Crohn’s & Colitis Congress™

Sessions Speakers Exhibitors Floor Plan Buzz

P042 - GREEN TEA EXTRACT, EGCG, INHIBITS DNMT-1, ER STRESS AND PREVENTS THE DEVELOPMENT OF INTESTINAL FIBROSIS IN MURINE CHRONIC TNBS-INDUCED COLITIS (Room Poster Hall)

We reported that increased endoplasmic reticulum (ER) stress signaling in subepithelial myofibroblasts (SEMF) isolated from patients with stricturing Crohn’s disease contributes to activation of TGF-β1 and fibrosis. Increased TGF-β1 results from increased interaction between GRP78 and latent TGF-beta and their translocation to the cell membrane where latent TGF-β1 is activated by αVβ3 integrin. Inhibition of ER stress by miR-199a is also lost due to DNA methyltransferase-1 (DNMT1)-induced promoter silencing of miR-199a-5p. AIM: To characterize ER stress in SEMF of mice after 8 weeks of chronic colitis, a model of fibrostenotic Crohn’s disease, and determine whether the natural DNMT-1 inhibitor, green tea extract (EGCG), inhibits ER stress and development of fibrosis. METHODS: C57BL/6J mice were treated for 8 weeks with escalating doses of intrarectal TNBS. Mice were given EGCG (10mg/kg/ir) in 0.9% PBS or PBS alone twice a week for 8 weeks. Collagen content was determined by Sircol collagen assay. SEMF isolated and placed into primary culture were used to prepare RNA for qRT-PCR, cell lysates for Western blot analysis of ER stress proteins and DNMT1, or for monolayer wound healing assay. RESULTS: Chronic TNBS colitis in mice results in a similar ER stress response in SEMF: increased GRP78 and XBP1, and DNMT-1 as that seen in SEMF of stricturing Crohn’s disease. EGCG decreased expression of DNMT-1 by 8 fold and protein levels by 65±3% and resulted in decreased GRP78 and XBP1 protein levels indicating an inhibition of ER stress. Collagen content in EGCG treated TNBS mice was reduced to 42±2 ng/ml compared to 135±6 ng/ml in control TNBS mice. MPO assay showed no significant inflammation after 8-week of TNBS. Wound healing decreased 60±3% with siRNA GRP78 knockdown. CONCLUSION: Chronic TNBS colitis in mice is a model of ER stress in SEMF of stricturing Crohn’s disease. Inhibition of ER stress with EGCG prevents TNBS-induced intestinal fibrosis in a mouse model.