TGF-ß1 alters esophageal epithelial barrier function by attenuation of claudin-7 in eosinophilic esophagitis.

Barrier dysfunction has been implicated in the pathophysiology of eosinophilic esophagitis (EoE). Transforming growth factor-ß1 (TGF-ß1), a potent pleiotropic molecule, is increased in EoE; however, no study has evaluated its influence on esophageal epithelial barrier. We hypothesized that TGF-ß1 regulates barrier dysfunction in EoE. We aimed to determine the role of TGF-ß1 in the epithelial barrier in models of EoE. To examine the impact of TGF-ß1 on esophageal barrier, immortalized human esophageal epithelial (EPC2-hTERT) cells were exposed to TGF-ß1 during the three-dimensional air-liquid interface (3D-ALI) model in vitro. TGF-ß1 exposure diminished EPC2-hTERT barrier function as measured by transepithelial electrical resistance (TEER) and 3 kDa Fluorescein isothiocyanate dextran paracellular flux (FITC Flux), and hematoxylin and eosin (H&E) assessment revealed prominent cellular separation. In analysis of epithelial barrier molecules, TGF-ß1 led to the specific reduction in expression of the tight-junction molecule, claudin-7 (CLDN7), and this was prevented by TGF-ß-receptor I inhibitor. Short hairpin ribonucleic acid (shRNA)-mediated CLDN7 knockdown diminished epithelial barrier function, whereas CLDN7 overexpression resulted in protection from TGF-ß1-mediated barrier dysfunction. In pediatric EoE biopsies CLDN7 expression was decreased and altered localization was observed with immunofluorescence analysis, and the TGF-ß1 downstream transcription factor, phosphorylated SMAD2/3 (pSMAD2/3), was increased. Our data suggest that TGF-ß1 participates in esophageal epithelial barrier dysfunction through CLDN7 dysregulation.

Tryptophan metabolite activation of the aryl hydrocarbon receptor regulates IL-10 receptor expression on intestinal epithelia.

IL-10 is a potent anti-inflammatory cytokine that inhibits the production of proinflammatory mediators. Signaling by IL-10 occurs through the IL-10 receptor (IL-10R), which is expressed in numerous cell types, including intestinal epithelial cells (IECs), where it is associated with development and maintenance of barrier function. Guided by an unbiased metabolomics screen, we identified tryptophan (Trp) metabolism as a major modifying pathway in interferon-γ (IFNγ)-dominant murine colitis. In parallel, we demonstrated that IFNγ induction of indoleamine 2,3-dioxygenase 1, an enzyme that catalyzes the conversion of Trp to kynurenine (Kyn), induces IL-10R1 expression. Based on these findings, we hypothesized that IL-10R1 expression on IEC is regulated by Trp metabolites. Analysis of the promoter region of IL-10R1 revealed a functional aryl hydrocarbon response element, which is induced by Kyn in luciferase-based IL-10R1 promoter assays. Additionally, this analysis confirmed that IL-10R1 protein levels were increased in response to Kyn in IEC in vitro. Studies using in vitro wounding assays revealed that Kyn accelerates IL-10-dependent wound closure. Finally, reduction of murine dextran sodium sulfate colitis through Kyn administration correlates with colonic IL-10R1 expression. Taken together, these results provide evidence on the importance of IL-10 signaling in intestinal epithelia and implicate AHR in the regulation of IL-10R1 expression in the colon.