Background: Coeliac disease (CD) is due to an inappropriate T cell mediated response to specific gluten peptides. Measured by interferon γ (IFN-γ) ELISPOT, about half of the gliadin specific T cells induced with in vivo wheat gluten exposure in HLA-DQ2+ CD are specific for an α/β-gliadin peptide (p57–73 QE65; QLQPFPQPELPYPQPQS) that includes two overlapping T cell epitopes (PFPQPELPY and PQPELPYPQ).
Aim: To define minimally substituted variants of p57–73 QE65 universally devoid of IFN-γ stimulatory capacity but capable of antagonising IFN-γ secretion from polyclonal T cells specific for p57–73 QE65.
Methods: Peripheral blood mononuclear cells collected from 75 HLA-DQ2+ CD patients after in vivo gluten challenge were used in overnight ELISPOT assays to screen 218 single or double substituted variants of p57–73 QE65 for cytokine stimulatory and antagonist activity.
Results: The region p60–71 (PFPQPELPYPQP) and especially p64–67 (PELP) was sensitive to substitution. Twelve substitutions in p64–67 stimulated no IFN-γ ELISPOT response. Among 131 partial agonists identified, 45 produced statistically significant inhibition of IFN-γ ELISPOT responses when cocultured in fivefold excess with p57–73 QE65 (n = 10). Four substituted variants of p57–73 QE65 were inactive by IFN-γ ELISPOT but consistently antagonised IFN-γ ELISPOT responses to p57–73 QE65, and also retained interleukin 10 stimulatory capacity similar to p57–73 QE65.
Conclusions: Altered peptide ligands of p57–73 QE65, identified using polyclonal T cells from multiple HLA-DQ2+ CD donors, have properties in vitro that suggest that a single substitution to certain α/β-gliadins could abolish their capacity to stimulate IFN-γ from CD4 T cells and also have anti-inflammatory or protective effects in HLA-DQ2+ CD.
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Published online first 18 November 2005
Supported by grants from Oxford University Challenge Seed Fund, Ramaciotti Foundation, and Cancer Council of Victoria. RPA is the DW Keir Fellow in Clinical Research and Lions Cancer Council Fellow, DvH is a Wellcome Clinician Scientist Fellow, and AVSH is a Wellcome Trust Principal Research Fellow. JTD holds a NHMRC postgraduate scholarship.
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