Cell density effect in the baculovirus-insect cells system: A quantitative analysis of energetic metabolism

• Published in: Biotechnology and bioengineering Vol. 104; no. 1; pp. 162 - 180
• Main Authors: Bernal, Vicente, Carinhas, Nuno, Yokomizo, Adriana Y, Carrondo, Manuel J.T, Alves, Paula M
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.09.2009; Wiley; Wiley Subscription Services, Inc
• Subjects: Amino acids; Animals; Baculoviridae - growth & development; Baculovirus; baculovirus production; Biological and medical sciences; Biotechnology; Cell Count; Cell culture; cell density effect; Cell Line; Cells; Energy Metabolism; Fundamental and applied biological sciences. Psychology; metabolic burden; metabolic flux analysis; Metabolic Networks and Pathways; Metabolism; protein production; Spodoptera; Spodoptera frugiperda; Viruses; Energetics; Insecta; Spodoptera frugiperda; Metabolism; Density; Protein; Virus; cell density effect; Baculoviridae; Arthropoda; Metabolic flux analysis; Lepidoptera; Production; Noctuidae; Invertebrata; metabolic burden; baculovirus production; Quantitative analysis; protein production
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.22364

The cell density effect (i.e., the drop in the specific productivity in the baculovirus-insect cells expression system when cells are infected at high cell densities) has been extensively described in the literature. In this article, a model for the central metabolism of serum-free suspension cultures of Spodoptera frugiperda Sf9 cells is proposed and used to investigate the metabolic basis for this phenomenon. The main metabolic pathways (glycolysis, pentose phosphate pathway, tricarboxylic acids cycle, glutaminolysis, and amino acids metabolism), cellular growth and energetics were considered. The analysis of the stoichiometric model allowed further understanding of the interplay of the consumption of carbon and nitrogen sources in insect cells. Moreover, metabolic flux analysis revealed that Sf9 cells undergo a progressive inhibition of central metabolism when grown to high cell densities, for which the incorporation of amino acids carbon backbones into the TCA cycle (mainly glutamine) and the down-regulation of glycolysis are partially responsible. Following infection by baculovirus and cellular division arrest, central energy metabolism depended on the infection strategy chosen (cell concentration at the moment of infection and multiplicity of infection), inhibition being observed at high cell densities. Interestingly, the energetic status of the culture correlated with the decrease in cellular production of baculovirus, meaning that there is room for process optimization through the application of metabolic engineering techniques. Biotechnol. Bioeng. 2009; 104: 162-180