Corroborating this hypothesis, a marked proliferation triggered by gliadin was reported in the peripheral blood of treated CD patients in the absence of gluten oral load, and accounted for predominantly by memory CD4+ T cells beta-catenin inhibitor [24–26]. In addition, CD8+ T lymphocytes reactive to a gliadin peptide and restricted by the HLA class I A2 molecule can be detected by the sensitive IFN-γ-ELISPOT assay in the peripheral blood of both treated and untreated CD patients who did not undergo

an in-vivo wheat gluten challenge [22]. Although our coeliac volunteers declared strict adherence to a gluten-free diet, we cannot exclude that for some of them an accidental gluten introduction might have occurred. It can be envisaged that occasional exposure to gluten could, in some cases, produce an increased frequency of gluten-reactive T cells detectable in the blood, associated presumably with the production of anti-tTG antibodies. However, although we found slight EMA/anti-tTG-positive titres in three patients, they showed no evident

differences in their response to gluten challenge compared Ribociclib to the EMA-negative subjects. In this study we compared the peripheral responses of 13 volunteers who underwent two separate wheat consumptions, separated by 3–10 months of a strict gluten-free diet. We found that the IFN-γ responses increased significantly in peripheral blood sampled 6 days after the second challenge and, unexpectedly, cells reactive to

whole gliadin were often more frequent than those observed in the first challenge, due most probably to the increased frequency of memory T cells activated upon the first gluten exposure. However, the relatively small Montelukast Sodium size of the patient cohort did not allow us to observe a statistically significant difference in the frequency of responsive cells at day 0 between the first and second challenges. Furthermore, there was no significant correlation between the specific PBMC responses to gluten and the time elapsed between the two wheat challenges. Overall, our findings suggest that a wash-out of at least 3 months is sufficient time to raise gluten-specific cells in the blood. Further studies are required to assess the memory phenotype and life turnover of circulating T cells raised during the gluten in-vivo exposure. To our knowledge, reproducibility of the short gluten challenge in the same study cohort has been poorly investigated. Importantly, we observed consistent responsiveness to the two short wheat challenges, either in terms of positive or negative responses, in 11 of 13 (85%) the patients. Raki et al. [7] reported a reduction of DQ2-α-I tetramer-positive T cells in the only patient subjected to a repeated challenge, suggesting recruitment of specific T cells in the gut after the first activation. Anderson et al.