Department of Biotechnology, Graduate School of Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, 464-8603, Japan.
Cytotechnology. 2000 Jul;33(1-3):147-55. doi: 10.1023/A:1008189802014.
We investigated the effect of cell adhesion on cellgrowth and productivity of recombinant protein inChinese hamster ovary (CHO) cells. Cells cultured innormal tissue culture dishes attached to the dishsurfaces and grew as a monolayer, while cells culturedin non-treated dishes proliferated in suspension assingle cells without adhering to the dish surfaces. On an agarose-coated dish surface, cell aggregatesformed without attaching to the dish. Growth rates inboth suspension cultures were slightly lower thanthose in monolayer culture. Cell cycle analysisindicated that the duration of the G(1) phase insuspension cultures was longer than that in monolayerculture, suggesting that attachment to the substratummainly affected the transition from the G(1) to theS phase. Consistent with this, CDK inhibitor p27,that inhibits the G(1)S transition, was induced inthe cells cultured in suspension.To assess the productivity of recombinant proteins,CHO cells were transfected with a plasmid containingmurine interferon gamma (mIFN-gamma) under thecontrol of the cytomegalovirus promoter. Insuspension culture, mIFN-gamma productivity wasslightly lower than that in the monolayer culture. When protein kinase C was activated by phorbol ester,mIFN-gamma production was enhanced in both themonolayer and suspension cultures. However, theproductivity in the suspension culture was lower thanthat in the adherent culture even in the presence ofhigh concentrations of phorbol ester. These resultssuggested that cell adhesion to the substratum affectsvarious features of CHO cells.
我们研究了细胞黏附对中国仓鼠卵巢（CHO）细胞中重组蛋白的细胞生长和生产效率的影响。在普通组织培养皿中培养的细胞黏附在皿表面并单层生长，而在未经处理的培养皿中培养的细胞在悬浮液中增殖，作为单个细胞而不黏附在皿表面。在琼脂糖涂层的皿表面上，细胞聚集体形成而不黏附在皿上。两种悬浮培养物的生长速度均略低于单层培养物。细胞周期分析表明，悬浮培养物中 G1 期的持续时间长于单层培养物，表明附着在基质上主要影响从 G1 期到 S 期的转变。与此一致，CDK 抑制剂 p27 诱导了悬浮培养物中的细胞，该抑制剂抑制 G1-S 期的转变。为了评估重组蛋白的生产效率，将含有鼠干扰素γ（mIFN-γ）的质粒转染到受巨细胞病毒启动子控制的 CHO 细胞中。在悬浮培养中，mIFN-γ的生产效率略低于单层培养。当蛋白激酶 C 通过佛波酯激活时，mIFN-γ的产生在单层和悬浮培养中均增强。然而，即使存在高浓度的佛波酯，悬浮培养物的生产效率也低于附着培养物。这些结果表明，细胞黏附到基质上影响 CHO 细胞的各种特征。
