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TISSUE-SPECIFIC STEM CELLS

A Novel Isolation Technique of Progenitor Cells in Human Corneal Epithelium Using Non-Tissue Culture Dishes

Departments of ^aCorneal Tissue Regeneration and
^dOphthalmology, Tokyo University Graduate School of Medicine, Tokyo, Japan;
^bLife Science Laboratory, Shimadzu Corporation, Kyoto, Japan;
^cDivision of Advanced Clinical Proteomics, Institute of Medical Science, University of Tokyo, Tokyo, Japan;
^eDepartment of Microbiology and Immunology, Keio University School of Medicine, Tokyo, Japan

Key Words. Progenitor • Corneal epithelium • Epidermal growth factor • Laminin-5

Correspondence: Satoru Yamagami, M.D., Ph.D., Corneal Tissue Regeneration, Tokyo University Graduate School of Medicine, Tokyo, Japan. Telephone: 81-3-5800-8660; Fax: 81-3-5800-9839; e-mail: syamagami-tky{at}umin.ac.jp

Received October 29, 2007; accepted for publication April 14, 2008.
First published online in STEM CELLS EXPRESS   April 24, 2008.

The existence of adult stem cells or progenitor cells in the human corneal epithelium (i.e., self-renewing squamous cells) has long been suggested, but these cells have not yet been isolated. Here we describe a novel isolation technique using non-tissue culture dishes to enrich progenitor cells, which are able to reconstitute a three-dimensional human corneal epithelial equivalent from single cells in serum-, feeder-, and bovine pituitary extract-free medium. These cells showed original tissue-committed differentiation, a high proliferative capacity, and limited self-renewal. Laminin-5 was measured by mass spectrometric analysis. Pretreatment of cells with anti-laminin-5 antibody demonstrated that laminin-5 was important in allowing corneal epithelial progenitor cells to adhere to non-tissue culture dishes. Hydrophilic tubes (used for cell collection throughout this study) are essential for efficient isolation of adherent corneal epithelial progenitor cells expressing laminin-5. These findings indicate that our new technique using non-tissue culture dishes allows the isolation of progenitor cells from human corneal limbal epithelium and that laminin-5 has a critical role in the adhesion of these cells.

Disclosure of potential conflicts of interest is found at the end of this article.