A novel isolation technique of progenitor cells in human corneal epithelium using non-tissue culture dishes.
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.
Yokoo
Seiichi
S
Department of Corneal Tissue Regeneration, Tokyo University Graduate School of Medicine, Tokyo, Japan.
Yamagami
Satoru
S
Shimada
Takashi
T
Usui
Tomohiko
T
Sato
Taka-aki
TA
Amano
Shiro
S
Araie
Makoto
M
Hamuro
Junji
J
D000328:Adult; D000368:Aged; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000096:biosynthesis*; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D019573:Epithelium, Corneal / Q000166:cytology* / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D016850:Limbus Corneae / Q000378:metabolism*; D008875:Middle Aged; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D000097794:Kalinin
D015815:Cell Adhesion Molecules; D000097794:Kalinin