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

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 ³ H-thymidine. The majority of T-cell lines reacted to the peptides studied. There was also cross-reactivity between wheat gluten-sensitive T-cell lines and the hordein, gliadin and secalin peptides. PTH, PTS, barley hordein and rye secalin-derived CD antigen-sensitive T-cell lines showed positive stimulation with PTG. ω-gliadin/C-hordein peptide and rye-derived peptide are immunogenic to gluten-sensitive T-cell lines and potentially present in wheat, rye and barley. Additional CD toxic peptides may be shared.

Keywords: antigen presentation; coeliac disease; gluten; small intestinal T-cell lines.

Figure 1

Each dot represents a stimulation index (S.I) for an individual assay. The lines represent the medians of the value for each tested antigen; ns indicates not significant (compared with PTG). PTG is peptic–tryptic gluten; PepG is ω‐gliadin/C‐hordein; peptide PepR is rye‐derived peptide (QPFPQPQQPIPQ); PTH is peptic–tryptic barley hordein; and PTR is peptic–tryptic rye secalin.

Figure 2

Stimulation indices (S.I) of small intestinal wheat gluten‐sensitive T‐cell lines (n = 14) tested in proliferation assays with three Triticeae prolamins (PTG, PTH, PTR) and two potential CD toxic peptides (PepG and PepR). Each bar corresponds to S.I obtained per antigen tested with 14 TCLs, labelled as WG1‐14 (patients studied with wheat gluten). A line at S.I of 2 across the bars distinguishes positive from negative results of proliferation assays. PTG is peptic–tryptic gluten; PepG is ω‐gliadin/C‐hordein; peptide PepR is rye‐derived peptide (QPFPQPQQPIPQ); PTH is peptic–tryptic barley hordein; and PTR is peptic–tryptic rye secalin.

Figure 3

Each dot represents a stimulation index (S.I) for an individual assay. The lines represent the medians; ns indicates not significant (compared with PTG). PTG is peptic–tryptic gluten; PepG is ω‐gliadin/C‐hordein; peptide PepR is rye‐derived peptide (QPFPQPQQPIPQ); and PTH is peptic–tryptic barley hordein.

Figure 4

Stimulation indices (S.I) of barley hordein‐sensitive T‐cell lines (n = 9) tested in proliferation assays with three Triticeae prolamins (PTG, PTH, PTR) and two potential CD toxic peptides (PepG and PepR). Each bar corresponds to the S.I obtained per antigen tested with nine TCLs, labelled as WB1‐9 (patients studied with barley hordein). A dotted line at S.I of 2 distinguishes positive from negative results of proliferation assays. PTG is peptic–tryptic gluten; PepG is ω‐gliadin/C‐hordein; peptide PepR is rye‐derived peptide (QPFPQPQQPIPQ); PTH is peptic–tryptic barley hordein; and PTR is peptic–tryptic rye secalin

Figure 5

Each dot represents a stimulation index (S.I) for an individual assay. PTG is peptic–tryptic gluten; PepG is ω‐gliadin/C‐hordein; peptide PepR is rye‐derived peptide (QPFPQPQQPIPQ); and PTR is peptic–tryptic rye secalin.