Seminar: Cell Sheet Tissue Engineering and Their Clinical Applications

1-s2.0-S1369702104002342-gr1 10th September 2007 – 11:30

Prof. Teruo Okano (Tokyo Women’s Medical University)



U.M. Grassano, Campus


My research group develops polymeric “intelligent surfaces” proven useful and published for applications in medicine [1-4] and biotechnology [5-8]. We have synthesized several temperature-responsive polymers of N-(isopropylacrylamide), and have attached these polymers to various surfaces. This new surface modification technology demonstrates several intelligent functions, including dramatic, reversible surface property alterations with slight temperature changes, useful for new applications. We use temperature-responsive cell culture to prepare novel living cell sheets [9, 10] that represent attractive, interesting new alternatives to tissue as scientific models and as living clinical tissue replacements.

Ocular trauma commonly causes corneal opacification and visual loss because of limbal stem cell deficiency and inability to regenerate the ocular surface. Although corneal transplantation is required in these cases, numbers of donor corneas are very limited. We have initiated clinical treatment to transplant our engineered ocular epithelial cell sheet cultured either from a patient’s biopsied limbal stem cells or oral mucosal cells. With intact native adhesive matrix, cell sheets bind structurally and functionally to eye stroma [11, 12] without sutures. In more than 30 human patients, significant improvement of visual acuity is observed beyond 36months to date.

Using cardiomyocyte heart muscle cells, we have demonstrated a novel tissue engineering methodology layering harvested cell sheets to construct 3-D functional cardiac tissues. When two pulsing cell sheets are layered,, the two sheets synchronize their pulsations through gap junctions between the sheets. We have succeeded in demonstrating long-term survival in vivo for these engineered pulsatile cardiac grafts [13-15]. Analogous cell sheet patches fabricated from autologous skeletal myoblasts are now on-going in large animal models. Cardiac tissue engineering based on our “Cell Sheet Engineering” [16-19] should prove useful for heart model fabrication and clinical cardiovascular tissue repair.

These new cell sheet manipulation techniques are promising for many cell types and tissue structures, and device shapes. We believe that two- and three-dimensional cell sheet manipulation – Cell Sheet Engineering – represents a useful, fundamental, generalized technique for next-generation tissue engineering and regenerative medicine.


  1. M. Yamato and T. Okano, “Cell Sheet Engineering”, Materials Today, 7 (5), 42-47(2004).

  2. J. Yang, M. Yamato and T. Okano, “Cell-sheet engineering using intelligent surfaces”, MRS Bulletin, 30(3), 189-193 (2005).

  3. J. Yang, M. Yamato, C. Kohno, A. Nishimoto, H. Sekine, F. Fukai, T. Okano, “Cell sheet engineering: recreating tissues without biodegradable scaffolds”, Biomaterials, 26 (33), 6415-6422 (2005).

  4. J. Yang, M. Yamato, K. Nishida, Y. Hayashida, T Shimizu, A. Kikuchi, Y. Tano and T. Okano, “Corneal epithelial stem cell delivery using cell sheet engineering: Notlost in transplantation”, Journal of Drug Targeting, 14 (7) 471-482 (2006).

  5. A. Kikuchi, and T. Okano, “7. Hydrogels :Stimuli-sensitive hydrogels”, Polymeric Drug Delivery Systems, Drugs and the pharmaceutical sciences, 148, 275-322, Edited by Glen S. Kwon, Taylor & Francis (2005),

  6. H. Kanazawa, Y. Matsushima and T. Okano, “Temperature-responsive chromatography”, Advances in Chromatography, Edited by P. R. Brown, E. Grushka, Marcel Dekker, Inc., 41, 311-336 (2001).

  7. N. Idoda, A. Kikuchi, J. Kobayashi, Y. Akiyama, K. Sakai and T. Okano, “Thermal modulated interaction of aqueous steroids using polymer-grafted capillaries”, Langmuir, 22 (1), 425-430 (2006).

