Email updates

Keep up to date with the latest news and content from BMC Biotechnology and BioMed Central.

Open Access Research article

Aggravation of cold-induced injury in Vero-B4 cells by RPMI 1640 medium – Identification of the responsible medium components

Gesine Pless-Petig1, Martin Metzenmacher1, Tobias R Türk23 and Ursula Rauen1*

Author affiliations

1 Institut für Physiologische Chemie, Universitätsklinikum Essen, Universität Duisburg-Essen, Hufelandstr. 55, 45122, Essen, Germany

2 Klinik für Nephrologie, Universitätsklinikum Essen, Universität Duisburg-Essen, Hufelandstr. 55, 45122, Essen, Germany

3 Present address: Medizinische Klinik 4 - Nephrologie und Hypertensiologie, Universität Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg, Ulmenweg 18, 91054, Erlangen, Germany

For all author emails, please log on.

Citation and License

BMC Biotechnology 2012, 12:73  doi:10.1186/1472-6750-12-73

Published: 10 October 2012

Abstract

Background

In modern biotechnology, there is a need for pausing cell lines by cold storage to adapt large-scale cell cultures to the variable demand for their products. We compared various cell culture media/solutions for cold storage of Vero-B4 kidney cells, a cell line widely used in biotechnology.

Results

Cold storage in RPMI 1640 medium, a recommended cell culture medium for Vero-B4 cells, surprisingly, strongly enhanced cold-induced cell injury in these cells in comparison to cold storage in Krebs-Henseleit buffer or other cell culture media (DMEM, L-15 and M199). Manufacturer, batch, medium supplements and the most likely components with concentrations outside the range of the other media/solutions (vitamin B12, inositol, biotin, p-aminobenzoic acid) did not cause this aggravation of cold-induced injury in RPMI 1640. However, a modified Krebs-Henseleit buffer with a low calcium concentration (0.42 mM), a high concentration of inorganic phosphate (5.6 mM), and glucose (11.1 mM; i.e. concentrations as in RPMI 1640) evoked a cell injury and loss of metabolic function corresponding to that observed in RPMI 1640. Deferoxamine improved cell survival and preserved metabolic function in modified Krebs-Henseleit buffer as well as in RPMI 1640. Similar Ca2+ and phosphate concentrations did not increase cold-induced cell injury in the kidney cell line LLC-PK1, porcine aortic endothelial cells or rat hepatocytes. However, more extreme conditions (Ca2+ was nominally absent and phosphate concentration raised to 25 mM as in the organ preservation solution University of Wisconsin solution) also increased cold-induced injury in rat hepatocytes and porcine aortic endothelial cells.

Conclusion

These data suggest that the combination of low calcium and high phosphate concentrations in the presence of glucose enhances cold-induced, iron-dependent injury drastically in Vero-B4 cells, and that a tendency for this pathomechanism also exists in other cell types.

Keywords:
Cell pausing; Cold storage; Iron chelator; Calcium; Phosphate; Preservation; Hypothermia