Labware

Application data for multiple layered cell-sheet (3D tissue constructs).

Construction of 3D tissue by layering cardiomyocyte cell-sheets (homogeneous cell-sheets)

• Place cardiomyocyte cell-sheet to another layer of cardiomyocyte cell-sheet.
• Since the cell-sheet retains viable cell surface proteins and channels, the vertical gap junction between cell-sheets can easily be established.

• Confocal image of 5 layered cardiomyocyte cell-sheets.

• Electric connectivity of the bi-layered cell-sheets has been established successfully.
• The vertical gap junction formation can be ahieved in 30 min.
• Synchronization was confirmed between the bi-layered cardiomyocyte cell-sheets.

References on 3D cardiomyocyte cell-sheets.

1. Masuda, S., Shimizu, T., Yamato, M., and Okano, T. (2008). Cell sheet engineering for heart tissue repair. Advanced drug delivery reviews 60, 277-285.
2. Shimizu, T., Sekine, H., Isoi, Y., Yamato, M., Kikuchi, A., and Okano, T. (2006). Long-term survival and growth of pulsatile myocardial tissue grafts engineered by the layering of cardiomyocyte sheets. Tissue engineering 12, 499-507.
3. Haraguchi, Y., Shimizu, T., Yamato, M., Kikuchi, A., and Okano, T. (2006). Electrical coupling of cardiomyocyte sheets occurs rapidly via functional gap junction formation. Biomaterials 27, 4765-4774.
4. Furuta, A., Miyoshi, S., Itabashi, Y., Shimizu, T., Kira, S., Hayakawa, K., Nishiyama, N., Tanimoto, K., Hagiwara, Y., Satoh, T., Fukada, K., Okano, T. and Ogawa, S. (2006). Pulsatile cardiac tissue grafts using a novel three-dimensional cell sheet manipulation technique functionally integrates with the host heart, in vivo. Circulation research 98, 705-712.
5. Shimizu, T., Yamato, M., Isoi, Y., Akutsu, T., Setomaru, T., Abe, K., Kikuchi, A., Umezu, M., and Okano, T. (2002). Fabrication of pulsatile cardiac tissue grafts using a novel 3-dimensional cell sheet manipulation technique and temperature-responsive cell culture surfaces. Circulation research 90, e40-e48.

Construction of 3D tissue (heterogeneous cell-sheets)

• Hepatocyte is co-cultured together with HAEC (human aortic endothelial cell) cell-sheet to replicate "in vivo"-like cell culture system.
• Co-cultured hepatocyte system maintained high level of albumin secretion (200ug/day) at day 15 whereas that of "hepatocyte only " system was very faint, suggesting that co-culture system resembles "in vivo"-like culture.
  (Data kindly provided by Dr. Ohashi (MD, Ph.D), of Tokyo Women's Medical Hospital)

References on co-culture system.

1. Ohashi, K., Yokoyama, T., Yamato, M., Kuge, H., Kanehiro, H., Tsutsumi, M., Amanuma, T., Iwata, H., Yang, J., Okano, T. and Nakajima, Y. (2007). Engineering functional two- and three-dimensional liver systems in vivo using hepatic tissue sheets. Nat Med 13, 880-885.
   Harimoto, M., Yamato, M., Hirose, M., Takahashi, C., Isoi, Y., Kikuchi, A., and Okano, T. (2002).
2. Novel approach for achieving double-layered cell sheets co-culture: overlaying endothelial cell sheets onto monolayer hepatocytes utilizing temperature-responsive culture dishes. Journal of biomedical materials research 62, 464-470.