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Autophagy Contributes to Homeostasis in Esophageal Epithelium Where High Autophagic Vesicle Level Marks Basal Cells With Limited Proliferation and Enhanced Self-Renewal Potential
Klochkova, Alena Karami, Adam L. Fuller, Annie D. Parham, Louis R. Panchani, Surali R. Natarajan, Shruthi Jackson, Jazmyne L. Mu, Anbin Tan, Yinfei Cai, Kathy Q.
Klochkova, Alena
Karami, Adam L.
Fuller, Annie D.
Parham, Louis R.
Panchani, Surali R.
Natarajan, Shruthi
Jackson, Jazmyne L.
Mu, Anbin
Tan, Yinfei
Cai, Kathy Q.
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Journal article
Date
2024-03-05
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Committee member
Department
Cancer and Cellular Biology
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DOI
https://doi.org/10.1016/j.jcmgh.2024.02.018
Abstract
Background & Aims
Autophagy plays roles in esophageal pathologies both benign and malignant. Here, we aim to define the role of autophagy in esophageal epithelial homeostasis.
Methods
We generated tamoxifen-inducible, squamous epithelial-specific Atg7 (autophagy related 7) conditional knockout mice to evaluate effects on esophageal homeostasis and response to the carcinogen 4-nitroquinoline 1-oxide (4NQO) using histologic and biochemical analyses. We fluorescence-activated cell sorted esophageal basal cells based on fluorescence of the autophagic vesicle (AV)-identifying dye Cyto-ID and then subjected these cells to transmission electron microscopy, image flow cytometry, three-dimensional organoid assays, RNA sequencing, and cell cycle analysis. Three-dimensional organoids were subjected to passaging, single-cell RNA sequencing, cell cycle analysis, and immunostaining.
Results
Genetic autophagy inhibition in squamous epithelium resulted in increased proliferation of esophageal basal cells under homeostatic conditions and also was associated with significant weight loss in mice treated with 4NQO that further displayed perturbed epithelial tissue architecture. Esophageal basal cells with high AV level (Cyto-IDHigh) displayed limited organoid formation capability on initial plating but passaged more efficiently than their counterparts with low AV level (Cyto-IDLow). RNA sequencing suggested increased autophagy in Cyto-IDHigh esophageal basal cells along with decreased cell cycle progression, the latter of which was confirmed by cell cycle analysis. Single-cell RNA sequencing of three-dimensional organoids generated by Cyto-IDLow and Cyto-IDHigh cells identified expansion of 3 cell populations and enrichment of G2/M-associated genes in the Cyto-IDHigh group. Ki67 expression was also increased in organoids generated by Cyto-IDHigh cells, including in basal cells localized beyond the outermost cell layer.
Conclusions
Autophagy contributes to maintenance of the esophageal proliferation-differentiation gradient. Esophageal basal cells with high AV level exhibit limited proliferation and generate three-dimensional organoids with enhanced self-renewal capacity.
Description
Citation
Alena Klochkova, Adam L. Karami, Annie D. Fuller, Louis R. Parham, Surali R. Panchani, Shruthi Natarajan, Jazmyne L. Jackson, Anbin Mu, Yinfei Tan, Kathy Q. Cai, Andres J. Klein-Szanto, Amanda B. Muir, Marie-Pier Tétreault, Xavier Graña, Kathryn E. Hamilton, Kelly A. Whelan, Autophagy Contributes to Homeostasis in Esophageal Epithelium Where High Autophagic Vesicle Level Marks Basal Cells With Limited Proliferation and Enhanced Self-Renewal Potential, Cellular and Molecular Gastroenterology and Hepatology, Volume 18, Issue 1, 2024, Pages 15-40, ISSN 2352-345X, https://doi.org/10.1016/j.jcmgh.2024.02.018. (https://www.sciencedirect.com/science/article/pii/S2352345X24000523)
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Elsevier
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Cellular and Molecular Gastroenterology and Hepatology, Vol. 18, Iss. 1
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