T-cell repertoires in refractory coeliac disease

Abstract

ObjectiveRefractory 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.DesignDNA 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.ResultsOn 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.ConclusionsTCRβ-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.

Clonal T cell receptor gene rearrangements in coeliac disease: implications for diagnosing refractory coeliac disease

“…A recent study utilising high-throughput sequencing of TCR-β genes to characterise the small intestinal T cell repertoire in CD and RCD detected frequent dominant clonotypes in RCDII cases along with lower numbers of other clonotypes (reduced repertoire) 25. Dominant TCR sequences/clonotypes were also noted in the biopsies of patients without CD and with RCDI, while patients with ACD had a significantly higher number of clonotypes (expanded repertoire) compared with CD patients on GFD, which might reflect bystander activation and expansion of T cells as a consequence of inflammation.…”

NKp46 is a diagnostic biomarker and may be a therapeutic target in gastrointestinal T-cell lymphoproliferative diseases: a CELAC study

“…Due to their normal cytology and intracellular expression of CD3, RCDII IEL can be easily mistaken for normal T-IEL. As a consequence, current diagnosis of RCDII is very challenging in the absence of flow cytometry and TCR clonality analyses 25. Herein, we show that a threshold of 25 NKp46+ IEL/100 EC can differentiate RCDII from uncomplicated CD and from non-malignant cases of CD refractory to GFD (RCDI) with high sensitivity and specificity.…”

Antigenic mimicry of ubiquitin by the gut bacteriumBacteroides fragilis: a potential link with autoimmune disease

“…A loss of epithelial integrity may precede gluten sensitivity in individuals with a genetic predisposition to coeliac disease . Interestingly, there is an increased risk of coeliac disease development with multiple gastrointestinal infections . The high incidence of anti‐BfUbb and anti‐Hubb antibodies in the coeliac serum category could therefore reflect early exposure to BfUbb and could be an additional aetiological factor for coeliac disease development.…”