Human 293 cell metabolism in low glutamine-supplied culture: interpretation of metabolic changes through metabolic flux analysis
| Autor: | Amine Kamen, J. Sabatié, Michel Perrier, M. Koehl, I. Nadeau |
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| Rok vydání: | 2000 |
| Předmět: |
Glutamine
Citric Acid Cycle Cell Culture Techniques Bioengineering Cell Count Biology Carbohydrate metabolism Kidney Applied Microbiology and Biotechnology Models Biological Cell Line Ammonia production Ammonia chemistry.chemical_compound Bioreactors Metabolic flux analysis Humans Lactic Acid Amino Acids Essential amino acid chemistry.chemical_classification Kidney metabolism Amino acid Culture Media Kinetics Glucose chemistry Biochemistry Cell Division Biotechnology |
| Zdroj: | Metabolic engineering. 2(4) |
| ISSN: | 1096-7176 |
| Popis: | Metabolic flux analysis is a useful tool to analyze cell metabolism. In this study, we report the use of a metabolic model with 34 fluxes to study the 293 cell, in order to improve its growth capacity in a DMEM/F12 medium. A batch, fed-batch with glutamine feeding, fed-batch with essential amino acids, and finally a fed-batch experiment with both essential and nonessential amino acids were compared. The fed-batch with glutamine led to a maximum cell density of 2.4x10(6) cells/ml compared to 1.8x10(6) cells/ml achieved in a batch mode. In this fed-batch with glutamine, it was also found that 2.5 mM ammonia was produced compared to the batch which had a final ammonia concentration of 1 mM. Ammonia was found to be growth inhibiting for this cell line at a concentration starting at 1 mM. During the fed-batch with glutamine, the flux analysis shows that a majority of amino acid fluxes and Kreb's cycle fluxes, except for glutamine flux, are decreased. This observation led to the conclusion that the main nutrient used is glutamine and that during the batch there is an overflow in the Kreb's cycle. Thus, a fed-batch with glutamine permits a better utilization of this nutrient. A fed-batch with essential amino acid without glutamine was also assayed in order to reduce ammonia production. The maximum cell density was increased further to 3x10(6) cells/ml and ammonia production was reduced below 1 mM. Flux analysis shows that the cells could adapt to a medium with low glutamine by increasing the amino acid fluxes toward the Kreb's cycle. Adding nonessential amino acids during this feeding strategy did not improve growth further and the nonessential amino acids accumulated in the medium. |
| Databáze: | OpenAIRE |
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