Estimation of the intracellular fluxes for a hybridoma cell line by material balances

Paredes, C.; Sanfeliu, Anna; Cárdenas, F.; Cairó, Jordi J.; Gòdia, Francesc

doi:10.1016/s0141-0229(98)00023-4

Cited by 25 publications

(32 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This possibly resulted in the increased flow of pyruvate flux towards the lactate production. This continuous regeneration of NADþ has been reported in other hybridoma cell lines (Paredes et al, 1998;. All these observations indicate that the pyruvate node may play an essential role in metabolic shifting between oxidative metabolism with high-energy production and increased substrate utilization leading to lactate overproduction through the maintenance of NADþ/NADH ratio.…”

Section: Glycolysissupporting

confidence: 73%

“…Recently, cybernetic modeling approach was combined with the MFA technique to elucidate multiple cell physiological states (Namjoshi et al, 2003). The changes in intracellular metabolic fluxes in response to the environmental variations have been further unveiled by combined MFA and in vitro 13 C NMR studies (Paredes et al, 1998;. For example, Paredes et al (1998) revealed that increase in lactate production was mainly due to the reduced activity of malate-aspartate shuttle in the cell metabolism.…”

Section: Introductionmentioning

confidence: 99%

“…The changes in intracellular metabolic fluxes in response to the environmental variations have been further unveiled by combined MFA and in vitro 13 C NMR studies (Paredes et al, 1998;. For example, Paredes et al (1998) revealed that increase in lactate production was mainly due to the reduced activity of malate-aspartate shuttle in the cell metabolism. The lumped reaction models have also been applied to designing the culture medium and feeding strategies (Xie and Wang, 1994a,b).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Elucidation of metabolism in hybridoma cells grown in fed‐batch culture by genome‐scale modeling

Selvarasu

Wong

Karimi

et al. 2008

Biotech & Bioengineering

View full text Add to dashboard Cite

Genome-scale modeling of mouse hybridoma cells producing monoclonal antibodies (mAb) was performed to elucidate their physiological and metabolic states during fed-batch cell culture. Initially, feed media nutrients were monitored to identify key components among carbon sources and amino acids with significant impact on the desired outcome, for example, cell growth and antibody production. The monitored profiles indicated rapid assimilation of glucose and glutamine during the exponential growth phase. Significant increase in mAb concentration was also observed when glutamine concentration was controlled at 0.5 mM as a feeding strategy. Based on the reconstructed genome-scale metabolic network of mouse hybridoma cells and fed-batch profiles, flux analysis was then implemented to investigate the cellular behavior and changes in internal fluxes during the cell culture. The simulated profile of the cell growth was consistent with experimentally measured specific growth rate. The in silico simulation results indicated (i) predominant utilization of glycolytic pathway for ATP production, (ii) importance of pyruvate node in metabolic shifting, and (iii) characteristic pattern in lactate to glucose ratio during the exponential phase. In future, experimental and in silico analyses can serve as a promising approach to identifying optimal feeding strategies and potential cell engineering targets as well as facilitate media optimization for the enhanced production of mAb or recombinant proteins in mammalian cells.

show abstract

Section: Glycolysissupporting

confidence: 73%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Elucidation of metabolism in hybridoma cells grown in fed‐batch culture by genome‐scale modeling

Selvarasu

Wong

Karimi

et al. 2008

Biotech & Bioengineering

View full text Add to dashboard Cite

show abstract

“…The model was developed from a model discussed in a previous article (Nadeau et al, 2000a and b) and used concepts described in reports from Bonarius et al (1996Bonarius et al ( , 1997Bonarius et al ( , 1998Nyberg et al (1999); Paredes et al (1998); Pissarra and Henriksen (1998); Savinell and Palsson (1992a, b, c); Schmidt et al (1998); Stephanopoulos (1998); Takiguchi et al (1997); Vanrolleghem and Heijnen (1998); Varma and Palsson (1994); Xie and Wang (1996a and b); and Zupke and Stephanopoulos (1995). Glycolysis, glutaminolysis, amino acid degradation, TCA cycle, protein production and internalization were considered as well as the pentose-phosphate cycle and compartmentalization of certain nutrients.…”

Section: Metabolic Modelmentioning

confidence: 99%

Low‐protein medium affects the 293SF central metabolism during growth and infection with adenovirus

