IMMUNOGLOBULIN A-COATED BACTERIA REPRESENT A UNIQUE SUBSET OF THE MICROBIOME IN PATIENTS WITH NEWLY DIAGNOSED INFLAMMATORY BOWEL DISEASE

BACKGROUND: B-cell lymphoma (BCL)2 family members induce or abolish apoptosis.Activation and differentiation of fibroblasts is linked to the expression of the BCL2 family.As fibroblasts are activated during fibrosis, decreasing BCL2 might represent a potential treatment approach.Fibrosis as a common problem in patients with Crohn's disease (CD) is resulting from an imbalance towards excessive fibrous tissue formation driven by fibroblasts.We investigated the impact of BCL2 repression on fibrogenesis.METHODS: Fibroblasts were stimulated with the BCL2 antagonist ABT-737 (AbbVie, USA, final concentration 0.01 -100 nM).BCL2, BCLXL, a-SMA and COL1A1 were determined by qPCR, IF and WB.The impact of BCL2 antagonist treatment on TGF b signaling pathways in primary human colonic fibroblasts was investigated using WB and IF. mRNA expression pattern in primary human colonic fibroblasts was determined by Next Generation Sequencing (NGS).For in vivo experiments, both the murine heterotopic transplantation model of intestinal fibrosis and the model of dextran sodium sulfate (DSS)-induced chronic colitis were used.Animals received BCL2 antagonist (50 mg/kg/day, intraperitoneally (i.p.)).Collagen layer thickness and hydroxyproline (HYP) content were determined.RESULTS: BCL2 and BCLXL were significantly decreased in primary human colonic fibroblasts upon administration of 1nM and 10nM BCL2 antagonist treatment compared to vehicle in a dose-dependent manner (0.68 ± 0.12 and 0.49 ± 0.04 vs. 1.00 ± 0.00, respectively, * p < 0.05 for BCL2, 0.41±0.26and 0.35±0.22 vs. 1.00 ± 0.00, respectively, * p < 0.05 for BCLXL).COL1A1 and a-SMA were decreased in both human and murine colonic fibroblasts upon BCL2 antagonist treatment compared to vehicle.WB analysis revealed a decrease of pERK1, pERK2 and SMAD3 from the TGFb signaling pathways in primary human fibroblasts in a dose dependent manner upon BCL2 antagonist treatment compared to vehicle.NGS and qPCR revealed a link to the transcription factors GATA6 and SOX9 for mediating pro-fibrotic effects and for reprogramming fibroblasts.Collagen layer thickness was significantly decreased in the mouse model of fibrosis upon BCL2 antagonist administration compared to vehicle (* p < 0.05).Decreased HYP content upon BCL2 antagonist administration confirmed the preventive effects of the BCL2 antagonist on intestinal fibrosis in vivo.CONCLUSION: The BCL2 antagonist changed the expression profile of BCL2 family members and prevented fibroblast differentiation into myofibroblasts.BCL2 antagonist administration partially prevented intestinal fibrogenesis in both the murine heterotopic transplantation model of intestinal fibrosis and the DSS-induced chronic colitis model and therefore may represent a potential treatment option against CD associated fibrosis.