Isolation of Multipotent Neural Crest-Derived Stem Cells from the Adult Mouse Cornea

Authors

  • Satoru Yoshida,

    1. Cornea Center, Tokyo Dental College, Ichikawa, Chiba, Japan
    2. Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
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  • Shigeto Shimmura M.D.,

    Corresponding author
    1. Cornea Center, Tokyo Dental College, Ichikawa, Chiba, Japan
    2. Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
    • Department of Ophthalmology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. Telephone: +81-3-3353-1211; Fax: +81-3-3359-8302
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  • Narihito Nagoshi,

    1. Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
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  • Keiichi Fukuda,

    1. Department of Regenerative Medicine and Advanced Cardiac Therapeutics, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
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  • Yumi Matsuzaki,

    1. Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
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  • Hideyuki Okano M.D.,

    Corresponding author
    1. Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
    • Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. Telephone: +81-3-3353-1211; Fax: +81-3-3357-5445
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  • Kazuo Tsubota

    1. Cornea Center, Tokyo Dental College, Ichikawa, Chiba, Japan
    2. Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan
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Abstract

We report the presence of neural crest-derived corneal precursors (COPs) that initiate spheres by clonal expansion from a single cell. COPs expressed the stem cell markers nestin, Notch1, Musashi-1, and ABCG2 and showed the side population cell phenotype. COPs were multipotent with the ability to differentiate into adipocytes, chondrocytes, as well as neural cells, as shown by the expression of β-III-tubulin, glial fibrillary acidic protein, and neurofilament-M. COP spheres prepared from E/nestin-enhanced green fluorescent protein (EGFP) mice showed induction of EGFP expression that was not originally observed in the cornea, indicating activation of the neural-specific nestin second intronic enhancer in culture. COPs were Sca-1+, CD34+, CD45, and c-kit. Numerous GFP+ cells were observed in the corneas of mice transplanted with whole bone marrow of transgenic mice ubiquitously expressing GFP; however, no GFP+ COP spheres were initiated from these mice. On the other hand, COP spheres from transgenic mice encoding P0-Cre/Floxed-EGFP as well as Wnt1-Cre/Floxed-EGFP were GFP+, indicating the neural crest origin of COPs, which was confirmed by the expression of the embryonic neural crest markers Twist, Snail, Slug, and Sox9. Taken together, these data indicate the existence of neural crest-derived, multipotent stem cells in the adult cornea.

Ancillary