Hasegawa Akiyuki, Haraguchi Yuji, Shimizu Tatsuya, Okano Teruo
Institute of Advanced Biomedical Engineering and Science, TWIns, Tokyo Women's Medical University, Tokyo, Japan.
J Biomed Mater Res A. 2015 Dec;103(12):3825-33. doi: 10.1002/jbm.a.35526.
Three-dimensional (3D) tissues can be reconstructed by cell sheet technology, and various clinical researches using these constructed tissues have already been initiated to regenerate damaged tissues. While 3D tissues can be easily fabricated by layering cell sheets, the attachment period for cell adhesion between a cell sheet and a culture dish, or double-layered cell sheets normally takes 20-30 min. This study proposed a more rapid fabrication system for bioengineered tissue using cell sheet technology and centrifugation. A C2C12 mouse myoblast sheet harvested from a temperature-responsive culture dish will attach tightly to a culture dish or another cell sheet at 37°C after a 20 min-incubation. However, the same cell sheet centrifuged (12-34 × g) for 3 min also attached tightly to a dish or another cell sheet at 37°C after only a 3 min-incubation. The manipulation time was reduced by approximately two-thirds by centrifugation. The rapid attachments were also cross-sectionally confirmed by optical coherence tomography. These rapidly constructed cell sheet-tissues using centrifugation showed active cell metabolism, cell viability, and very high production of vascular endothelial growth factor, like those prepared by the conventional method; indicating complete cell sheet-attachment without any cell damage. This new system will be a powerful tool in the fields of cell sheet-based tissue engineering and regenerative medicine, and accelerate the use of cell sheets in clinical applications.
三维（3D）组织可通过细胞片技术进行重建，并且已经启动了各种使用这些构建组织的临床研究以再生受损组织。虽然通过堆叠细胞片可以轻松制造3D组织，但细胞片与培养皿或双层细胞片之间细胞黏附的附着期通常需要20 - 30分钟。本研究提出了一种使用细胞片技术和离心法的生物工程组织快速制造系统。从温度响应培养皿收获的C2C12小鼠成肌细胞片在37°C孵育20分钟后会紧密附着于培养皿或另一细胞片。然而，同样的细胞片在37°C下离心（12 - 34×g）3分钟后，仅孵育3分钟也会紧密附着于培养皿或另一细胞片。通过离心，操作时间减少了约三分之二。快速附着情况也通过光学相干断层扫描进行了横截面确认。这些使用离心法快速构建的细胞片组织表现出活跃的细胞代谢、细胞活力以及与传统方法制备的组织一样非常高的血管内皮生长因子产生量；表明细胞片完全附着且没有任何细胞损伤。这种新系统将成为基于细胞片的组织工程和再生医学领域的有力工具，并加速细胞片在临床应用中的使用。
