Metabolism of PER.C6™ cells cultivated under fed-batch conditions at low glucose and glutamine levels

• Published in: Biotechnology and bioengineering Vol. 94; no. 1; pp. 139 - 150
• Main Authors: MARANGA, Luis, GOOCHEE, Charles F
• Format: Journal Article
• Language: English
• Published: New York, NY Wiley 05.05.2006
• Subjects: Adenoviridae - genetics; Amino Acids - metabolism; Ammonia - metabolism; Biological and medical sciences; Bioreactors; Biotechnology; Cell Count; Cell Culture Techniques; Cell Line, Transformed; Cell Survival; Cell Transformation, Viral; Culture Media - chemistry; Culture Media, Serum-Free; Energy Metabolism; Fundamental and applied biological sciences. Psychology; Glucose - metabolism; Glutamine - metabolism; Humans; Lactic Acid - metabolism; Oxygen - metabolism; Retina - cytology; Retina - embryology; Retina - growth & development; Retina - metabolism; Retina - virology; Time Factors; Human; Cell culture; Cell line; fed-batch; metabolic shift; PER.C6# cells; Glucose; Metabolism; Semicontinuous; Glutamine
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.20890

This is the first study to examine PER.C6 cell glucose/energy and glutamine metabolism with fed-batch cultures at controlled low glutamine, low glucose, and simultaneous low glucose and low glutamine levels. PER.C6(TM) cell metabolism was investigated in serum-free suspension bioreactors at two-liter scale. Control of glucose and/or glutamine concentrations had a significant effect on cellular metabolism leading to an increased efficiency of nutrient utilization, altered byproduct synthesis, while having no effect on cell growth rate. Cultivating cells at a controlled glutamine concentration of 0.25 mM reduced q(Gln) and q(NH(4)(+)) by approximately 30%, q(Ala) 85%, and q(NEAA) 50%. The fed-batch control of glutamine also reduced the overall accumulation of ammonium ion by approximately 50% by minimizing the spontaneous chemical degradation of glutamine. No major impact upon glucose/energy metabolism was observed. Cultivating cells at a glucose concentration of 0.5 mM reduced q(Glc) about 50% and eliminated lactate accumulation. Cells exhibited a fully oxidative metabolism with Y(O(2)/Glc) of approximately 6 mol/mol. However, despite no increase in q(Gln), an increased ammonium ion accumulation and Y(NH(4)(+)/Gln) were also observed. Effective control of lactate and ammonium ion accumulation by PER.C6 cells was achieved using fed-batch with simultaneously controlled glucose and glutamine. A fully oxidative glucose metabolism and a complete elimination of lactate production were obtained. The q(Gln) value was again reduced and, despite an increased q(NH(4)(+)) compared with batch culture, ammonium ion levels were typically lower than corresponding ones in batch cultures, and the accumulation of non-essential amino acids (NEAA) was reduced about 50%. In conclusion, this study shows that PER.C6 cell metabolism can be confined to a state with improved efficiencies of nutrient utilization by cultivating cells in fed-batch at millimolar controlled levels of glucose and glutamine. In addition, PER.C6 cells fall into a minority category of mammalian cell lines for which glutamine plays a minor role in energy metabolism.