Single-cell RNA sequencing of mast cells in eosinophilic esophagitis reveals heterogeneity, local proliferation, and activation that persists in remission.

BACKGROUND: Mast cells (MCs) are pleiotropic cells that accumulate in the esophagus of patients with eosinophilic esophagitis (EoE) and are thought to contribute to disease pathogenesis, yet their properties and functions in this organ are largely unknown. OBJECTIVES: This study aimed to perform a comprehensive molecular and spatial characterization of esophageal MCs in EoE. METHODS: Esophageal biopsies obtained from patients with active EoE, patients with EoE in histologic remission, and individuals with histologically normal esophageal biopsies and no history of esophageal disease (ie, control individuals) were subject to single-cell RNA sequencing, flow cytometry, and immunofluorescence analyses. RESULTS: This study probed 39,562 single esophageal cells by single-cell RNA sequencing; approximately 5% of these cells were MCs. Dynamic MC expansion was identified across disease states. During homeostasis, TPSAB1highAREGhigh resident MCs were mainly detected in the lamina propria and exhibited a quiescent phenotype. In patients with active EoE, resident MCs assumed an activated phenotype, and 2 additional proinflammatory MC populations emerged in the intraepithelial compartment, each linked to a proliferating MKI67high cluster. One proinflammatory activated MC population, marked as KIThighIL1RL1highFCER1Alow, was not detected in disease remission (termed "transient MC"), whereas the other population, marked as CMA1highCTSGhigh, was detected in disease remission where it maintained an activated state (termed "persistent MC"). MCs were prominent producers of esophageal IL-13 mRNA and protein, a key therapeutic target in EoE. CONCLUSIONS: Esophageal MCs comprise heterogeneous populations with transcriptional signatures associated with distinct spatial compartmentalization and EoE disease status. In active EoE, they assume a proinflammatory state and locally proliferate, and they remain activated and poised to reinitiate inflammation even during disease remission.

Desmoplakin and periplakin genetically and functionally contribute to eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is a chronic allergic inflammatory disease with a complex underlying genetic etiology. Herein, we conduct whole-exome sequencing of a multigeneration EoE pedigree (discovery set) and 61 additional multiplex families with EoE (replication set). A series of rare, heterozygous, missense variants are identified in the genes encoding the desmosome-associated proteins DSP and PPL in 21% of the multiplex families. Esophageal biopsies from patients with these variants retain dilated intercellular spaces and decrease DSP and PPL expression even during disease remission. These variants affect barrier integrity, cell motility and RhoGTPase activity in esophageal epithelial cells and have increased susceptibility to calpain-14-mediated degradation. An acquired loss of esophageal DSP and PPL is present in non-familial EoE. Taken together, herein, we uncover a pathogenic role for desmosomal dysfunction in EoE, providing a deeper mechanistic understanding of tissue-specific allergic responses.

Functional role of kallikrein 5 and proteinase-activated receptor 2 in eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is a chronic, food antigen-driven, inflammatory disease of the esophagus and is associated with impaired barrier function. Evidence is emerging that loss of esophageal expression of the serine peptidase inhibitor, kazal type 7 (SPINK7), is an upstream event in EoE pathogenesis. Here, we provide evidence that loss of SPINK7 mediates its pro-EoE effects via kallikrein 5 (KLK5) and its substrate, protease-activated receptor 2 (PAR2). Overexpression of KLK5 in differentiated esophageal epithelial cells recapitulated the effect of SPINK7 gene silencing, including barrier impairment and loss of desmoglein-1 expression. Conversely, KLK5 deficiency attenuated allergen-induced esophageal protease activity, modified commensal microbiome composition, and attenuated eosinophilia in a murine model of EoE. Inhibition of PAR2 blunted the cytokine production associated with loss of SPINK7 in epithelial cells and attenuated the allergen-induced esophageal eosinophilia in vivo. Clinical samples substantiated dysregulated PAR2 expression in the esophagus of patients with EoE, and delivery of the clinically approved drug α1 antitrypsin (A1AT, a protease inhibitor) inhibited experimental EoE. These findings demonstrate a role for the balance between KLK5 and protease inhibitors in the esophagus and highlight EoE as a protease-mediated disease. We suggest that antagonizing KLK5 and/or PAR2 has potential to be therapeutic for EoE.