Characterization of rabbit limbal epithelial side population cells using RNA sequencing and single-cell qRT-PCR

Sumako Kameishi, Terumasa Umemoto, Yu Matsuzaki, Masako Fujita, Teruo Okano, Takashi Kato, Masayuki Yamato

    Research output: Contribution to journalArticle

    4 Citations (Scopus)

    Abstract

    Corneal epithelial stem cells reside in the limbus, a transitional zone between the cornea and conjunctiva, and are essential for maintaining homeostasis in the corneal epithelium. Although our previous studies demonstrated that rabbit limbal epithelial side population (SP) cells exhibit stem cell-like phenotypes with Hoechst 33342 staining, the different characteristics and/or populations of these cells remain unclear. Therefore, in this study, we determined the gene expression profiles of limbal epithelial SP cells by RNA sequencing using not only present public databases but also contigs that were created by de novo transcriptome assembly as references for mapping. Our transcriptome data indicated that limbal epithelial SP cells exhibited a stem cell-like phenotype compared with non-SP cells. Importantly, gene ontology analysis following RNA sequencing demonstrated that limbal epithelial SP cells exhibited significantly enhanced expression of mesenchymal/endothelial cell markers rather than epithelial cell markers. Furthermore, single-cell quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) demonstrated that the limbal epithelial SP population consisted of at least two immature cell populations with endothelial- or mesenchymal-like phenotypes. Therefore, our present results may propose the presence of a novel population of corneal epithelial stem cells distinct from conventional epithelial stem cells.

    Original languageEnglish
    Pages (from-to)704-709
    Number of pages6
    JournalBiochemical and Biophysical Research Communications
    Volume473
    Issue number3
    DOIs
    Publication statusPublished - 2016 May 6

    Fingerprint

    Side-Population Cells
    RNA Sequence Analysis
    Polymerase chain reaction
    Transcription
    Stem cells
    Reverse Transcription
    Epithelial Cells
    RNA
    Rabbits
    Polymerase Chain Reaction
    Stem Cells
    Transcriptome
    Cells
    Population
    Phenotype
    Endothelial cells
    Gene expression
    Ontology
    Corneal Epithelium
    Genes

    Keywords

    • Corneal epithelial stem cells
    • Gene expression profile
    • Limbal epithelial side population
    • RNA sequencing
    • Single-cell qRT-PCR

    ASJC Scopus subject areas

    • Biochemistry
    • Biophysics
    • Cell Biology
    • Molecular Biology

    Cite this

    Characterization of rabbit limbal epithelial side population cells using RNA sequencing and single-cell qRT-PCR. / Kameishi, Sumako; Umemoto, Terumasa; Matsuzaki, Yu; Fujita, Masako; Okano, Teruo; Kato, Takashi; Yamato, Masayuki.

    In: Biochemical and Biophysical Research Communications, Vol. 473, No. 3, 06.05.2016, p. 704-709.

    Research output: Contribution to journalArticle

    Kameishi, Sumako ; Umemoto, Terumasa ; Matsuzaki, Yu ; Fujita, Masako ; Okano, Teruo ; Kato, Takashi ; Yamato, Masayuki. / Characterization of rabbit limbal epithelial side population cells using RNA sequencing and single-cell qRT-PCR. In: Biochemical and Biophysical Research Communications. 2016 ; Vol. 473, No. 3. pp. 704-709.
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