Influence of Bcl-2 on cell death during the cultivation of a Chinese hamster ovary cell line expressing a chimeric antibody

Tey, Beng Ti; Singh, R. P.; Piredda, Lucia; Piacentini, Mauro; Al‐Rubeai, Mohamed

doi:10.1002/(sici)1097-0290(20000405)68:1<31::aid-bit4>3.0.co;2-l

Cited by 126 publications

(63 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, in the present study no evidence of apoptosis was observed by fluorescence microscopy, in fact ammonia levels studied did not have a detrimental affect on either HepZ proliferation or survival. During HepZ batch culture, the cumulative ammonia levels recorded exceeded cytotoxic levels in hybridoma and myeloma cultures, with a concentration reaching 7.4 mM recorded (not shown) (Mecille and Massie 1994;Singh et al 1994;Tey et al 2000b). No adverse affects were observed on increasing ammonia levels to 14 mM, indicating that HepZ have retained the ability to metabolise excess ammonia via ureogenesis.…”

Section: Discussionmentioning

confidence: 90%

“…Since its discovery as a regulator of apoptosis, bcl-2 has been seen as the prototype apoptosis suppressor gene and has been successfully overexpressed in many cell types including Burkitts lymphoma (Singh et al 1996), hybridoma (Simpson et al 1997(Simpson et al , 1998(Simpson et al , 1999Ishaque and AlRubeai 2002), myeloma (Suzuki et al 1997;Tey et al 2000a), CHO (Fussenegger et al 2000;Tey et al 2000b) and hepatic cells (Lacronique et al 1996). However, the assertion that over-expression of bcl-2 is the answer to the apoptosis problem in biotechnology has come under increasing scrutiny.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Apoptosis and Its Suppression in Hepatocytes Culture

Mukwena

Al‐Rubeai

2004

Cytotechnology

View full text Add to dashboard Cite

In order to achieve the goal of developing extracorporeal liver support devices, it is necessary to optimise bioprocess environment such that viability and function are maximised. Optimising culture medium composition and controlling the constitution of the cellular microenvironment within the bioreactor have for many years been considered vital to achieving these aims. Coupled to this is the need to understand apoptosis, the prime suspect in the demise of animal cultures, including those of hepatocytes. Results presented here show that absent nutrients including glucose and amino acids play a substantial part in the induction of apoptosis. The use of chemical apoptosis inhibitors was utilised to investigate key components of hepatic apoptosis where caspases, predominantly caspase 8, were implicated in staurosporine (STS)-induced HepZ apoptosis. Caspase 9 and 3 activation although recorded was of less significance. Interestingly, these results were not consistent with those of mitochondrial membrane depolarisation where inhibition of caspase activation appeared to drive depolarisation. Inhibition of mitochondrial permeability transition and use of anti-oxidants was unsuccessful in reducing apoptosis, caspase activation and mitochondrial membrane depolarisation. In further studies, the anti-apoptotic gene bcl-2 was over-expressed in HepZ, resulting in a cell line that was more robust and resistant to death induced by glucose and cystine deprivation and treatment with STS. Bcl-2 did not however show significant cytoprotectivity where apoptosis was stimulated by deprivation of glutamine and serum. Overall, results indicated that although apoptosis can be curbed by use of chemical inhibitors and genetic manipulation, their success is dependent on apoptotic stimuli.

show abstract

Section: Discussionmentioning

confidence: 90%

Section: Discussionmentioning

confidence: 99%

Apoptosis and Its Suppression in Hepatocytes Culture

Mukwena

Al‐Rubeai

2004

Cytotechnology

View full text Add to dashboard Cite

show abstract

“…In general most researchers have observed increased maximum cell number, enhanced viability, extended culture duration, protection from viral and sodium butyrate mediated apoptosis and in certain cases increased titers following expression of Bcl family members. (Fassnacht et al 1999;Tey et al 2000;Goswami et al 1999;Simpson et al 1999;Fussenegger et al 2000;Mastrangelo et al 2000a, b;Meents et al 2002;Chiang and Sisk 2005;Kim and Lee 2002;Sung and Lee 2005;Sauerwald et al 2006;Ifandi and Al-Rubeai 2005). Viral homologues of Bcl-2 including E1B19K have also been used for the inhibition of apoptosis in mammalian cell cultures (Wong et al 2006;Figueroa et al 2006), where E1B19K inhibits the apoptotic activity of p53 (Debbas and White 1993;Lowe and Ruley 1993).…”

Section: Mirna Targets Regulating Apoptosismentioning

confidence: 99%

MicroRNAs: recently discovered key regulators of proliferation and apoptosis in animal cells

Gammell

2007

Cytotechnology

View full text Add to dashboard Cite

The relatively recent discovery of miRNAs has added a completely new dimension to the study of the regulation of gene expression. The mechanism of action of miRNAs, the conservation between diverse species and the fact that each miRNA can regulate a number of targets and phenotypes clearly indicates the importance of these molecules. In this review the current state of knowledge relating to miRNA expression and gene regulation is presented, outlining the key morphological and biochemical features controlled by miRNAs with particular emphasis on the key phenotypes that impact on cell growth in bioreactors, namely proliferation and apoptosis.

show abstract

“…S6322-1VL; Sigma-Aldrich, Inc.) 3 days before harvesting for fusion. The spleen was harvested, and 1.56 · 10 7 splenocytes were fused to NS0 Bcl2 cells (43,44) with Polyethylene Glycol 1500 (catalog no. 10 783 641 001; Roche) by methods described previously.…”

Section: Methodsmentioning

confidence: 99%