  8. K. Itoga, J. Kobayashi, M. Yamato, A. Kikuchi and T.Okano, “Maskless liquid-crystal-display projection photolithography for improved design flexidility of cellular micropatterns”, Biomaterials, 27 (15), 3005-3009 (2006).

  9. N. Yamada, T. Okano, H. Sakai, F. Karikusa, Y. Sawasaki and Y. Sakurai, “Thermo-Responsive Polymeric Surfaces; Control of Attachment and Detachment of Cultured Cells”, Macromol. Chem. Rapid Commun., 11(11), 571-576 (1990).

  10. T. Okano, N. Yamada, H. Sakai and Y. Sakurai, “A Novel Recovery System for Cultured Cells Using Plasma-Treated Polystyrene Dishes Grafted with Poly (N-isopropylacrylamide)”, J. Biomed. Mater. Res., 27(10), 1243-1251 (1993)

  11. K. Nishida, M. Yamato, Y. Hayashida, K. Watanabe, N. Maeda, H. Watanabe, S. Nagai, A. Kikuchi, Y. Tano and T. Okano, “Functional bioengineered corneal epithelial sheet grafts from corneal stem cells expanded ex vivo on a temperature-responsive cell culture surface”, Transplantation, 77(3), 379-385 (2004).

  12. K. Nishida, M. Yamato, Y. Hayashida, K. Watanabe, K. Yamamoto, E. Adachi, S. Nagai, A. Kikuchi, N. Maeda, H. Watanabe, T. Okano and Y. Tano, “Corneal reconstruction with tissue-engineered cell sheets composed of autologous oral mucosal epithelium”, N. Engl. J. Med., 351 (12), 1187-1196 (2004).

  13. T. Shimizu, M. Yamato, A. Kikuchi and T. Okano, “Two-dimensional manipulation of cardiac myocyte sheets utilizing temperature-responsive culture dishes augments the pulsatile amplitude”, Tissue Engineering, 7(2), 141-151 (2001).

  14. T. Shimizu, M. Yamato, Y. Isoi, T. Akutsu, T. Setomaru, K. Abe, A. Kikuchi, M. Umezu and T. Okano, “Fabrication of pulsatile cardiac tissue grafts using a novel 3-dimensional cell sheet manipulation technique and temperature-responsive cell culture surfaces”, Circ Res. 90:e40-e48 (2002).

  15. Y. Haraguchi, T. Shimizu, M. Yamato, A Kikuchi and, T. Okano, “Electrical coupling of cardiomyocyte sheets occurs rapidly via functional gap junction formation”, Biomaterials, 27 (27), 4765-4774 (2006).

  16. H. Sekine, T. Shimizu, S. Kosaka, E. Kobayashi and T. Okano, “Cardiomyocyte bridging between hearts and bio-engineered myocardial tissues with mesenchymal transition of mesothelial cells”, J. Heart Lung Transplant., 25 (3), 324-332 (2006).

  17. T. Shimizu, H. Sekine, J. Yang, Y. Isoi, M. Yamato, A. Kikuchi, E. Kobayashi and T. Okano, “Polysurgery of cell sheet grafts overcomes diffusion limits to produce thick, vascularized myocardial tissues”, FASEB J., 20 (6), 708-710 (2006).

  18. H. Hata, G. Matsumiya, S. Miyagawa, H. Kondo, N. Kawaguchi, N. Matsuura, T. Shimizu, T. Okano, H. Matsuda Hikaru and Y. Sawa, “Grafted skeletal myoblast sheets attenuate myocardial remodeling in pacing-induced canine heart failure model”, J. Thorac. Cardiovasc. Surg., 132 (4) 918-924 (2006).

  19. Y. Miyahara, N. Nagaya, M. Kataoka, B. Yanagawa, K. Tanaka, H. Hao, K. Ishino, H. Ishida, T. Shimizu, K. Kangawa, S. Sano, T. Okano, S. Kitamura and H. Mori, “Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction”, Nature Med., 12 (4), 459-465 (2006).

Related Information:
Tokyo Women’s Medical University

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