In celiac disease (CeD), gluten activates adaptive immune cells that cause damage to the small intestinal mucosa. Histological evaluation of intestinal biopsies allows for grading of disease severity. CeD can effectively be treated with a life‐long gluten‐free diet. Gluten challenge of treated CeD patients is used to confirm diagnosis and to test drug efficacy in clinical trials, but patients respond with different magnitudes to the same gluten challenge. In this study of 19 well‐treated CeD patients, proteome analysis of total tissue or isolated epithelial cell compartment from formalin‐fixed paraffin embedded biopsies collected before and after 14‐day gluten challenge demonstrates that patients with strong mucosal response to challenge have signs of ongoing tissue inflammation already before challenge. This low‐level tissue inflammation at baseline is paralleled by increased gluten specific CD4+ T‐cell frequencies in the gut and presence of a low‐level blood inflammatory profile. Thus, apparently well‐treated CeD is frequently not entirely quiescent, with presence of low‐grade inflammation and antigluten immunity in the gut mucosa. Histology assessment alone appears insufficient to judge full recovery and gut mucosal healing of CeD patients. The findings raise a concern whether a seemingly proper gluten‐free diet is able to curb gut inflammation in all CeD patients.

Objective To shed light on gut mucosa processes provoked by gluten exposure in coeliac disease, we performed a quantitative proteomic analysis of duodenal tissue from well-treated coeliac patients before and after gluten challenge. Design We extracted and digested proteins from formalin-fixed paraffin-embedded tissue of 19 coeliac disease patients who had been challenged orally with gluten for 14 days. Protein identification and quantification was done by label-free quantitative mass spectrometry-based proteomics from total tissue and from laser capture microdissected epithelial cell layer. Proteomics data were compared with clinical, serological and histological data. Results At baseline, all patients were in clinical and mucosal remission (Marsh 0-1) except one (Marsh 3). After challenge, five patients reached Marsh 3 scores. Proteome analysis categorised these five and additionally two patients as responders. Already at baseline, responder patients differed from the remaining patients in their gut tissue protein composition with altered levels of inflammatory and enterocyte function proteins - the same proteins that changed upon gluten challenge. Patients classified as responders from the proteomic analysis also differed from the remaining patients at baseline, with mild crypt hyperplasia and a slight increase in blood inflammatory parameters and gluten specific CD4+ T-cell frequencies. Conclusion Despite clinical and histological remission, coeliac disease patients that develop a mucosal response after 14-day gluten challenge have already at baseline altered protein compositions of their gut tissue with signs of ongoing inflammation. Thus, apparently well-treated coeliac disease is frequently not fully quiescent with presence of low-grade anti-gluten immunity in gut mucosa