Single-Cell RNA Transcriptome Helps Define the Limbal/Corneal Epithelial Stem/Early Transit Amplifying Cells and How Autophagy Affects This Population.
Invest Ophthalmol Vis Sci
; 60(10): 3570-3583, 2019 08 01.
Article
em En
| MEDLINE
| ID: mdl-31419300
ABSTRACT
Purpose:
Single-cell RNA-sequencing (scRNA-seq) was used to interrogate the relatively rare stem (SC) and early transit amplifying (TA) cell populations in limbal/corneal epithelia from wild-type and autophagy-compromised mice.Methods:
We conducted scRNA-seq on ocular anterior segmental tissue from wild-type and beclin 1-deficient (beclin1+/-) mice, using a 10X Gemomics pipeline. Cell populations were distinguished by t-distributed stochastic neighbor embedding. Seurat analysis was conducted to compare gene expression profiles between these two groups of mice. Differential protein expression patterns were validated by immunofluorescence staining and immunoblotting.Results:
Unbiased clustering detected 10 distinct populations three clusters of mesenchymal and seven clusters of epithelial cells, based on their unique molecular signatures. A discrete group of mesenchymal cells expressed genes associated with corneal stromal SCs. We identified three limbal/corneal epithelial cell subpopulations designated as stem/early TA, mature TA, and differentiated corneal epithelial cells. Thioredoxin-interacting protein and PDZ-binding kinase (PBK) were identified as novel regulators of stem/early TA cell quiescence. PBK arrested corneal epithelial cells in G2/M phase of the cell cycle. Beclin1+/- mice displayed a decrease in proliferation-associated (Ki67, Lrig1) and stress-response (H2ax) genes. The most increased gene in beclin1+/- mice was transcription factor ATF3, which negatively regulates limbal epithelial cell proliferation.Conclusions:
Establishment of a comprehensive atlas of genes expressed by stromal and epithelial cells from limbus and cornea forms the foundation for unraveling regulatory networks among these distinct tissues. Similarly, scRNA-seq profiling of the anterior segmental epithelia from wild-type and autophagy-deficient mice provides new insights into how autophagy influences proliferation in these tissues.
Texto completo:
1
Bases de dados:
MEDLINE
Assunto principal:
Autofagia
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RNA
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Limbo da Córnea
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Epitélio Corneano
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Células-Tronco Mesenquimais
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Transcriptoma
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
Invest Ophthalmol Vis Sci
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Estados Unidos