Department of Mechanical Engineering, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, TWIns, 8-1 Kawada-cho, Shinjuku-ku, Tokyo, Japan.
Sci Rep. 2020 Jun 11;10(1):9468. doi: 10.1038/s41598-020-66375-1.
Proteinases that digest the extracellular matrix are usually used to harvest cells from culture vessels in a general culture process, which lowers the initial adhesion rate in regenerative medicine. Cell sheet engineering is one of the most important technologies in this field, especially for transplantation, because fabricated cell sheets have rich extracellular matrixes providing strong initial adhesion. Current cell sheet fabrication relies on temperature-responsive polymer-coated dishes. Cells are cultured on such specialized dishes and subjected to low temperature. Thus, we developed a simple but versatile cell sheet fabrication method using ubiquitous culture dishes/flasks without any coating or temperature modulation. Confluent mouse myoblasts (C2C12 cell line) were exposed to ultrasonic vibration from underneath and detached as cell sheets from entire culture surfaces. Because of the absence of low temperature, cell metabolism was statically increased compared with the conventional method. Furthermore, viability, morphology, protein expression, and mRNA expression were normal. These analyses indicated no side effects of ultrasonic vibration exposure. Therefore, this novel method may become the standard for cell sheet fabrication. Our method can be easily conducted following a general culture procedure with a typical dish/flask, making cell sheets more accessible to medical experts.
在一般的培养过程中，通常使用消化细胞外基质的蛋白酶从培养容器中收获细胞，这会降低再生医学中细胞的初始黏附率。细胞片工程是该领域最重要的技术之一，特别是对于移植而言，因为所制备的细胞片具有丰富的细胞外基质，提供了强大的初始黏附力。目前的细胞片制备依赖于对温度响应的聚合物涂层培养皿。细胞在这种特殊的培养皿上培养并在低温下处理。因此，我们开发了一种简单但通用的细胞片制备方法，使用无处不在的培养皿/培养瓶，无需任何涂层或温度调节。将汇合的小鼠成肌细胞（C2C12 细胞系）暴露在底部的超声振动下，并从整个培养表面作为细胞片脱落。由于没有低温，与传统方法相比，细胞代谢处于静态增加状态。此外，细胞活力、形态、蛋白质表达和 mRNA 表达均正常。这些分析表明超声振动暴露没有副作用。因此，这种新方法可能成为细胞片制备的标准。我们的方法可以很容易地按照带有典型培养皿/培养瓶的一般培养程序进行，使细胞片更容易被医学专家获得。
