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

BMC Biotechnology

Gesine Pless-Petig²  Martin Metzenmacher²  Tobias R Türk¹  Ursula Rauen² 

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

[2] Institut für Physiologische Chemie, Universitätsklinikum Essen, Universität Duisburg-Essen, Hufelandstr. 55, 45122, Essen, Germany

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

Others  :  1134761
DOI  :  10.1186/1472-6750-12-73

 received in 2012-07-26, accepted in 2012-10-04,  发布年份 2012

【 摘 要 】

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.

【 授权许可】

   
2012 Pless-Petig et al.; licensee BioMed Central Ltd.

【 图 表 】

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