Vriezen N, Romein B, Luyben K C, van Dijken J P
Department of Microbiology and Enzymology, Kluyver Laboratory for Biotechnology, Delft University of Technology, Julianalaan 67, 2628BC Delft, The Netherlands.
Biotechnol Bioeng. 1997 May 5;54(3):272-86. doi: 10.1002/(SICI)1097-0290(19970505)54:3<272::AID-BIT8>3.0.CO;2-C.
Glutamine is a major source of energy, carbon, and nitrogen for mammalian cells. The amount of glutamine present in commercial mammalian cell media is, however, not necessarily balanced with cell requirements. Therefore, the effects of glutamine limitation on the physiology of two mammalian cell lines were studied in steady-state chemostat cultures fed with IMDM medium with 5% serum. The cell lines used were MN12, a mouse-mouse hybridoma, and SP2/0-Ag14, a mouse myeloma often used in hybridoma fusions. Cultures, grown at a fixed dilution rate of 0.03 h(-1), were fed with media containing glutamine concentrations ranging from 0.5 to 4 mmol L(-1). Biomass dry weight and cell number were linearly proportional to the glutamine concentrations fed, between 0.5 and 2 mmol L(-1), and glutamine was completely consumed by both cell lines. From this it was concluded that glutamine was the growth-limiting substrate in this concentration range and that the standard formulation of IMDM medium contains a twofold excess of glutamine. In glutamine-limited cultures, the specific rates of ammonia and alanine production were low compared to glutamine-excess cultures containing 4 mmol L(-1) glutamine in the feed medium. The specific consumption rates of nearly all amino acids decreased with increasing glutamine feed, indicating that, in their metabolic function, they may partially be replaced by glutamine. Both cell lines reacted similarly to differences in glutamine feeding in all aspects investigated, except for glucose metabolism, In SP2/0-Ag14 glutamine feed concentrations did not affect the specific glucose consumption, whereas in MN12 this parameter increased with increasing amounts of glutamine fed. This systematic study using controlled culture conditions together with a detailed analysis of culture data shows that, although cells may react similarly in many aspects, cell-line-specific characteristics may be encountered even with respect to fundamental physiological responses like the interaction of the glutamine and glucose metabolism.
谷氨酰胺是哺乳动物细胞能量、碳和氮的主要来源。然而，商业哺乳动物细胞培养基中谷氨酰胺的含量不一定与细胞需求相平衡。因此，在添加5%血清的IMDM培养基的稳态恒化器培养中，研究了谷氨酰胺限制对两种哺乳动物细胞系生理的影响。所用的细胞系是MN12（一种小鼠-小鼠杂交瘤）和SP2/0-Ag14（一种常用于杂交瘤融合的小鼠骨髓瘤）。以固定稀释率0.03 h(-1)培养的细胞，用含有0.5至4 mmol L(-1)谷氨酰胺浓度的培养基进行投喂。在0.5至2 mmol L(-1)之间，生物量干重和细胞数量与投喂的谷氨酰胺浓度呈线性比例关系，且两种细胞系都将谷氨酰胺完全消耗。由此得出结论，在该浓度范围内谷氨酰胺是生长限制底物，且IMDM培养基的标准配方中谷氨酰胺含量过量两倍。在谷氨酰胺限制的培养中，与在投喂培养基中含有4 mmol L(-1)谷氨酰胺的谷氨酰胺过量培养相比，氨和丙氨酸的比生产速率较低。几乎所有氨基酸的比消耗速率都随着谷氨酰胺投喂量的增加而降低，这表明在其代谢功能中，它们可能部分被谷氨酰胺替代。除了葡萄糖代谢外，在所有研究的方面，两种细胞系对谷氨酰胺投喂差异的反应相似。在SP2/0-Ag14中，谷氨酰胺投喂浓度不影响比葡萄糖消耗，而在MN12中，该参数随着投喂谷氨酰胺量的增加而增加。这项使用受控培养条件并结合培养数据详细分析的系统研究表明，尽管细胞在许多方面可能反应相似，但即使在谷氨酰胺和葡萄糖代谢相互作用等基本生理反应方面，也可能存在细胞系特异性特征。
