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Original article
T-cell repertoires in refractory coeliac disease
  1. Julia Ritter1,
  2. Karin Zimmermann1,
  3. Korinna Jöhrens1,
  4. Stefanie Mende1,
  5. Anke Seegebarth1,
  6. Britta Siegmund2,
  7. Steffen Hennig3,
  8. Kremena Todorova4,
  9. Andreas Rosenwald5,
  10. Severin Daum2,
  11. Michael Hummel1,
  12. Michael Schumann2,6,7
  1. 1Institute of Pathology, Charité—University Medicine, Berlin, Germany
  2. 2Department of Gastroenterology, Infectious Diseases and Rheumatology, Charité—University Medicine, Berlin, Germany
  3. 3HS Diagnomics GmbH, Berlin, Germany
  4. 4Center for Tumor Medicine, Charité—University Medicine, Berlin, Germany
  5. 5Institute of Pathology, University of Würzburg, and Comprehensive Cancer Center Mainfranken (CCCMF), Würzburg, Germany
  6. 6Berlin Institute of Health, Berlin, Germany
  7. 7Berlin-Brandenburg School for Regenerative Therapies, Berlin, Germany
  1. Correspondence to Professor Michael Hummel, Institute of Pathology, Charité—University Medicine, Berlin D-12200, Germany, michael.hummel{at}charite.de

Abstract

Objective Refractory coeliac disease (RCD) is a potentially hazardous complication of coeliac disease (CD). In contrast to RCD type I, RCD type II is a precursor entity of enteropathy-associated T-cell lymphoma (EATL), which is associated with clonally expanding T-cells that are also found in the sequentially developing EATL. Using high-throughput sequencing (HTS), we aimed to establish the small-intestinal T-cell repertoire (TCR) in CD and RCD to unravel the role of distinct T-cell clonotypes in RCD pathogenesis.

Design DNA extracted from duodenal mucosa specimens of controls (n=9), active coeliacs (n=10), coeliacs on a gluten-free diet (n=9), RCD type I (n=8), RCD type II (n=8) and unclassified Marsh I cases (n=3) collected from 2002 to 2013 was examined by TCRβ-complementarity-determining regions 3 (CDR3) multiplex PCR followed by HTS of the amplicons.

Results On average, 106 sequence reads per sample were generated consisting of up to 900 individual TCRβ rearrangements. In RCD type II, the most frequent clonotypes (ie, sequence reads with identical CDR3) represent in average 42.6% of all TCRβ rearrangements, which was significantly higher than in controls (6.8%; p<0.01) or RCD type I (6.7%; p<0.01). Repeat endoscopies in individual patients revealed stability of clonotypes for up to several years without clinical symptoms of EATL. Dominant clonotypes identified in individual patients with RCD type II were unique and not related between patients. CD-associated, gliadin-dependent CDR3 motifs were only detectable at low frequencies.

Conclusions TCRβ-HTS analysis unravels the TCR in CD and allows detailed analysis of individual TCRβ rearrangements. Dominant TCRβ sequences identified in patients with RCD type II are unique and not homologous to known gliadin-specific TCR sequences, supporting the assumption that these clonal T-cells expand independent of gluten stimulation.

  • T-CELL RECEPTOR
  • SMALL INTESTINE
  • COELIAC DISEASE
  • LYMPHOMA
  • MOLECULAR PATHOLOGY

This is an Open Access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/

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Footnotes

  • Contributors Study concept and design: JR, BS, AR, SD, MH, MS; acquisition of data: JR, SM, AS, SH, MH; analysis and interpretation of data: JR, KZ, KJ, SD, MH, MS; manuscript preparation: JR, BS, AR, SD, MH, MS, statistical analysis: JR, KZ.

  • Funding Moniek Cranmehr, Hans-Henning Müller, Erika Berg, Claudia Heldt, Ulrike Dethlefs for active support. We received funding from the Deutsche Zöliakie Gesellschaft, Dr. Schär and the Clinical Scientist programme of the Charité. The sponsors were neither involved in study design nor the collection, analysis or interpretation of data.

  • Competing interests None declared.

  • Ethics approval EA4/016/14.

  • Provenance and peer review Not commissioned; externally peer reviewed.

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