Automatic fabrication of 3-dimensional tissues using cell sheet manipulator technique
Automatic fabrication of 3-dimensional tissues using cell sheet manipulator technique
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DOI：
10.1016/j.biomaterials.2013.12.014
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发表时间：
2014-03-01
期刊：
影响因子：
14
通讯作者：
Okano, Teruo
中科院分区：
文献类型：
--
作者：
Kikuchi, Tetsutaro;Shimizu, Tatsuya;Okano, Teruo
自动化制造是组织工程治疗产品变得通用和经济的关键。在这里，我们开发了一种自动细胞片堆叠装置，利用我们的细胞片操纵器技术制造三维组织工程构建体，其中从温度响应培养皿收获的细胞片堆叠成多层细胞片。通过优化的堆叠条件和细胞接种条件，该装置最终能够重复地使五层的人骨骼肌成肌细胞（HSMM）片的厚度约为70-80 μ m在100分钟内。组织切片和共聚焦形貌的五层HSMM片显示分层结构，没有分层。在使用胰蛋白酶消化的细胞计数中，一层、三层和五层HSMM片材中的活细胞数与堆叠过程后1 h的接种细胞数相当;然而，在随后的5天静态培养后，五层HSMM片材的活细胞数略有下降，而一层和三层HSMM片材保持其活细胞数。这表明在静态培养中保持组织存在厚度限制。我们的结论是，通过结合我们的细胞片操纵器技术和工业机器人技术，我们可以创建一个安全，具有成本效益的制造系统，能够从细胞片生产组织工程产品。(C)2013爱思唯尔有限公司保留所有权利。
Automated manufacturing is a key for tissue-engineered therapeutic products to become common-place and economical. Here, we developed an automatic cell sheet stacking apparatus to fabricate 3-dimensional tissue-engineered constructs exploiting our cell sheet manipulator technique, where cell sheets harvested from temperature-responsive culture dishes are stacked into a multilayered cell sheet. By optimizing the stacking conditions and cell seeding conditions, the apparatus was eventually capable of reproducibly making five-layer human skeletal muscle myoblast (HSMM) sheets with a thickness of approximately 70-80 mu m within 100 min. Histological sections and confocal topographies of the five-layer HSMM sheets revealed a stratified structure with no delamination. In cell counts using trypsinization, the live cell numbers in one-, three- and five-layer HSMM sheets were equivalent to the seeded cell numbers at 1 h after the stacking processes; however, after subsequent 5-day static cultures, the live cell numbers of the five-layered HSMM sheets decreased slightly, while one- and three-layer HSMM sheets maintained their live cell numbers. This suggests that there are thickness limitations in maintaining tissues in a static culture. We concluded that by combining our cell sheet manipulator technique and industrial robot technology we can create a secure, cost-effective manufacturing system able to produce tissue-engineered products from cell sheets. (C) 2013 Elsevier Ltd. All rights reserved.