Identification of GP2, the major zymogen granule membrane glycoprotein, as the autoantigen of pancreatic antibodies in Crohn’s disease
- D Roggenbuck1,
- G Hausdorf2,
- L Martinez-Gamboa2,
- D Reinhold3,
- T Büttner1,
- P R Jungblut4,
- T Porstmann5,
- M W Laass6,
- J Henker6,
- C Büning7,
- E Feist2,
- K Conrad8
- 1GA Generic Assays GmbH, Dahlewitz, Germany
- 2Department of Rheumatology and Clinical Immunology, Charité Universitätsmedizin Berlin, Berlin, Germany
- 3Institute of Molecular and Clinical Immunology, Otto-von-Guericke University Magdeburg, Magdeburg, Germany
- 4Max Planck Institute for Infection Biology, Berlin, Germany
- 5Seramun Diagnostica GmbH, Heidesee, Germany
- 6Clinic of Pediatrics, Technical University Dresden, Dresden, Germany
- 7Department of Gastroenterology, Hepatology, and Endocrinology, Charité Universitätsmedizin Berlin, Berlin, Germany
- 8Institute of Immunology, Technical University Dresden, Dresden, Germany
- Correspondence to Dr K Conrad, Institute of Immunology, Technical University Dresden, Fetscherstraße 74, 01307 Dresden, Germany; k_conrad{at}mail.zih.tu-dresden.de
- Revised 29 May 2009
- Accepted 9 June 2009
- Published Online First 22 June 2009
Abstract
Backround and aims: The aetiopathogenesis of Crohn’s disease, an inflammatory bowel disease (IBD), is not yet fully understood. Autoimmune mechanisms are thought to play a role in the development of Crohn’s disease, but the target antigens and the underlying pathways have not been sufficiently identified.
Methods: Based on data from immunoblotting and matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry, the major antigenic target of pancreatic autoantibodies (PABs), which are specific for Crohn’s disease, was identified. Specificity of autoantibody reactivity was confirmed by enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence (IIF) using purified rat and human recombinant GP2 synthesised in transiently transfected mammalian HEK 293 cells. Real-time polymerase chain reaction (rt-PCR) and IIF were used to detect mRNA and antigen localisation in human colon biopsies.
Results: The major zymogen granule membrane glycoprotein 2 (GP2) was identified as the autoantigen of PABs in Crohn’s disease. PAB-positive sera from patients with Crohn’s disease (n = 42) displayed significantly higher IgG reactivity to rat GP2 in ELISA than either PAB-negative sera (n = 31), or sera from patients with ulcerative colitis (n = 49), or sera from blood donors (n = 69) (p<0.0001, respectively). Twenty-eight (66%) and 18 (43%) of 42 PAB-positive sera demonstrated IgG and IgA reactivity to human recombinant GP2 in IIF, respectively. Patients with PAB-negative Crohn’s disease (n = 31) were not reactive. GP2 mRNA transcription was significantly higher in colon biopsies from patients with Crohn’s disease (n = 4) compared to patients with ulcerative colitis (n = 4) (p = 0.0286). Immunochemical staining confirmed GP2 expression in human colon biopsies from patients with Crohn’s disease.
Conclusion: Anti-GP2 autoantibodies constitute novel Crohn’s disease-specific markers, the quantification of which could significantly improve the serological diagnosis of IBD. The expression of GP2 in human enterocytes suggests an important role for anti-GP2 response in the pathogenesis of Crohn’s disease.
Footnotes
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Funding This work was supported by the Brandenburg Ministry of Economics and European Union grant 80130073.
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Competing interests DR is a shareholder of GA Generic Assays GmbH. TP is a shareholder of both Seramun Diagnostica GmbH and GA Generic Assays GmbH. Both companies are diagnostic manufacturers. The remaining authors have no competing interests.
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Provenance and peer review Not commissioned; externally peer reviewed.
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Ethics approval This study was approved by the local ethics committee of the Medical Faculty, Technical University Dresden, on 3 May 2007. The animals used in this study were handled according to German regulations concerning the protection of animals.
