b,^⁎
^a Division of Immunopathology, Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria
^b Christian Doppler Laboratory for Allergy Research, Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria
^c Department of Otorhinolaryngology, Medical University of Vienna, Vienna, Austria
^d Institute of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, Vienna, Austria
^e Department of Clinical Microbiology, Medical University of Vienna, Vienna, Austria
^f Department of Dermatology and Allergy, Division of Immunodermatology and Allergy Research, Hannover Medical School, Hannover, Germany
^g Department of Molecular Biology, Salzburg University, Salzburg, Austria
^h Clinical Research Division of Molecular and Clinical Allergotoxicology, Department of Dermatology and Allergy, Technische Universität München, Munich, Germany
ⁱ Department of Dermatology and Allergy, Technische Universität München, Munich, Germany
^j Department of Dermatology and Allergy, University of Bonn, Bonn, Germany

Reprint requests: Rudolf Valenta, MD, Division of Immunopathology, Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.

Abstract

Background

Staphylococcus aureus superinfections occur in more than 90% of patients with atopic dermatitis (AD) and aggravate skin inflammation. S aureus toxins lead to tissue damage and augment T-cell–mediated skin inflammation by a superantigen effect.

Objective

To characterize IgE-reactive proteins from S aureus.

Methods

A genomic S aureus library was screened with IgE from patients with AD for DNA clones coding for IgE-reactive antigens. One was identified as fibronectin-binding protein (FBP). Recombinant FBP was expressed in Escherichia coli, purified, and tested for specific IgE reactivity in patients with AD. Its allergenic activity was studied in basophil activation experiments and T-cell cultures. The in vivo allergenic activity was investigated by sensitizing mice.

Results

Using IgE from patients with AD for screening of a genomic S aureus library, an IgE-reactive DNA clone was isolated that coded for FBP. Recombinant FBP was expressed in E coli and purified. It reacted specifically with IgE from patients with AD and exhibited allergenic activity in basophil degranulation assays. FBP showed specific T-cell reactivity requiring antigen presentation and induced the secretion of proinflammatory cytokines from PBMCs. Mice sensitized with FBP mounted FBP-specific IgE responses, showed FBP-specific basophil degranulation as well as FBP-specific T-cell proliferation, and mixed T_h2/T_h1 cytokine secretion.

Conclusion

Evidence is provided that specific humoral and cellular immune responses to S aureus antigens dependent on antigen presentation represent a novel mechanism for S aureus–induced skin inflammation in AD. Furthermore, FBP may be used for the development of novel diagnostic and therapeutic strategies for S aureus infections.

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Key words : Staphylococcus aureus, atopic dermatitis, bacterial antigen, bacterial allergen, allergy, IgE, superinfection, superantigen

Abbreviations used : AD, APC, Con A, FBP, FbpA, r, RBL

Plan

Methods

Characterization of patients with AD

IgE immunoscreening of a genomic S aureus library with sera from patients with AD

Expression and purification of recombinant S aureus fibronectin-binding protein in Escherichia coli

IgE and IgG subclass reactivity of recombinant fibronectin-binding protein, IgE epitope mapping, and basophil degranulation experiments

FBP-specific lymphoproliferation and cytokine responses

Assay for measuring superantigen activity

FBP-specific T-cell proliferation using viable or fixed antigen-presenting cells

Allergic sensitization of mice with rFBP

Statistical analysis of data

Results

IgE-based isolation of a DNA encoding an S aureus FBP from a genomic S aureus library

Expression and purification of S aureus FBP in E coli

S aureus FBP is an IgE-reactive protein with allergenic activity containing major IgE epitopes in the DUF814 domain

S aureus FBP-specific IgG subclass response in patients with AD is indicative of a T_h2 response

S aureus FBP requires antigen presentation to induce specific lymphoproliferative responses and does not act as superantigen

S aureus FBP induces the release of proinflammatory cytokines IL-6, IFN-γ, and TNF-⍺ in cultured PBMCs

S aureus FBP induces an allergic immune response in mice

FBP-sensitized mice exhibit strong lymphoproliferative and T_h2 cytokine responses

Discussion

Supported by grants F1804, F1815, and F1818 from the Austrian Science Fund; the Christian Doppler Research Association, Vienna, Austria; and a research grant from Biomay, Vienna, Austria. B.J.-S. is supported by grant P20011-B13 from the Austrian Science Fund.

Disclosure of potential conflict of interest: V. Niederberger receives research support from the Austrian Science Fund (FWF). J. Thalhamer has consultant arrangements with Biomay AG and receives research support from Biomay AG, Christian Doppler Forschungsgesellschaft, and the Austrian Science Foundation. S. Weidinger receives research support from the German Research Council and the Bundesministerium für Bildung und Forschung. N. Novak receives research support from the German Research Council and is a lecturer for Phadia. R. Valenta receives research support from the Austrian Science Fund, the Christian Doppler Research Association, Biomay, and Phadia and has provided legal consultation or expert witness testimony in cases related to allergy therapy and allergy diagnosis. The rest of the authors have declared that they have no conflict of interest.

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Vol 128 - N° 1

P. 82 - juillet 2011 Retour au numéro

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Staphylococcus aureus fibronectin-binding protein specifically binds IgE from patients with atopic dermatitis and requires antigen presentation for cellular immune responses - 11/08/11

Abstract

Background

Objective

Methods

Results

Conclusion

Plan

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