primary prevention of coeliac disease - Associazione Italiana ...

A DOMINANT EPITOPE IN GLUTEN PEPTIDES ?
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exploited to search for “toxic” gluten peptides . A-gliadin, the first fully sequenced
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wheat gliadin protein , has been used as the archetypal “toxic gluten” protein source
for most of these peptides (see Fig. 1). More recently, recombinant gliadins and
peptides corresponding to parts of cDNA-derived gliadin sequences have been studied
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. In view of the diversity of model systems, some with no relationship to coeliac
disease (for example fetal intestinal explants), it is not surprising that a variety of
different peptides have been defined as toxic. In other studies, non-specific markers of
toxicity (for example epithelial cell height) have been used as surrogates for
immunological toxicity. If the primary hypothesis is that coeliac disease is a T cell
mediated disease, rational identification of “toxic” gluten peptides in coeliac disease
should ideally begin with assays designed to detect HLA-DQ2 restricted CD4 T cells
that secrete Th1-associated cytokines (for example interferon gamma).
Intestinal T cell clones for identification of toxic gluten peptides
More recently, intestinal T cell clones from duodenal biopsies of coeliac patients on
gluten free diet pulsed with protease-digested gliadin have been used to search for
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gliadin-specific T cell epitopes . These studies have revealed that intestinal T cell
clones raised from gliadin-pulsed coeliac intestinal biopsies predominantly recognise
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deamidated gliadin epitopes . T cell epitopes generally correspond to gliadin peptides
deamidated by tissue transglutaminase (tTG). tTG is induced with inflammation and
apoptosis, and irreversibly cross links proteins and peptides via glu-lys isopeptide
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bonds, or directly deamidates glutamine residues to glutamate . Intestinal tissue,
particularly if inflamed, is rich in tTG. Introduction of glutamate in gliadin T cell
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epitopes by tTG greatly enhances their binding to HLA-DQ2 .
T cell epitope mapping in immune-mediated diseases
In human immune-mediated diseases, T cell epitopes have been mapped using
synthetic 15-20mers overlapping by 10 residues spanning known antigenic proteins in
assays of peripheral blood T cells. In coeliac disease, it has been contended that gliadin-
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specific peripheral blood T cells are qualitatively different from intestinal T cells .
Gliadin-specific intestinal T cell clones are generally HLA-DQ2 restricted while
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peripheral blood T cell clones are HLA-DR, -DQ or -DP restricted . However, these
studies were performed before the realization that deamidation of gliadin was
important for T cell recognition. Hence, contemporary understanding of toxic gluten
peptides in the context of coeliac disease as an HLA-DQ2 associated CD4 T cellmediated
disease has relied upon identification of epitopes of intestinal T cell clones
using protease-digested gliadin with or without deamidation by tTG.
Unfortunately, it is impossible to know whether epitopes of T cell clones are
dominant or subdominant, or whether a clone is specific for only a part of a polyclonal T
cell response in vivo. Furthermore, recent studies to identify epitopes of gliadin T cell
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clones have used chymotrypsin-digested gliadins even though chymotrysin
selectively cleaves peptide bonds following bulky hydrophobic aminoacids (the same
aminoacids known to be anchor residues at the N - and C-terminal end of the HLA-DQ2
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binding motif ), raising the possibility that bioactive peptides may be artefactually