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Coeliac disease: immunogenicity studies of barley hordein and rye secalin-derived peptides.

International journal of experimental pathology (2016-09-24)
Widya A Wahab, Tanja Šuligoj, Julia Ellis, Beatriz Côrtez-Real, Paul J Ciclitira
ABSTRACT

Coeliac disease (CD) is an inflammatory disorder of the small intestine. It includes aberrant adaptive immunity with presentation of CD toxic gluten peptides by HLA-DQ2 or DQ8 molecules to gluten-sensitive T cells. A ω-gliadin/C-hordein peptide (QPFPQPEQPFPW) and a rye-derived secalin peptide (QPFPQPQQPIPQ) were proposed to be toxic in CD, as they yielded positive responses when assessed with peripheral blood T-cell clones derived from individuals with CD. We sought to assess the immunogenicity of the candidate peptides using gluten-sensitive T-cell lines obtained from CD small intestinal biopsies. We also sought to investigate the potential cross-reactivity of wheat gluten-sensitive T-cell lines with peptic-tryptic digested barley hordein (PTH) and rye secalin (PTS). Synthesised candidate peptides were deamidated with tissue transglutaminase (tTG). Gluten-sensitive T-cell lines were generated by culturing small intestinal biopsies from CD patients with peptic-tryptic gluten (PTG), PTH or PTS, along with autologous PBMCs for antigen presentation. The stimulation indices were determined by measuring the relative cellular proliferation via incorporation of

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Sigma-Aldrich
HEPES, 1 M, pH 7.0-7.6, sterile-filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Transglutaminase from guinea pig liver, lyophilized powder, ≥1.5 units/mg protein
Sigma-Aldrich
Pepsin−Agarose from porcine gastric mucosa, lyophilized powder, ≥30 units/mg dry solid
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Trypsin Agarose, buffered aqueous suspension, from bovine pancreas trypsin