Nadeau

Gilbert²,

Jacob³

et al. 2001

Biotech & Bioengineering

View full text Add to dashboard Cite

In this study the metabolism of 293SF cells grown in serum-free and low-protein medium was analyzed. This cell line is known for its ability to replicate recombinant adenovirus, mainly used in gene therapy applications. A complete model composed of the main glycolytic, glutaminolytic, and amino acids pathways, as well as the internalization fluxes of certain compounds into the mitochondria, is used for metabolic flux calculations. The pentose-phosphate cycle is also added to the biochemical reactions set and was independently measured with labeled 14C-glucose. Different feeding strategies in two different media were analyzed with the model, and the theoretical ATP production was also calculated. The two media were similar in their glucose and amino acid composition, but one contained BSA at 1g/L whereas the other had a very low protein content. Use of low-protein medium resulted in up to fourfold higher adenoviral vector production. In this medium, glucose utilization was more efficient, as it entered the TCA cycle more efficiently. Also, lower glutamine and amino acids consumption were observed as well as lower lactate and ammonia production. This increased TCA activity led to a twofold higher ATP production in the low-protein medium.

show abstract

“…When there is sufficient Gln in the medium, Gln metabolism proceeds mainly through the a-ketoglutarate and alanine pathway. When Gln is restricted, Gln metabolism begins to switch to generate a-ketoglutaric acid and the ammonia pathway, since the ammonia pathway produces more ATP, thereby alleviating the decline in energy substrates (Kuwae and Ohda 2005;Paredes et al 1998). The co-consumption of lactate and Ala after 250 h of cultivation suggests that the decrease in residual glucose concentration caused a limited availability of pyruvate from the glycolytic pathway and then led to the simultaneous consumption of lactate and Ala. Lactate and Ala can be metabolized via pyruvate as a source of energy to yield ATP.…”

Section: Discussionmentioning

confidence: 99%

Bioprocess development for the production of mouse-human chimeric anti-epidermal growth factor receptor vIII antibody C12 by suspension culture of recombinant Chinese hamster ovary cells

Deng

Wang

et al. 2011

Cytotechnology

View full text Add to dashboard Cite

The mouse-human chimeric anti-epidermal growth factor receptor vIII (EGFRvIII) antibody C12 is a promising candidate for the diagnosis of hepatocellular carcinoma (HCC). In this study, 3 processes were successfully developed to produce C12 by cultivation of recombinant Chinese hamster ovary (CHO-DG44) cells in serum-free medium. The effect of inoculum density was evaluated in batch cultures of shaker flasks to obtain the optimal inoculum density of 5 9 10 5 cells/mL. Then, the basic metabolic characteristics of CHO-C12 cells were studied in stirred bioreactor batch cultures. The results showed that the limiting concentrations of glucose and glutamine were 6 and 1 mM, respectively. The culture process consumed significant amounts of aspartate, glutamate, asparagine, serine, isoleucine, leucine, and lysine. Aspartate, glutamate, asparagine, and serine were particularly exhausted in the early growth stage, thus limiting cell growth and antibody synthesis. Based on these findings, fedbatch and perfusion processes in the bioreactor were successfully developed with a balanced amino acid feed strategy. Fed-batch and especially perfusion culture effectively maintained high cell viability to prolong the culture process. Furthermore, perfusion cultures maximized the efficiency of nutrient utilization; the mean yield coefficient of antibody to consumed glucose was 44.72 mg/g and the mean yield coefficient of glutamine to antibody was 721.40 mg/g. Finally, in small-scale bioreactor culture, the highest total amount of C12 antibody (1,854 mg) was realized in perfusion cultures. Therefore, perfusion culture appears to be the optimal process for small-scale production of C12 antibody by rCHO-C12 cells.

show abstract

Estimation of the intracellular fluxes for a hybridoma cell line by material balances

Cited by 25 publications

References 24 publications

Elucidation of metabolism in hybridoma cells grown in fed‐batch culture by genome‐scale modeling

Elucidation of metabolism in hybridoma cells grown in fed‐batch culture by genome‐scale modeling

Low‐protein medium affects the 293SF central metabolism during growth and infection with adenovirus

Bioprocess development for the production of mouse-human chimeric anti-epidermal growth factor receptor vIII antibody C12 by suspension culture of recombinant Chinese hamster ovary cells

Contact Info

Product

Resources

About