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Cross-reactivity of autoantibodies from patients with epidermolysis bullosa acquisita with murine collagen VII

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Abstract

The pathomechanism of antibody-mediated tissue damage in autoimmune diseases can be best studied in experimental models by passively transferring specific autoantibodies into animals. The reproduction of the disease in animals depends on several factors, including the cross-reactivity of patient autoantibodies with the animal tissue. Here, we show that autoantibodies from patients with epidermolysis bullosa acquisita (EBA), a subepidermal autoimmune blistering disease, recognize multiple epitopes on murine collagen VII. Indirect immunofluorescence microscopy revealed that EBA patients’ IgG cross-reacts with mouse skin. Overlapping, recombinant fragments of murine collagen VII were used to characterize the reactivity of EBA sera and to map the epitopes on the murine antigen by ELISA and immunoblotting. The patients’ autoantibody binding to murine collagen VII triggered pathogenic events as demonstrated by a complement fixing and an ex vivo granulocyte-dependent dermal–epidermal separation assay. These findings should greatly facilitate the development of improved disease models and novel therapeutic strategies.

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Acknowledgments

The authors acknowledge support by grants from the Deutsche Forschungsgemeinschaft SI-1281/2-1, SI-1281/4-1 and BIOSS-B13 (CS), from the Medical Faculty of the University of Freiburg (C.S.) and by an ERASMUS stipend (V.F.). We thank Dr. Leena Bruckner-Tuderman, Freiburg, Germany, for critical reading of the manuscript and helpful advice.

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Authors and Affiliations

  1. Department of Dermatology, University of Freiburg, Freiburg, Germany

    Kinga Csorba, Alina Sesarman, Eva Oswald, Vasile Feldrihan, Anja Fritsch & Cassian Sitaru

  2. Faculty of Biology, University of Freiburg, Freiburg, Germany

    Kinga Csorba & Eva Oswald

  3. Department of Dermatology, Kurume University, Kurume, Japan

    Takashi Hashimoto

  4. Universitäts-Hautklinik, Hauptstr. 7, 79104, Freiburg, Germany

    Cassian Sitaru

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  1. Kinga Csorba
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  2. Alina Sesarman
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Correspondence to Cassian Sitaru.

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Supplementary Fig. 1

Expression of the recombinant GST-fusion and His-tagged fragments of murine collagen VII. (a) Equimolar amounts of GST (lane 2), GST-mCVII-1 (lane 3), GST-mCVII-2 (lane 4), GST-mCVII-3 (lane 5), GST-mCVII-4 (lane 6), GST-mCVII-5 (lane 7), GST-mCVII-Z (lane 8), GST-mCVII-Cr (lane 9) as well as (b) His-DHFR (lane 2), His-mCVII-1 (lane 3), His-mCVII-2 (lane 4), His-mCVII-3 (lane 5), His-mCVII-4 (lane 6), His-mCVII-5 (lane 7), His-mCVII-Z (lane 8) and His-mCVII-Cr (lane 9) were separated by 12% SDS-PAGE and stained with BioSafe Coomassie blue. The GST and His labelled proteins migrated according to their expected molecular weight. (c) The His-DHFR (lane 1) and the His tagged recombinant murine collagen VII fragments (lanes 2-8) were separated on a 12% polyacrylamide gel. Immunoblot analysis with a mouse IgG raised against RGS-His (Qiagen), in a first step, and with a HRP conjugated anti mouse antibody (BioRad) in a second step resulted in detection of all His labelled proteins. (TIF 13.2 kb)

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Csorba, K., Sesarman, A., Oswald, E. et al. Cross-reactivity of autoantibodies from patients with epidermolysis bullosa acquisita with murine collagen VII. Cell. Mol. Life Sci. 67, 1343–1351 (2010). https://doi.org/10.1007/s00018-009-0256-3

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  • DOI: https://doi.org/10.1007/s00018-009-0256-3

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