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Originally published In Press as doi:10.1167/iovs.08-2262 on January 10, 2009
 (Investigative Ophthalmology and Visual Science. 2009;50:2109-2115.)
© 2009 by The Association for Research in Vision and Ophthalmology, Inc.
doi:10.1167/iovs.08-2262
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The Use of Human Mesenchymal Stem Cell–Derived Feeder Cells for the Cultivation of Transplantable Epithelial Sheets

Masahiro Omoto,1,2 Hideyuki Miyashita,1,2 Shigeto Shimmura,1,3 Kazunari Higa,3 Tetsuya Kawakita,1 Satoru Yoshida,1,3 Michael McGrogan,4 Jun Shimazaki,1,3 and Kazuo Tsubota1,3

1From the Department of Ophthalmology, Keio University School of Medicine, Tokyo, Japan; the 3Department of Ophthalmology, Tokyo Dental College, Chiba, Japan; and 4SanBio Inc, Mountain View, California.

PURPOSE. To report the efficacy of human bone marrow–derived mesenchymal stem cells as a source of feeder cells for the cultivation of transplantable corneal epithelial cell sheets.

METHODS. Human mesenchymal stem cells (marrow adherent stem cells; MASCs) were cultured in {alpha}-modified Eagle’s medium with 10% serum and were treated with mitomycin C. Expression of cytokines in MASCs was confirmed by reverse transcription-polymerase chain reaction. Human limbal epithelial cells were cocultured with MASCs or 3T3 feeder cells to compare colony-forming efficiency (CFE). Limbal epithelial cells were cultured on MASCs or 3T3 feeder cells at the air-liquid interface to allow stratification, and stratified epithelial sheets were analyzed by immunohistochemistry against cytokeratin 3 (K3), K15, p63{alpha}, and ABCG2. Rabbit limbal epithelial cell sheets were cultivated with MASC feeder cells and transplanted to the ocular surface of the limbal-deficient rabbits. Epithelial grafts were observed by slit lamp microscopy for 4 weeks and then evaluated by histology and immunohistochemistry against K3 and K4.

RESULTS. MASC feeder cells expressed keratinocyte growth factor, hepatocyte growth factor, and N-cadherin. The CFE of human limbal epithelial cells was similar in MASC and 3T3 feeder groups. Stratified cell sheets were successfully cultivated with MASC feeder cells expressing K3, K15, p63{alpha}, and ABCG2. Transplanted epithelial sheets regenerated the corneal phenotype in limbal-deficient rabbits.

CONCLUSIONS. MASC-derived feeder cells are suitable for the engineering of epithelial sheets, avoiding the use of potentially hazardous xenologic feeder cells.




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E. A. Meyer-Blazejewska, F. E. Kruse, K. Bitterer, C. Meyer, C. Hofmann-Rummelt, P. H. Wunsch, and U. Schlotzer-Schrehardt
Preservation of the Limbal Stem Cell Phenotype by Appropriate Culture Techniques
Invest. Ophthalmol. Vis. Sci., February 1, 2010; 51(2): 765 - 774.
[Abstract] [Full Text] [PDF]


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