Characterising one of the DQ2 candidate epitopes in coeliac disease: A-gliadin 51-70 toxicity assessed using an organ culture system

OBJECTIVE: To investigate, using an organ culture system, in -vitro toxicity of region 51-70 of A-gliadin (SQQPYLQLQPFPQPQLPYSQ), a peptide overlapping some of the sequences recently characterized as DQ2-restricted T-cell epitopen in coeliac disease. METHODS: Jejunal biopsies obtained from each of ten coeliac patients (eight treated, two untreated) and two non-coeliac patients were cultured in vitro for 18 h in the presence of A-gliadin amino acids 51-70 (200 microg/ml) , organ culture medium only, peptic-tryptic digest of gliadin (1 mg/ml) or ovalbulim (1 mg/ml), the last two acting as positive and negative controls, respectively. Morphometric analysis involved measuring the cell height of 30 enterocytes, selected at random form the middle third of different villi for each section. Mean enterocyte cell heights (ECH) were compared with values for specimens cultured in medium alone. Levels of tissue transglutaminase antibody in biopsy supernatants were assessed by enzyme-linked immunosorbent assay (ELISA). RESULTS: In eight of ten coeliac patients, A-gliadin 51-70 was significantly toxic, causing a 30% decrease in ECH when compared with medium alone. In two of ten subjects, the peptide did not show any toxic effect. In all ten cases, we found that both positive and negative controls worked as expected. The peptide was non-toxic in the non-coeliac individuals. Tissue transglutaminase antibody titre in the supernatant was not found to be related to mucosal damage. CONCLUSIONS: We showed that the peptide corresponding to amino acids 51-70 of A-gliadin is characterized by in-vitro toxicity to he jejunal coeliac mucosa, correlating with recent findings of an immunological role of similar peptides. The lack of response in two of ten subjects suggests that this epitope may not be relevant in all cases of coeliac disease.