Resolution of Epithelial Dysfunction in Eosinophilic Esophagitis is Mediated by a HIF-1α-CD73-Adenosine Signalling Axis

Abstract

Background: Inflammatory hypoxia is induced by eosinophilic inflammation, while a decreased expression in the key hypoxic regulator, hypoxia inducible transcription factor (HIF-1α) has been shown in Eosinophilic Esophagitis (EoE) patients, contributing to maladaptive hypoxic responses in the esophageal epithelium. Recent publications have reported attenuated CD73/ecto-5'-nucleotidase expression in EoE patients, which is a known extracellular adenosine producing ecto-enzyme and a direct HIF-1α target.

Objectives: We hypothesised that dysregulated CD73 facilitated epithelial wound healing and barrier dysfunction in EoE and was modulated by HIF-1α and mediated through impaired extracellular adenosine signalling.

Methods: In vitro scratch assays and 3-dimensional air liquid interface (3D-ALI) cultures were carried on EPC2-hTERT cells out to evaluate wound healing and barrier responses in the esophageal epithelium. Pharmacological CD73 inhibition (α,β-methylene adenosine-5-diphosphate (APCP)), ADORA2B adenosine receptor activation (BAY60-6583) and HIF-1α stabilization (DMOG) were also used. In vivo, the L2-IL5^OXA, a HIF-1α overexpressing L2-IL5^OXA (L2-IL5⁺/HIF-1A-dPA^+/+/K14Cre⁺)^OXA and ADORA2B agonist-BAY60-6583 (L2-IL5^OXABAY) EoE mouse models were used.

Results: Pharmacologic studies demonstrated that CD73 inhibition resulted in defective wound healing responses in scratch assays and decreased epithelial barrier in 3D-ALI cultures. Activation of downstream adenosine A2B receptor (ADORA2B) signalling improved wound healing responses and barrier functions via restoration of fibronectin (FN1) and occludin (OCLN) expression. In vivo studies in L2-IL5^OXA EoE mouse models recapitulated in vitro findings of improved epithelial barrier integrity characterised by increased expression of occludin following ADORA2B agonism and HIF-1α stabilisation.

Conclusions: Overall, our study suggests defective HIF-1α signalling in extended hypoxia is a key driver of CD73 downregulation in EoE, leading to impaired extracellular adenosine signalling, defective wound healing and barrier dysfunction, establishing the possibility for ADORA2B agonism as a potential novel therapeutic approach for EoE.

Keywords: CD73; Eosinophilic esophagitis; HIF-1α; barrier integrity; extracellular adenosine; wound healing.