CCR10+ epithelial cells from idiopathic pulmonary fibrosis lungs drive remodeling
David M. Habiel, Milena S. Espindola, Isabelle C. Jones, Ana Lucia Coelho, Barry Stripp, Cory M. Hogaboam
David M. Habiel, Milena S. Espindola, Isabelle C. Jones, Ana Lucia Coelho, Barry Stripp, Cory M. Hogaboam
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Research Article Cell biology Pulmonology

CCR10+ epithelial cells from idiopathic pulmonary fibrosis lungs drive remodeling

Abstract

Idiopathic pulmonary fibrosis (IPF) is a devastating fibrotic lung disease of unknown etiology and limited therapeutic options. In this report, we characterize what we believe is a novel CCR10+ epithelial cell population in IPF lungs. There was a significant increase in the percentage of CCR10+ epithelial cells in IPF relative to normal lung explants and their numbers significantly correlated to lung remodeling in humanized NSG mice. Cultured CCR10-enriched IPF epithelial cells promoted IPF lung fibroblast invasion and collagen 1 secretion. Single-cell RNA sequencing analysis showed distinct CCR10+ epithelial cell populations enriched for inflammatory and profibrotic transcripts. Consistently, cultured IPF but not normal epithelial cells induced lung remodeling in humanized NSG mice, where the number of CCR10+ IPF, but not normal, epithelial cells correlated with hydroxyproline concentration in the remodeled NSG lungs. A subset of IPF CCR10hi epithelial cells coexpress EphA3 and ephrin A signaling induces the expression of CCR10 by these cells. Finally, EphA3+CCR10hi epithelial cells induce more consistent lung remodeling in NSG mice relative to EphA3–CCR10lo epithelial cells. Our results suggest that targeting epithelial cells, highly expressing CCR10, may be beneficial in IPF.

Authors

David M. Habiel, Milena S. Espindola, Isabelle C. Jones, Ana Lucia Coelho, Barry Stripp, Cory M. Hogaboam

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Figure 8

EphA3 signaling induces CCR10 and EphA3 transcript expression and promotes TGF-β1–induced VIM and FN1 transcript expression in IPF relative to normal epithelial cells.

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EphA3 signaling induces CCR10 and EphA3 transcript expression and promot...
(A) Normal and IPF CRC-cultured epithelial cells were stained with fluorescently conjugated anti-EpCAM, -CCR10, and -EphA3 antibodies and analyzed by flow cytometry. Shown are representative dot plots from normal (top) and IPF (bottom) epithelial cultures depicting EpCAM+CCR10+EphA3+ cells. (B) Shown is the percentage of EpCAM+CCR10+EphA3+ cells in normal and IPF epithelial cultures. n = 3–4/group. Data shown are the mean ± SEM. *P ≤ 0.05 via unpaired parametric 2-tailed t test. (C) Magnetic sorting of EphA3+ or EphA3– cells followed by qPCR analysis for various lineage markers. n = 6/group. Data shown are the mean ± SEM. **P ≤ 0.01 via 2-way ANOVA corrected with Bonferroni’s test. (D) RNA was extracted from PEB- or CRC-cultured epithelial cells and qPCR analysis was performed for EphA3. Shown is the average EphA3 expression in CRC-cultured epithelial cells normalized to PEB-cultured cells. n = 3/group. Data shown are the mean ± SEM. *P ≤ 0.05 via unpaired parametric 2-tailed t test with Welch’s correction. (E–J) PEB-cultured normal (n = 2; black bars) and IPF (n = 4; white bars) epithelial cells were stimulated with 20 nM preclustered EFNA5-FC, 10 nM TGF-β, or both for 24 hours. As a control, cells were stimulated with saline (vehicle). After stimulation, RNA was extracted, and qPCR analysis was performed for various transcripts. Depicted is the average expression of CCR10 (E), EphA3 (F), KRT5 (G), COL17A1 (H), FN1 (I), and VIM (J) transcripts in stimulated epithelial cells relative to vehicle-treated cells. Data shown are the mean ± SEM. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001 normal versus IPF via 2-way ANOVA corrected with Tukey’s test. IPF, idiopathic pulmonary fibrosis.