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Published Online: 7 November 2014

Elimination of Remaining Undifferentiated Induced Pluripotent Stem Cells in the Process of Human Cardiac Cell Sheet Fabrication Using a Methionine-Free Culture Condition

Publication: Tissue Engineering Part C: Methods
Volume 21, Issue Number 3


Cardiac tissue engineering is a promising method for regenerative medicine. Although we have developed human cardiac cell sheets by integration of cell sheet-based tissue engineering and scalable bioreactor culture, the risk of contamination by induced pluripotent stem (iPS) cells in cardiac cell sheets remains unresolved. In the present study, we established a novel culture method to fabricate human cardiac cell sheets with a decreased risk of iPS cell contamination while maintaining viabilities of iPS cell-derived cells, including cardiomyocytes and fibroblasts, using a methionine-free culture condition. When cultured in the methionine-free condition, human iPS cells did not survive without feeder cells and could not proliferate or form colonies on feeder cells or in coculture with cells for cardiac cell sheet fabrication. When iPS cell-derived cells after the cardiac differentiation were transiently cultured in the methionine-free condition, gene expression of OCT3/4 and NANOG was downregulated significantly compared with that in the standard culture condition. Furthermore, in fabricated cardiac cell sheets, spontaneous and synchronous beating was observed in the whole area while maintaining or upregulating the expression of various cardiac and extracellular matrix genes. These findings suggest that human iPS cells are methionine dependent and a methionine-free culture condition for cardiac cell sheet fabrication might reduce the risk of iPS cell contamination.

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Published In

cover image Tissue Engineering Part C: Methods
Tissue Engineering Part C: Methods
Volume 21Issue Number 3March 2015
Pages: 330 - 338
PubMed: 25245976


Published in print: March 2015
Published ahead of print: 11 November 2014
Published online: 7 November 2014
Published ahead of production: 23 September 2014
Accepted: 3 September 2014
Received: 10 April 2014


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Katsuhisa Matsuura
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.
Department of Cardiology, Tokyo Women's Medical University, Tokyo, Japan.
Fumiko Kodama
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.
Kasumi Sugiyama
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.
Tatsuya Shimizu
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.
Nobuhisa Hagiwara
Department of Cardiology, Tokyo Women's Medical University, Tokyo, Japan.
Teruo Okano
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.


Address correspondence to:Katsuhisa Matsuura, MD, PhDInstitute of Advanced Biomedical Engineering and ScienceTokyo Women's Medical University8-1 Kawada-choShinjukuTokyo 162-8666Japan
E-mail: [email protected]

Disclosure Statement

Teruo Okano is a founder and director of the board of Cell Seed, Inc., licensing technologies and patents from Tokyo Women's Medical University. Teruo Okano and Tatsuya Shimizu are stakeholders of Cell Seed, Inc. Tokyo Women's Medical University is receiving research funds from CellSeed, Inc. Teruo Okano, Tatsuya Shimizu, and Katsuhisa Matsuura are inventors of bioreactor systems.

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