Review of the Study “Esophageal ILC2s Mediate Abnormal Epithelial Remodeling in Eosinophilic Esophagitis via AREG-EGFR Signaling”

Background Overview

Eosinophilic Esophagitis (EoE) is a chronic allergic disease that has shown a significant increase in prevalence in recent years. It is characterized by eosinophil infiltration and epithelial thickening, severely impairing swallowing function. The pathological features of EoE include basal cell hyperproliferation, epithelial thickening, and fibrosis, which significantly alter the morphology and function of the esophagus. Historically, EoE pathogenesis was attributed primarily to the adaptive immune system, particularly Th2-mediated immune responses. However, recent research on Group 2 Innate Lymphoid Cells (ILC2s) has revealed their critical role in type 2 inflammation. ILC2s are activated by alarmins such as IL-25 and IL-33 and produce cytokines like IL-4, IL-5, IL-13, and Amphiregulin (AREG). These molecules regulate epithelial cell proliferation and differentiation, playing a vital role in maintaining mucosal barrier integrity.

While the role of ILC2s in respiratory pathophysiology, such as allergic asthma, is relatively well studied, their specific role in EoE pathogenesis remains unclear. ILC2s could exacerbate disease progression by promoting epithelial remodeling through the secretion of AREG, which binds to the EGFR (Epidermal Growth Factor Receptor). Thus, this study aims to elucidate whether ILC2s drive abnormal epithelial remodeling in EoE through the AREG-EGFR signaling pathway, offering new therapeutic targets for the treatment of EoE.

Study Context

This study was conducted by Minyeong Lim and colleagues from several renowned Korean research institutions, including Seoul National University College of Medicine, Ewha Womans University, Jeju National University College of Medicine, and Hanyang University College of Medicine. The article was published in the journal Cellular & Molecular Immunology in 2025, with the initial manuscript submitted in May 2024 and the online publication completed in December 2024. The corresponding author is Hye Young Kim.

Research Workflow and Methods

a. Workflow and Experimental Design

The study utilized various experimental approaches, including animal models, in vitro experiments, and human sample analyses. The key aspects are described below:

1. Characterization of ILC2 Distribution and Features:

   • Flow cytometry was used to analyze ILC2 populations in healthy mouse esophageal and lung tissues. Results showed that ILC2s were significantly more frequent in the esophagus than in the lungs.
   • Immunofluorescence staining of human esophageal tissues from healthy individuals and EoE patients revealed a marked increase in CD3e-KLRG1+ ILC2s in the esophagus of EoE patients. In contrast, ILC2s were scarce in healthy individuals and those with Gastroesophageal Reflux Disease (GERD).

2. Validation of IL-33-Induced EoE Mouse Model:

   • Acute and chronic EoE models were established in mice through intranasal administration of recombinant IL-33 (rmIL-33), which induced hallmark EoE features such as eosinophil infiltration, basal cell hyperplasia, and epithelial thickening.
   • In Rag1 knockout and Rag2/IL2rg double knockout mice, the absence of ILCs alleviated pathological epithelial changes induced by IL-33.
   • Red fluorescent protein (tdTomato)-labeled IL-5 reporters were employed to study ILC2 responses to IL-33 stimulation. Activation of ILC2s was observed, with clustering near the epithelial barrier.

3. Role of the AREG-EGFR Signaling Axis:

   • AREG expression was confirmed to be primarily derived from KLRG1+ ILC2s in the IL-33 model. Immunohistochemistry revealed that AREG binds to EGFR, activating phosphorylation signaling pathways that promote basal cell proliferation.
   • In vitro experiments using human esophageal epithelial cell lines (CP-A and HET-1A) demonstrated that AREG significantly induced EGFR phosphorylation and downstream activation of AKT and ERK signaling pathways, leading to cell proliferation. The EGFR inhibitor Erlotinib completely blocked these effects.
   • Injection of recombinant AREG into normal mice replicated EoE-like symptoms, further confirming the critical role of AREG in pathological epithelial remodeling.

4. Therapeutic Potential Evaluation:

   • Intervention with Erlotinib or neutralizing anti-AREG antibodies reduced epithelial thickening and eosinophilic infiltration in the EoE mouse model.
   • In 3D organoid cultures co-cultured with primary ILC2s, basal cell proliferation was promoted by ILC2-derived AREG and was completely suppressed upon the addition of anti-AREG or EGFR inhibitors.

5. Validation with Human Samples:

   • Single-cell RNA sequencing data from EoE patients revealed that ILC2s were the major source of AREG. Immunofluorescence staining of patient tissues highlighted elevated levels of activated ILC2s (KLRG1+), AREG, and phosphorylated EGFR in EoE samples, providing translational relevance to human disease.

b. Key Findings

• Esophageal ILC2s were significantly increased and localized near the epithelial border in EoE pathology.
• Experiments demonstrated that ILC2s secrete AREG, which binds to EGFR to activate downstream signaling pathways, causing basal cell proliferation and epithelial thickening.
• Therapeutic interventions targeting the AREG-EGFR axis successfully reduced epithelial remodeling and inflammation in the murine EoE model.
• Human sample analyses confirmed the key role of ILC2s and AREG-EGFR signaling in esophageal pathology, consistent with findings in mice.

c. Conclusions and Implications

The study established that ILC2s play a critical role in driving abnormal epithelial remodeling in EoE through the AREG-EGFR signaling axis. This finding offers valuable insights into EoE pathogenesis and proposes novel therapeutic targets beyond traditional type 2 inflammation pathways. Therapies that inhibit AREG or EGFR signaling could effectively mitigate epithelial remodeling, providing an innovative strategy for managing EoE and potentially other related allergic diseases.

d. Highlights and Novelty

• Highlighted the role of ILC2s in esophageal immunity and their central contribution to EoE pathology.
• Provided mechanistic insights into how ILC2s mediate basal cell hyperplasia through the AREG-EGFR signaling pathway.
• Proposed and validated the therapeutic potential of targeting ILC2s, AREG, or EGFR to treat EoE-related epithelial abnormalities.

e. Additional Insights

The research comprehensively verified the critical role of the ILC2-AREG-EGFR axis in multiple organisms, including mouse models and human patients. Cross-species validation suggests that targeting this signaling pathway may have broader implications for other inflammation-driven epithelial diseases.

Summary

Through extensive experimentation and multi-faceted analyses, this study elucidated the previously unrecognized role of ILC2s in mediating abnormal epithelial remodeling in EoE through the AREG-EGFR axis. Targeting this pathway provides a promising therapeutic avenue, potentially revolutionizing current EoE management strategies. Additionally, this discovery broadens the understanding of esophageal immunology and epithelial interactions in disease, laying the groundwork for further clinical translation and therapeutic development.