Kato Yuma, Kim Mee-Hae, Kino-Oka Masahiro
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
J Biosci Bioeng. 2018 Jun;125(6):736-740. doi: 10.1016/j.jbiosc.2018.01.002. Epub 2018 Feb 2.
Understanding the fundamental mechanisms that govern the growth kinetics of human induced pluripotent stem cells (hiPSCs) contributes to culture design strategies to improve large-scale production. Two hiPSC lines (Tic and 253G1) were cultured under static and dynamic suspension conditions, and growth kinetics were compared during early (24-48 h), middle (48-72 h), and late (72-96 h) stages. In 2D static culture, similar growth profiles were observed for both hiPSC lines. However, there were significant differences in growth profile patterns and aggregate morphologies between hiPSC lines grown in 3D static and dynamic cultures. Based on immunostaining comparing the two hiPSC lines, surface distribution of collagen type I was observed in aggregates of the Tic line, but not in those of the 253G1 line. Compared to that in 3D static culture, the numbers of cells at 96 h were significantly decreased in 3D dynamic culture. The apparent specific growth rate (μ) of the Tic line was maintained continuously throughout culture, whereas that of the 253G1 line decreased gradually with culture until the late phase, at which time this parameter was reduced to μ = (0.85 ± 0.71) × 10 h. This indicates that during the growth of hiPSCs in 3D dynamic culture, cells were damaged by liquid flow, which disrupted the cell-synthesized extracellular matrix (ECM). These results demonstrate that cell-synthesized ECM is an important factor affecting cell growth and morphology, and that changes to the ECM within aggregates lead to reduced growth abilities in dynamic culture.
了解调控人诱导多能干细胞（hiPSC）生长动力学的基本机制有助于制定优化大规模生产的培养设计策略。将两条hiPSC系（Tic和253G1）分别在静态和动态悬浮条件下培养，并比较其在早期（24 - 48小时）、中期（48 - 72小时）和晚期（72 - 96小时）的生长动力学。在二维静态培养中，两条hiPSC系均观察到相似的生长曲线。然而，在三维静态和动态培养中生长的hiPSC系之间，其生长曲线模式和聚集体形态存在显著差异。基于对两条hiPSC系的免疫染色比较，在Tic系的聚集体中观察到I型胶原的表面分布，而在253G1系的聚集体中未观察到。与三维静态培养相比，三维动态培养中96小时时的细胞数量显著减少。Tic系的表观比生长速率（μ）在整个培养过程中持续保持，而253G1系的表观比生长速率在培养后期逐渐下降，此时该参数降至μ = (0.85 ± 0.71) × 10 h⁻¹。这表明在三维动态培养中hiPSC生长期间，细胞受到液流损伤，从而破坏了细胞合成的细胞外基质（ECM）。这些结果表明，细胞合成的ECM是影响细胞生长和形态的重要因素，并且聚集体内ECM的变化导致动态培养中生长能力降低。
