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通过焓平衡验证的化学计量生长方程检测培养动物细胞不断变化的底物需求。

Detection of the changing substrate requirements of cultured animal cells by stoichiometric growth equations validated by enthalpy balances.

作者信息

Guan Y H, Kemp R B

机构信息

Institute of Biological Sciences, University of Wales, Aberystwyth, UK.

出版信息

J Biotechnol. 1999 Apr 15;69(2-3):95-114. doi: 10.1016/s0168-1656(99)00007-3.

Abstract

As part of an overall aim to base the feeding of substrates to cultured animal cells on their actual metabolic needs, we have developed a stoichiometric approach centred on the macronutrients in the medium. Heat flux records the overall metabolic activity and therefore was the sensitive indicator of changing metabolic requirement. Analyses were made of the experimental measurements on two engineered cell lines in batch culture, the 2C11-12 macrophage hybridoma cell capable of the respiratory burst and the CHO320 constitutively producing human interferon-gamma. The crux was to construct simplified stoichiometric equations for the growth reactions to represent metabolic activity as it changed with time. Beforehand, it was essential to select the appropriate components for the equations. The choice was then justified by constructing enthalpy balances in which the ratio of heat flux to enthalpy flux must be close to unity for validation. By combining the stoichiometric approach with heat flow measurements, it was shown both theoretically and experimentally that the set of stoichiometric coefficients constituting a validated growth equation has a one-to-one corresponding relationship to the metabolic activity of the average cell population. Thus, a strategy was established for feeding the cells at any one time with the correct ratio of the major substrates, glucose and glutamine, in response to metabolic requirements that change with time.

摘要

作为根据培养动物细胞的实际代谢需求提供底物这一总体目标的一部分,我们开发了一种以培养基中的常量营养素为中心的化学计量方法。热通量记录了总体代谢活性,因此是代谢需求变化的敏感指标。对分批培养的两种工程细胞系进行了实验测量分析,一种是能够产生呼吸爆发的2C11 - 12巨噬细胞杂交瘤细胞,另一种是组成型产生人干扰素 - γ的CHO320细胞。关键在于构建生长反应的简化化学计量方程,以表示代谢活性随时间的变化。在此之前,为方程选择合适的组分至关重要。然后通过构建焓平衡来证明这种选择的合理性,在焓平衡中,热通量与焓通量的比值必须接近1才能得到验证。通过将化学计量方法与热流测量相结合,从理论和实验上都表明,构成有效生长方程一组化学计量系数与平均细胞群体的代谢活性具有一一对应的关系。因此,建立了一种策略,可根据随时间变化的代谢需求,在任何时刻以正确的比例向细胞提供主要底物葡萄糖和谷氨酰胺。

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