Quantitative proteomics of coeliac gut during 14-day gluten challenge: low-level baseline inflammation despite clinical and histological normality predicts subsequent response

Abstract

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

Pivotal Role of Inflammation in Celiac Disease

“…New observations have been obtained and published challenging gluten in vivo in GFD-CD patients by analyzing before and after gluten introduction on several different pathways such as proliferation, inflammation and differentiation at the intestinal level [25][26][27]. Dotsenko et al, demonstrated that the intestinal morphology in patients on a strict GFD was similar to that measured in control subjects [25].…”

“…Another recent paper by Stamnaes J. et al [26,27] shows by proteomics analysis that before being challenged in GFD-CD patients, epithelial inflammation is already in the intestines, with changes indicative of minor crypt hyperplasia and low-grade inflammation in the serum. After challenge with gluten, an increase in the same proteins, already altered at base line in GFD-CD responders, was found.…”

“…In conclusion, studies on the GFD-CD population have helped to give an important insight into the pathogenesis of CD, but their meaning is still controversial. Some researchers regard the alterations found in GFD-CD patients as an effect of residual amounts of gluten in the diet [26]. Although small amounts of gluten contaminant cannot be ruled out from food and the CD intestines may be sensitive even to small amounts of gliadin, new insights from compartments far away from the intestines and intestinal organoids point to the presence of constitutive alterations in CD, as discussed in the next paragraphs.…”

A Cumulative Effect of Food and Viruses to Trigger Celiac Disease (CD): A Commentary on the Recent Literature

“…Gliadin and the undigested gliadin peptide p. 31–43 have also been shown to be able to induce inflammation in CD. In particular this peptide and the peptic tryptic product of gliadin can induce NFkB and the other markers of inflammation in vitro and in vivo [ 25 , 32 , 33 ].…”

Pediatric Celiac Disease Patients Show Alterations of Dendritic Cell Shape and Actin Rearrangement

“…Constitutive alterations of CD cells, in the absence of a gliadin challenge, have been described in different systems and both in vivo and in vitro assays [ 35 , 36 , 37 , 38 ]. These alterations point to a derangement of signaling, proliferation and the cytoskeleton, leading to inflammation, innate immunity activation and cellular stress in the enterocytes and fibroblasts of CD patients.…”