Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Limbal and Central Corneal Epithelia

ABSTRACT: Limbal and central regoins of the rat cornea epithelium were used to constract 2 SAGE libraries, in order to identify candidate genes to distinguish stem cell from differentiated central cornea epithelium cells. Keywords: gene expression SAGE-based, count cornea epithelium was scraped from central cornea and limbal Epithelium regions of 6 week male Wister rat 16 and 48 eye (respectively).

ORGANISM(S): Rattus norvegicus

SUBMITTER: Hagit Ulanovsky

PROVIDER: E-GEOD-4121 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Serial analysis of gene expression (SAGE) in the rat limbal and central corneal epithelium.

Adachi Wakako W Ulanovsky Hagit H Li Yan Y Norman Barbara B Davis Janine J Piatigorsky Joram J

Investigative ophthalmology & visual science 20060901 9

<h4>Purpose</h4>To identify genes preferentially expressed in the stem-cell-rich limbal epithelium of the rat cornea.<h4>Methods</h4>The limbal and central corneal epithelial cells of 6-week-old rats were isolated by microdissection. Serial analysis of gene expression (SAGE) libraries were constructed and analyzed, and in situ hybridization, reverse transcription-polymerase chain reaction (RT-PCR) and cDNA cloning were conducted by conventional procedures.<h4>Results</h4>The rat limbal and centr ...[more]

PMID: 16936091

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Project description:Limbal stem cells (LSCs), known as corneal epithelial stem cells, are located at the basal epithelial layer of the corneal limbus and serve an important function in maintaining the homeostasis of the corneal epithelium. Several putative molecular markers of LSCs have been previously identified. However, the specificity of these markers remains largely controversial. To address this gap in the current understanding of LSCs, we performed a transcriptome profiling of heterogeneous corneal limbal basal cells using single-cell transcriptomics technology to identify LSCs and their exclusive markers. We isolated limbal basal cells from two young donors, constructed scRNA-seq libraries, generated RNA-sequencing data using the 10x Genomics platform, and then finally obtained the transcriptome of about 18,000 individual single-cells using Cell ranger (https://10xgenomics.com). Next, we performed quality control, filtering and data integration using Seurat package (https://satijalab.org/seurat), about 16,400 cells were retained for further downstream analysis such as dimensional reduction, unsupervised clustering, and Differentially expressed gene selection, trajectory analysis and visualization. We identified 11 unique clusters of cells assigned to a putative cell type based on published known biomarkers for differentiation, proliferation and putative epithelial stem cells. As a results, we found 2 Terminally Differentiated Cell (TDC), 3 Post-Mitotic Cell (PMC), 1 Transient Amplifying Cell (TAC), 2 Limbal Progenitor Cell (LPC), Putative Limbal Stem Cell (LSC) and 2 Melanocyte (MC) sub-cell type clusters. Furthermore, we confirmed the trajectory order of LSC differentiation for 9 clusters using Pseudotemporal and Functional PCA analysis. Lastly, we validated each assigned cell subtypes and determined their location in human corneal limbus tissue with 9 markers using RNAscope We were able to reveal the heterogeneity of corneal limbal basal epithelium by defining novel dynamic trajectories for cell types in a pseudotemporal manner. This approach allowed us to identify the distinct clusters of LSCs and progenitors with exclusively expressed markers, and might apply for translational research on regenerating a normal corneal epithelium and restoring vision.

Dataset Information

Limbal and Central Corneal Epithelia

Publications

Serial analysis of gene expression (SAGE) in the rat limbal and central corneal epithelium.

Similar Datasets

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