Authors
S. HU (1), N. BLÉTARD (2), C. MASSOT (3), N. PIERRE (1), F. QUESADA CALVO (1), G. PAULISSEN (1), G. MAZZUCCHELLI (4), N. SMARGIASSO (5), D. BAIWIR (6), E. DE PAUW (5), P. DELVENNE (2), M. MEUWIS (7), E. LOUIS (3) / [1] GIGA-R, Univeristy of Liège, , Liège, Belgium, Translational Gastroenterology , [2] CHU de Liège, Liège, Belgium, Pathological anatomy and cytology, [3] CHU de Liège -GIGA-R, Liège, Belgium, Hepato-Gastroenterology and Digestive Oncology and Translational Gastroenterology, [4] University of Liège, , Belgium, Laboratory of Mass Spectrometry, Chemsitry dep. , [5] University of Liège, , Belgium, Laboratory of Mass Spectrometry, Chemistry dep., [6] GIGA facility Uliège, Liège, Belgium, GIGA-proteomics, [7] CHU de Liège -GIGA-R, Liège, Belgium, Hepato-Gastroenterology and Digestive Oncology Department and Translational Gastroenterology
Introduction
Intestinal fibrosis in Crohn’s Disease (CD) is complex and its initiation and progression are linked to chronic inflammation and lead to bowel damages. Bordering epithelium involvement in this process remains unravelled.
Aim
Our purpose was to address proteomic changes occurring in the bordering epithelium of regions with increasing degrees of sub-mucosal inflammation and fibrosis. Proteins differentially abundant could be potential actors eliciting fibrogenic pathways and potential therapeutic targets.
Methods
Formalin fixed paraffin embedded tissue sections from CD patients with ileal stenosis (n=5) were treated by laser capture microdissection to isolate bordering epithelial cells. Paired regions were selected in Normal (NL), inflammatory and mildly fibrotic (IF1) as well as inflammatory and moderately to severely fibrotic (IF2/3) areas. Protein digests were analysed by label free proteomics. Immunohistochemical (IHC) evaluation on independent CD cases was done (n=32). Moreover, the epithelial colonic cell line HT29 was used for in vitro culture experiments in order to characterise the selected protein in the epithelium and its potential impact on a simple model of intestinal fibrosis.
Results
Proteomics identified 1249 proteins from which 257 showed a distribution varying significantly with the degree of fibrosis. Anteriority gradient protein homolog 2 (AGR2), related to endoplasmic reticulum (ER) stress and mucus secretion, was the most significant one. AGR2 IHC confirmed a significant increase in ileal CD (n=19) with inflammation and a stronger one with fibrosis. In the colon (n=13) there was no significant difference between simple inflammation and inflammation with fibrosis. ER stress induction in HT29 cells enabled AGR2 increase as measured by RT-Q-PCR and WB, while TGFβ, the main fibrogenic elicitor, induced a decrease of this protein.
Conclusions
AGR2 increase in the epithelium of fibrotic CD ileum and in epithelial cell line upon ER stress induction suggest a potential role for epithelium and ER stress (including AGR2) in the development of surrounding submucosal fibrosis in ileal CD.
