Stable preparation of in vivo transplantable periodontal ligament-derived mesenchymal stem cell sheets in thermoresponsive culture dishes with tunable cell detachability

Kazuki Morita; Masamichi Nakayama; Jiacheng Wang; Satoru Onizuka; Masahiro Hatasa; Yujin Ohsugi; Yosuke Tsuchiya; Hiromi Niimi; Anhao Liu; Hideaki Sakai; Teruo Okano; Takanori Iwata

doi:10.1016/j.reth.2025.01.004

Stable preparation of in vivo transplantable periodontal ligament-derived mesenchymal stem cell sheets in thermoresponsive culture dishes with tunable cell detachability

Regen Ther. 2025 Jan 13:28:345-357. doi: 10.1016/j.reth.2025.01.004. eCollection 2025 Mar.

Authors

Kazuki Morita¹, Masamichi Nakayama², Jiacheng Wang¹, Satoru Onizuka³, Masahiro Hatasa^{1

4}, Yujin Ohsugi^{5

6

7}, Yosuke Tsuchiya¹, Hiromi Niimi⁸, Anhao Liu¹, Hideaki Sakai⁹, Teruo Okano^{2

9}, Takanori Iwata¹

Affiliations

¹ Department of Periodontology, Graduate School of Medical and Dental Sciences, Institute of Science Tokyo (Science Tokyo), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
² Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, 8-1 Kawadacho, Shinjuku-ku, Tokyo 162-8666, Japan.
³ Division of Periodontology, Department of Oral Function, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka, 803-8580, Japan.
⁴ Liver Center, Saga University Hospital, Saga, Saga, Japan.
⁵ Section of Vascular Cell Biology, Joslin Diabetes Center, Harvard Medical School, One Joslin Place, Boston, MA, 02215, USA.
⁶ Department of Oral Biology, Graduate School of Medical and Dental Sciences, Institute of Science Tokyo (Science Tokyo), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
⁷ Oral Science Center, Institute of Science Tokyo (Science Tokyo), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
⁸ Department of Lifetime Oral Health Care Science, Graduate School of Medical and Dental Sciences, Institute of Science Tokyo (Science Tokyo), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
⁹ Organization of Cell Sheet Tissue Engineering Regenerative Medicine Initiatives (CSTERM), Ark Mori Building 36F, 1-12-32 Akasaka, Minato-ku, Tokyo 107-6036, Japan.

Abstract

Tissue engineering plays a pivotal role in the advancement of regenerative medicine. Thermoresponsive culture dishes, coated with specialized polymers that control cell adhesion through temperature fluctuations, enable the processing of cells into sheets for medical applications while maintaining their intact state. Cell sheets prepared using these culture dishes have been incorporated into several commercial pharmaceutical products. However, controlling the detachability of cell sheets using conventional thermoresponsive culture dishes remains a challenge, and often leads to unexpected detachment during cultivation. In this study, we developed a thermoresponsive culture dish with tunable cell detachability using a thermoresponsive block copolymer, poly(butyl methacrylate)-b-poly(N-isopropylacrylamide) (PBMA-PIPAAm), which is a specialized polymer that allows precise control of the amount of surface-immobilized polymer and polymer layer thickness. Culturing periodontal ligament-derived mesenchymal stem cells on these dishes demonstrated fully tunable detachability without compromising cell properties compared to conventional thermoresponsive dishes (UpCell®). Thermoresponsive PBMA-PIPAAm-coated culture dishes enable the complete on-demand detachment of transplantable cell sheets, thereby avoiding unexpected detachment that may increase production costs and reduce technical hurdles in the manufacturing process. The PBMA-PIPAAm coating method has the potential to contribute to biomedical and clinical applications of mesenchymal stem cell sheets.

Keywords: Cell sheet; Periodontal ligament derived mesenchymal stem cell; Periodontitis; Polymer coating method; Thermoresponsive culture dish.