Epithelial HIF-1α/claudin-1 axis regulates barrier dysfunction in eosinophilic esophagitis
Joanne C. Masterson, … , Sean P. Colgan, Glenn T. Furuta
Joanne C. Masterson, … , Sean P. Colgan, Glenn T. Furuta
Published August 1, 2019; First published July 2, 2019
Citation Information: J Clin Invest. 2019;129(8):3224-3235. https://doi.org/10.1172/JCI126744.
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Research Article Gastroenterology

Epithelial HIF-1α/claudin-1 axis regulates barrier dysfunction in eosinophilic esophagitis

Abstract

Epithelial barrier dysfunction is a significant factor in many allergic diseases, including eosinophilic esophagitis (EoE). Infiltrating leukocytes and tissue adaptations increase metabolic demands and decrease oxygen availability at barrier surfaces. Understanding of how these processes impact barrier is limited, particularly in allergy. Here, we identified a regulatory axis whereby the oxygen-sensing transcription factor HIF-1α orchestrated epithelial barrier integrity, selectively controlling tight junction CLDN1 (claudin-1). Prolonged experimental hypoxia or HIF1A knockdown suppressed HIF-1α–dependent claudin-1 expression and epithelial barrier function, as documented in 3D organotypic epithelial cultures. L2-IL5OXA mice with EoE-relevant allergic inflammation displayed localized eosinophil oxygen metabolism, tissue hypoxia, and impaired claudin-1 barrier via repression of HIF-1α/claudin-1 signaling, which was restored by transgenic expression of esophageal epithelial-targeted stabilized HIF-1α. EoE patient biopsy analysis identified a repressed HIF-1α/claudin-1 axis, which was restored via pharmacologic HIF-1α stabilization ex vivo. Collectively, these studies reveal HIF-1α’s critical role in maintaining barrier and highlight the HIF-1α/claudin-1 axis as a potential therapeutic target for EoE.

Authors

Joanne C. Masterson, Kathryn A. Biette, Juliet A. Hammer, Nathalie Nguyen, Kelley E. Capocelli, Bejan J. Saeedi, Rachel F. Harris, Shahan D. Fernando, Lindsay B. Hosford, Caleb J. Kelly, Eric L. Campbell, Stefan F. Ehrentraut, Faria N. Ahmed, Hiroshi Nakagawa, James J. Lee, Eóin N. McNamee, Louise E. Glover, Sean P. Colgan, Glenn T. Furuta

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Figure 4

CLDN1 is a direct HIF-1α target gene, and claudin-1 protein is mislocalized in active EoE patients.

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CLDN1 is a direct HIF-1α target gene, and claudin-1 protein is mislocali...
(A) Proximal human CLDN1 promoter sequence identified 2 potential hypoxia-responsive elements (HREs). Transcription start site, +1. ChIP using a HIF-1α antibody on cells exposed to 4 hours of hypoxia compared with normoxia, followed by PCR using primers spanning HRE2 and HRE1 sites, was normalized to PCRs performed on ChIP product generated using an isotype IgG control antibody (n = 3). Luciferase activity in lysates transfected with full-length CLDN1 promoter (WT) compared with cells transfected with either mutated HRE site 1 or mutated HRE site 2. Luciferase data normalized to total cell protein content (n = 6). Statistical significance was assessed using an ordinary 1-way ANOVA with correction for multiple comparisons. *P < 0.05, **P < 0.01. (B) Esophageal pinch biopsies from patients undergoing endoscopy were obtained and assessed for CLDN1 expression by real-time RT-PCR (n = 6–10 per group). Statistical significance was assessed using the nonparametric Kruskal-Wallis test with correction for multiple comparisons. *P < 0.05, **P < 0.01. (C) Representative photomicrographs of claudin-1 immunolocalization. Scale bars: 100 μm. (D) Arrows depict dilated intercellular spaces in H&E-stained FFPE pinch biopsies from active EoE subjects compared with normal controls. Data are expressed as means ± SEM and represent a minimum of 3 experimental repeats.