Single-cell RNA-Seq of human esophageal epithelium in homeostasis and allergic inflammation
Mark Rochman, Ting Wen, Michael Kotliar, Phillip J. Dexheimer, Netali Ben-Baruch Morgenstern, Julie M. Caldwell, Hee-Woong Lim, Marc E. Rothenberg
Mark Rochman, Ting Wen, Michael Kotliar, Phillip J. Dexheimer, Netali Ben-Baruch Morgenstern, Julie M. Caldwell, Hee-Woong Lim, Marc E. Rothenberg
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Research Article Immunology Inflammation

Single-cell RNA-Seq of human esophageal epithelium in homeostasis and allergic inflammation

Abstract

Inflammation of the esophageal epithelium is a hallmark of eosinophilic esophagitis (EoE), an emerging chronic allergic disease. Herein, we probed human esophageal epithelial cells at single-cell resolution during homeostasis and EoE. During allergic inflammation, the epithelial differentiation program was blocked, leading to loss of KRT6hi differentiated populations and expansion of TOP2hi proliferating, DSPhi transitioning, and SERPINB3hi transitioning populations; however, there was stability of the stem cell–enriched PDPNhi basal epithelial compartment. This differentiation program blockade was associated with dysregulation of transcription factors, including nuclear receptor signalers, in the most differentiated epithelial cells and altered NOTCH-related cell-to-cell communication. Each epithelial population expressed genes with allergic disease risk variants, supporting their functional interplay. The esophageal epithelium differed notably between EoE in histologic remission and controls, indicating that remission is a transitory state poised to relapse. Collectively, our data uncover the dynamic nature of the inflamed human esophageal epithelium and provide a framework to better understand esophageal health and disease.

Authors

Mark Rochman, Ting Wen, Michael Kotliar, Phillip J. Dexheimer, Netali Ben-Baruch Morgenstern, Julie M. Caldwell, Hee-Woong Lim, Marc E. Rothenberg

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Figure 3

Functional and molecular properties of the human esophageal basal layer cells in homeostasis.

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Functional and molecular properties of the human esophageal basal layer ...
(A) Schematic representation of the experimental design to purify and characterize basal layer cells (upper left). Podoplanin (PDPN) staining depicting basal layer localization from the Human Protein Atlas (HPA) (lower left). A representative FACS plot shows the gating strategy to purify PDPNhi for basal cells and PDPNlo for suprabasal cells (right). (B) IHC images are screenshots from HPA of the representative markers for basal and suprabasal cells identified by differential analysis of bulk RNA-Seq data comparing PDPNhi and PDPNlo cells; see the HPA for magnification of original images (4). (C) Average expression of top 20 PDPNhi and PDPNlo marker genes projected onto UMAP of quiescent and proliferating cells from healthy controls distinguishing basal and suprabasal cells; the dotted line separates quiescent and proliferating cells. (D) Cell cycle status of cells inferred by Seurat projected onto UMAP of quiescent and proliferating cells. (E) Ex vivo colony-forming assay for sorted basal and suprabasal cells. Each line represents cells from a biopsy obtained from a distinct individual. A total number of colonies per 2000 seeded cells are plotted. Representative images of the high-power microscopic field show colony growth of basal PDPNhi cells (arrows; upper image, ×10 magnification). Cells in the PDPNlo image are mitotically inactivated murine fibroblasts that served as feeder layer cells for colony growth. (F) Immunofluorescence images of the colonies grown from the PDPNhi cells. Scale bar: 50 μM.