The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line
One of the most important limitations of mammalian cells-based bioprocesses, and particularly hybridoma cell lines, is the accelerated metabolism related to glucose and glutamine consumption. The high uptake rates of glucose and glutamine (i.e., the main sources of carbon, nitrogen and energy) lead...
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MDPI AG
2021-07-01
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| Online Access: | https://www.mdpi.com/2076-3417/11/14/6258 |
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| author | Iván Martínez-Monge Pere Comas David Catalán-Tatjer Jordi Prat Antoni Casablancas Carlos Paredes Martí Lecina Jordi Joan Cairó |
| author_facet | Iván Martínez-Monge Pere Comas David Catalán-Tatjer Jordi Prat Antoni Casablancas Carlos Paredes Martí Lecina Jordi Joan Cairó |
| author_sort | Iván Martínez-Monge |
| collection | DOAJ |
| description | One of the most important limitations of mammalian cells-based bioprocesses, and particularly hybridoma cell lines, is the accelerated metabolism related to glucose and glutamine consumption. The high uptake rates of glucose and glutamine (i.e., the main sources of carbon, nitrogen and energy) lead to the production and accumulation of large amounts of lactate and ammonia in culture broth. Lactate and/or ammonia accumulation, together with the depletion of the main nutrients, are the major causes of apoptosis in hybridoma cell cultures. The KB26.5 hybridoma cell line, producing an IgG<sub>3</sub>, was engineered with BHRF1 (KB26.5-BHRF1), an Epstein–Barr virus-encoded early protein homologous to the antiapoptotic protein Bcl-2, with the aim of protecting the hybridoma cell line from apoptosis. Surprisingly, besides achieving effective protection from apoptosis, the expression of BHRF1 modified the metabolism of the hybridoma cell line. Cell physiology and metabolism analyses of the original KB26.5 and KB26.5-BHRF1 revealed an increase of cell growth rate, a reduction of glucose and glutamine consumption, as well as a decrease in lactate secretion in KB26.5-BHRF1 cells. A flux balance analysis allowed us to quantify the intracellular fluxes of both cell lines. The main metabolic differences were identified in glucose consumption and, consequently, the production of lactate. The lactate production flux was reduced by 60%, since the need for NADH regeneration in the cytoplasm decreased due to a more than 50% reduction in glucose uptake. In general terms, the BHRF1 engineered cell line showed a more efficient metabolism, with an increase in biomass volumetric productivity under identical culture conditions. |
| first_indexed | 2024-03-10T09:46:50Z |
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| institution | Directory Open Access Journal |
| issn | 2076-3417 |
| language | English |
| last_indexed | 2024-03-10T09:46:50Z |
| publishDate | 2021-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Applied Sciences |
| spelling | doaj.art-51ee68f70b814f5888d58b7287a218d82023-11-22T03:06:40ZengMDPI AGApplied Sciences2076-34172021-07-011114625810.3390/app11146258The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell LineIván Martínez-Monge0Pere Comas1David Catalán-Tatjer2Jordi Prat3Antoni Casablancas4Carlos Paredes5Martí Lecina6Jordi Joan Cairó7The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, DK-2800 Kongens Lyngby, DenmarkChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainThe Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, DK-2800 Kongens Lyngby, DenmarkChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainChemical, Biological and Environmental Engineering Department, Campus Bellatera, Universitat Autònoma de Barcelona, Edifici Q, Carretera de Cerdanyola, s/n., 08193 Bellaterra, SpainOne of the most important limitations of mammalian cells-based bioprocesses, and particularly hybridoma cell lines, is the accelerated metabolism related to glucose and glutamine consumption. The high uptake rates of glucose and glutamine (i.e., the main sources of carbon, nitrogen and energy) lead to the production and accumulation of large amounts of lactate and ammonia in culture broth. Lactate and/or ammonia accumulation, together with the depletion of the main nutrients, are the major causes of apoptosis in hybridoma cell cultures. The KB26.5 hybridoma cell line, producing an IgG<sub>3</sub>, was engineered with BHRF1 (KB26.5-BHRF1), an Epstein–Barr virus-encoded early protein homologous to the antiapoptotic protein Bcl-2, with the aim of protecting the hybridoma cell line from apoptosis. Surprisingly, besides achieving effective protection from apoptosis, the expression of BHRF1 modified the metabolism of the hybridoma cell line. Cell physiology and metabolism analyses of the original KB26.5 and KB26.5-BHRF1 revealed an increase of cell growth rate, a reduction of glucose and glutamine consumption, as well as a decrease in lactate secretion in KB26.5-BHRF1 cells. A flux balance analysis allowed us to quantify the intracellular fluxes of both cell lines. The main metabolic differences were identified in glucose consumption and, consequently, the production of lactate. The lactate production flux was reduced by 60%, since the need for NADH regeneration in the cytoplasm decreased due to a more than 50% reduction in glucose uptake. In general terms, the BHRF1 engineered cell line showed a more efficient metabolism, with an increase in biomass volumetric productivity under identical culture conditions.https://www.mdpi.com/2076-3417/11/14/6258hybridomagenome-scale metabolic modelantiapoptotic genemitochondrial transportBHRF1 |
| spellingShingle | Iván Martínez-Monge Pere Comas David Catalán-Tatjer Jordi Prat Antoni Casablancas Carlos Paredes Martí Lecina Jordi Joan Cairó The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line Applied Sciences hybridoma genome-scale metabolic model antiapoptotic gene mitochondrial transport BHRF1 |
| title | The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line |
| title_full | The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line |
| title_fullStr | The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line |
| title_full_unstemmed | The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line |
| title_short | The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line |
| title_sort | effect of the expression of the antiapoptotic bhrf1 gene on the metabolic behavior of a hybridoma cell line |
| topic | hybridoma genome-scale metabolic model antiapoptotic gene mitochondrial transport BHRF1 |
| url | https://www.mdpi.com/2076-3417/11/14/6258 |
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