Resolution of epithelial dysfunction in eosinophilic esophagitis is mediated by an HIF-1α-CD73-adenosine signaling axis

Abstract

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

Objectives: We sought to check the hypothesis that dysregulated CD73 facilitated epithelial wound healing and barrier dysfunction in EoE and was modulated by HIF-1α and mediated through impaired extracellular adenosine signaling.

Methods: In vitro scratch assays and 3-dimensional air-liquid interface cultures were carried out on EPC2-hTERT cells to evaluate wound healing and barrier responses in the esophageal epithelium. Pharmacological CD73 inhibition (α,β-methylene adenosine-5-diphosphate), adenosine receptor A2B (ADORA2B) activation (BAY60-6583), and HIF-1α stabilization (dimethyloxalylglycine) were also used. In vivo, the L2-IL5^OXA, an 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 3-dimensional air-liquid interface cultures. Activation of downstream ADORA2B signaling 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 characterized by increased expression of occludin following ADORA2B agonism and HIF-1α stabilization.

Conclusions: Overall, our study suggests that defective HIF-1α signaling in extended hypoxia is a key driver of CD73 downregulation in EoE, leading to impaired extracellular adenosine signaling, 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.