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Single-cell RNA sequencing identifies diverse roles of epithelial cells in idiopathic pulmonary fibrosis
Yan Xu, … , Barry R. Stripp, Jeffrey A. Whitsett
Yan Xu, … , Barry R. Stripp, Jeffrey A. Whitsett
Published December 8, 2016
Citation Information: JCI Insight. 2016;1(20):e90558. https://doi.org/10.1172/jci.insight.90558.
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Research Article Inflammation

Single-cell RNA sequencing identifies diverse roles of epithelial cells in idiopathic pulmonary fibrosis

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Abstract

Idiopathic pulmonary fibrosis (IPF) is a lethal interstitial lung disease characterized by airway remodeling, inflammation, alveolar destruction, and fibrosis. We utilized single-cell RNA sequencing (scRNA-seq) to identify epithelial cell types and associated biological processes involved in the pathogenesis of IPF. Transcriptomic analysis of normal human lung epithelial cells defined gene expression patterns associated with highly differentiated alveolar type 2 (AT2) cells, indicated by enrichment of RNAs critical for surfactant homeostasis. In contrast, scRNA-seq of IPF cells identified 3 distinct subsets of epithelial cell types with characteristics of conducting airway basal and goblet cells and an additional atypical transitional cell that contributes to pathological processes in IPF. Individual IPF cells frequently coexpressed alveolar type 1 (AT1), AT2, and conducting airway selective markers, demonstrating “indeterminate” states of differentiation not seen in normal lung development. Pathway analysis predicted aberrant activation of canonical signaling via TGF-β, HIPPO/YAP, P53, WNT, and AKT/PI3K. Immunofluorescence confocal microscopy identified the disruption of alveolar structure and loss of the normal proximal-peripheral differentiation of pulmonary epithelial cells. scRNA-seq analyses identified loss of normal epithelial cell identities and unique contributions of epithelial cells to the pathogenesis of IPF. The present study provides a rich data source to further explore lung health and disease.

Authors

Yan Xu, Takako Mizuno, Anusha Sridharan, Yina Du, Minzhe Guo, Jie Tang, Kathryn A. Wikenheiser-Brokamp, Anne-Karina T. Perl, Vincent A. Funari, Jason J. Gokey, Barry R. Stripp, Jeffrey A. Whitsett

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Figure 7

Immunofluorescence confocal microscopy identifies atypical epithelial cell differentiation in IPF.

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Immunofluorescence confocal microscopy identifies atypical epithelial ce...
(A) Peripheral samples of normal and IPF lung tissue were stained for epithelial cell markers used to identify AT2 (HTII-280 and ABCA3), AT1 (HOPX), ciliated (TUBA4A), goblet (MUC5B) cells. Yellow staining indicates coexpression of the proteins. HTII-280 and ABCA3, normally restricted to peripheral/alveolar epithelial cells in normal lung, were expressed in IPF lesions; cystic lesions were variably lined by hyperplastic AT2 cells that stained for ABCA3 in close proximity to MUC5B (goblet) or TUBA4A (ciliated) stained cells. Abnormally shaped epithelial cells variably staining for HOPX, ABCA3, and HTII-280 were characteristic of IPF tissues that generally lacked normal squamous AT1 cells. (B) Epithelial cells expressing conducting airway and alveolar epithelial cell markers were found in close proximity in the IPF lesions (e.g., TP63, KRT14 and MUC5B) and (ABCA3 and HOPX) respectively are shown. Figures are representative of n = 3–5 control and 9 IPF samples, except for KRT14 (n = 3). Images were obtained at ×10 magnification (scale bars: 200 μm). Insets in yellow boxes are at ×60 magnification
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