The influence of Bcl-2 expression on the suppression of apoptosis during the cultivation of an NS0 cell line expressing a chimeric antibody was investigated. Following selection of transfectants in medium containing G418, Western analysis revealed evidence of some up-regulation of endogenous Bcl-2 expression even in the control vector transfectants. Cultivation of the two cell lines in suspension batch cultures clearly demonstrated the enhanced robustness of the bcl-2 vector transfected cells. Suppression of apoptosis resulted in an approximately 20% increase in maximum viable cell number, and a doubling in culture duration compared to the control transfected cells. However, despite the significant affect on viability, Bcl-2 expression did not result in an increase in final antibody titre in comparison with the control cell line. Exposure of cells to various nutrient limited conditions further emphasised the influence of Bcl-2 on cell survival. After 3 days of exposure to serum, glucose, glutamate and asparagine deprivation, the viable cell number and viability were significantly higher in the bcl-2 transfected cell line. When control cells were deprived of all amino acids, there was a complete loss of viability and viable cell number within 3 days. By contrast, the bcl-2 transfected cell line retained greater than 75% of the initial viable cell number and about 70% viability. In response to exposure to 8 mM thymidine (a cytostatic agent) the control cell line underwent complete loss of viability and viable cell number after 6 days. This compared with 18 days for complete loss of viability in the bcl-2 transfected cell line. As under batch culture conditions, there was no difference between the two cell lines in final antibody titre, which indicated that MAb synthesis is limited by nutrient availability during the latter stages of culture in both cases. When fed batch cultures were carried out using a concentrated essential amino acid feed, the bcl-2 cell line exhibited a 60% increase in maximum viable cell number and a 50% increase in culture duration, when compared to the control cell line. Moreover, the bcl-2 cell line exhibited a greater than 40% increase in maximum antibody titre. (C) 2000 Elsevier Science B.V. All rights reserved.

Tey, B., Singh, R., Piredda, L., Piacentini, M., Al Rubeai, M. (2000). Bcl-2 mediated suppression of apoptosis in myeloma NS0 cultures. JOURNAL OF BIOTECHNOLOGY, 79(2), 147-159 [10.1016/S0168-1656(00)00223-6].

Bcl-2 mediated suppression of apoptosis in myeloma NS0 cultures

PIREDDA, LUCIA;PIACENTINI, MAURO;
2000-01-01

Abstract

The influence of Bcl-2 expression on the suppression of apoptosis during the cultivation of an NS0 cell line expressing a chimeric antibody was investigated. Following selection of transfectants in medium containing G418, Western analysis revealed evidence of some up-regulation of endogenous Bcl-2 expression even in the control vector transfectants. Cultivation of the two cell lines in suspension batch cultures clearly demonstrated the enhanced robustness of the bcl-2 vector transfected cells. Suppression of apoptosis resulted in an approximately 20% increase in maximum viable cell number, and a doubling in culture duration compared to the control transfected cells. However, despite the significant affect on viability, Bcl-2 expression did not result in an increase in final antibody titre in comparison with the control cell line. Exposure of cells to various nutrient limited conditions further emphasised the influence of Bcl-2 on cell survival. After 3 days of exposure to serum, glucose, glutamate and asparagine deprivation, the viable cell number and viability were significantly higher in the bcl-2 transfected cell line. When control cells were deprived of all amino acids, there was a complete loss of viability and viable cell number within 3 days. By contrast, the bcl-2 transfected cell line retained greater than 75% of the initial viable cell number and about 70% viability. In response to exposure to 8 mM thymidine (a cytostatic agent) the control cell line underwent complete loss of viability and viable cell number after 6 days. This compared with 18 days for complete loss of viability in the bcl-2 transfected cell line. As under batch culture conditions, there was no difference between the two cell lines in final antibody titre, which indicated that MAb synthesis is limited by nutrient availability during the latter stages of culture in both cases. When fed batch cultures were carried out using a concentrated essential amino acid feed, the bcl-2 cell line exhibited a 60% increase in maximum viable cell number and a 50% increase in culture duration, when compared to the control cell line. Moreover, the bcl-2 cell line exhibited a greater than 40% increase in maximum antibody titre. (C) 2000 Elsevier Science B.V. All rights reserved.
2000
Pubblicato
Rilevanza internazionale
Articolo
Sì, ma tipo non specificato
Settore BIO/06 - ANATOMIA COMPARATA E CITOLOGIA
English
protein bcl 2; antibody titer; apoptosis; article; cell count; cell growth; cell survival; cell viability; controlled study; gene expression regulation; human; human cell; priority journal; Ammonia; Animals; Antibodies, Monoclonal; Apoptosis; Cell Division; Genes, bcl-2; Humans; Mice; Multiple Myeloma; Tumor Cells, Cultured
Tey, B., Singh, R., Piredda, L., Piacentini, M., Al Rubeai, M. (2000). Bcl-2 mediated suppression of apoptosis in myeloma NS0 cultures. JOURNAL OF BIOTECHNOLOGY, 79(2), 147-159 [10.1016/S0168-1656(00)00223-6].
Tey, B; Singh, R; Piredda, L; Piacentini, M; Al Rubeai, M
Articolo su rivista
File in questo prodotto:
Non ci sono file associati a questo prodotto.

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2108/46356
Citazioni
  • ???jsp.display-item.citation.pmc??? ND
  • Scopus 62
  • ???jsp.display-item.citation.isi??? 69
social impact