May 2004
Volume 45, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2004
Harvest of bioengineered human corneal endothelial cell sheet cultivated on temperature–responsive culture dish
Author Affiliations & Notes
  • T. Sumide
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • K. Nishida
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • M. Yamato
    Institute of Advanced Biomedical Engineering & Science, Tokyo Women's Medical University, Tokyo, Japan
  • T. Ide
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • N. Maeda
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • H. Watanabe
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • A. Kikuchi
    Institute of Advanced Biomedical Engineering & Science, Tokyo Women's Medical University, Tokyo, Japan
  • T. Okano
    Institute of Advanced Biomedical Engineering & Science, Tokyo Women's Medical University, Tokyo, Japan
  • Y. Tano
    Department of Ophthalmology, Osaka University Medical School, Osaka, Japan
  • Footnotes
    Commercial Relationships  T. Sumide, None; K. Nishida, CellSheed C, P; M. Yamato, CellSheed C, P; T. Ide, None; N. Maeda, None; H. Watanabe, None; A. Kikuchi, None; T. Okano, CellSheed C, P; Y. Tano, None.
  • Footnotes
    Support  Grant (15390530) from the Ministry of Education, Culture, Sports, Science and Technology, Japan
Investigative Ophthalmology & Visual Science May 2004, Vol.45, 4787. doi:
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      T. Sumide, K. Nishida, M. Yamato, T. Ide, N. Maeda, H. Watanabe, A. Kikuchi, T. Okano, Y. Tano; Harvest of bioengineered human corneal endothelial cell sheet cultivated on temperature–responsive culture dish . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4787.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : Purpose: Previously, we reported our temperature–responsive culture system that allowed us to fabricate and harvest, in various cell lines, a single, intact cell sheet retaining junctional proteins as well as extracellular matrix proteins. In this study, we have investigated a method to culture and harvest the human corneal endothelial cell (HCEC) sheets from the temperature–responsive dish for clinical use and studied some physiologic characteristics. Methods: HCECs from the remainder of USA donor corneas after penetrating keratoplasty were seeded on temperature–responsive culture dishes, and cultivated at 37°C in a 10% CO2 atmosphere. Several weeks later, fabricated cell sheets were harvested by low temperature treatment. The Na/K ATPase pump sites in the harvested sheet were examined by immunohistochemistry and quantified by a 3H–ouabain binding assay. Results: The cultivated HCECs could be harvested intact as a monolayer. In these sheets, the Na/K ATPase pump sites were located at the lateral borders of the cells. The 3H–ouabain binding assay demonstrated that the number of the pump sites in the sheet is similar to that in the human corneal endothelium in vivo. Conclusions: We have developed a novel method to fabricate and harvest cultivated HCEC sheets using our temperature–responsive culture system. Location and number of Na/K ATPase pump sites in the sheet are similar to in vivo. This bioengineered tissue construct has potential applications for ocular reconstructive surgery.

Keywords: cornea: endothelium • NaK ATPase • transplantation 
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