Tharmalingam T, Ghebeh H, Wuerz T, Butler M
Department of Microbiology, University of Manitoba, Manitoba, Winnipeg, Canada R3T 2N2.
Mol Biotechnol. 2008 Jun;39(2):167-77. doi: 10.1007/s12033-008-9045-8.
The addition of the non-ionic surfactant, Pluronic F-68, to serum-free CHO cultures causes multi-functional effects that enhance cell yield in agitated cultures and reduce cell adhesion in stationary cultures. Three independent CHO cell lines were subjected to high liquid shear in assay systems that either included or excluded a liquid-gas interface. In the absence of Pluronic, there was a loss in cell viability in either assay system, although there was an intrinsic variability in sensitivity of the cell lines to shear damage. Supplementation with Pluronic prevented loss of cell viability, indicating protection in either a gas sparged or bubble-free environment. However, we found no evidence of long-term protection of cells once Pluronic was removed. Pluronic was capable of repairing trypsin-damaged cells as evidenced by enhanced growth, reduced membrane porosity, and improved robustness under liquid shear. The proportion of adherent cells was reduced to a minimal level by the presence of Pluronic although its effect was rapidly reversible with a high proportion (70%) of adherent cells observed within a few culture passages of its removal. The observed effects of Pluronic on these cultures are compatible with a mechanism in which the polymer forms a protective layer on the cell membrane, which has a significantly lower hydrophobicity.
向无血清的中国仓鼠卵巢(CHO)细胞培养物中添加非离子表面活性剂普朗尼克F-68会产生多种功能效应,可提高搅拌培养中的细胞产量,并减少静置培养中的细胞黏附。在包含或不包含气液界面的检测系统中,对三种独立的CHO细胞系施加高液体剪切力。在没有普朗尼克的情况下,两种检测系统中的细胞活力均会丧失,尽管细胞系对剪切损伤的敏感性存在内在差异。添加普朗尼克可防止细胞活力丧失,表明在通气或无气泡环境中均有保护作用。然而,我们发现一旦去除普朗尼克,就没有证据表明细胞能得到长期保护。普朗尼克能够修复胰蛋白酶损伤的细胞,这表现为细胞生长增强、膜孔隙率降低以及在液体剪切下的稳健性提高。普朗尼克的存在使贴壁细胞比例降至最低水平,不过其效果是迅速可逆的,在去除普朗尼克后的几代培养中,观察到有高比例(70%)的贴壁细胞。观察到的普朗尼克对这些培养物的影响与一种机制相符,即该聚合物在细胞膜上形成一层疏水性显著较低的保护层。