Kanie Kei, Sakai Teppei, Imai Yuta, Yoshida Kei, Sugimoto Ayako, Makino Hodaka, Kubo Hirotsugu, Kato Ryuji
Department of Basic Medicinal Sciences, Graduate School of Pharmaceutical Sciences, Nagoya University, Furocho, Chikusa-ku, Nagoya, 464-8601, Japan.
Ogino Memorial Laboratory, Nihon Kohden Corporation, TWIns (Tokyo Women's Medical University-Waseda University Joint Institution for Advanced Biomedical Sciences), 8-1, Kawata-cho, Shinjyuku-ku, Tokyo, 162-8666, Japan.
Regen Ther. 2019 Jun 6;12:27-35. doi: 10.1016/j.reth.2019.05.002. eCollection 2019 Dec 15.
The development of induced pluripotent stem cell (iPSC) techniques has solved various limitations in cell culture including cellular proliferation and potency. Hence, the expectations on wider applications and the quality of manufactured iPSCs are rapidly increasing. To answer such growing expectations, enhancement of technologies to improve cell-manufacturing efficiency is now a challenge for the bioengineering field. Mechanization of conventional manual operations, aimed at automation of cell manufacturing, is quickly advancing. However, as more processes are being automated in cell manufacturing, it is need to be more critical about influential parameters that may not be as important in manual operations. As a model of such parameters, we focused on the effect of mechanical vibration, which transmits through the vessel to the cultured iPSCs. We designed 7 types of vertical vibration conditions in cell culture vessels using a vibration calibrator. These conditions cover a wide range of potential situations in cell culture, such as tapping or closing an incubator door, and examined their effects by continuous passaging (P3 to P5). Detailed evaluation of cells by time-course image analysis revealed that vibrations can enhance cell growth as an early effect but can negatively affect cell adhesion and growth profile after several passages as a delayed effect. Such unexpected reductions in cell quality are potentially critical issues in maintaining consistency in cell manufacturing. Therefore, this work reveals the importance of continuous examination across several passages with detailed, temporal, quantitative measurements obtained by non-invasive image analysis to examine when and how the unknown parameters will affect the cell culture processes.
诱导多能干细胞（iPSC）技术的发展解决了细胞培养中的各种限制，包括细胞增殖和潜能。因此，对iPSC更广泛应用和制造质量的期望正在迅速增加。为了回应这种不断增长的期望，提高细胞制造效率的技术提升现在是生物工程领域面临的一项挑战。旨在实现细胞制造自动化的传统手动操作的机械化正在迅速推进。然而，随着细胞制造中越来越多的过程实现自动化，需要更加关注在手动操作中可能不那么重要的影响参数。作为这类参数的一个模型，我们关注机械振动的影响，这种振动会通过容器传递给培养的iPSC。我们使用振动校准器在细胞培养容器中设计了7种垂直振动条件。这些条件涵盖了细胞培养中广泛的潜在情况，如轻拍或关闭培养箱门，并通过连续传代（P3至P5）来研究它们的影响。通过时间进程图像分析对细胞进行详细评估发现，振动作为早期效应可以促进细胞生长，但作为延迟效应，在几次传代后会对细胞黏附和生长情况产生负面影响。这种细胞质量的意外下降可能是维持细胞制造一致性的关键问题。因此，这项工作揭示了通过非侵入性图像分析获得详细的、随时间的、定量测量结果，对多次传代进行持续检查的重要性，以研究未知参数何时以及如何影响细胞培养过程。
