title: Aggravation of cold-induced injury in Vero-B4 cells by RPMI 1640 medium - identification of the responsible medium components.
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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Institut für Physiologische Chemie, Universitätsklinikum Essen, Universität Duisburg-Essen, Hufelandstr, 55, 45122, Essen, Germany.
authors.forename: Gesine
authors.initials: G
authors.lastname: Pless-Petig
authors.forename: Martin
authors.initials: M
authors.lastname: Metzenmacher
authors.forename: Tobias R
authors.initials: TR
authors.lastname: Türk
authors.forename: Ursula
authors.initials: U
authors.lastname: Rauen
mesh_terms: D000241:Adenosine / Q000494:pharmacology; D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000737:chemistry / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002522:Chlorocebus aethiops; D003470:Culture Media / Q000494:pharmacology*; D003676:Deferoxamine / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D005978:Glutathione / Q000494:pharmacology; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects; D007328:Insulin / Q000494:pharmacology; D018374:LLC-PK1 Cells; D019188:Organ Preservation Solutions / Q000494:pharmacology; D010710:Phosphates / Q000737:chemistry / Q000494:pharmacology; D011887:Raffinose / Q000494:pharmacology; D051381:Rats; D013552:Swine; D013696:Temperature; D014709:Vero Cells
chemical_list: D003470:Culture Media; D007328:Insulin; D019188:Organ Preservation Solutions; D010710:Phosphates; C055206:University of Wisconsin-lactobionate solution; D000493:Allopurinol; D005978:Glutathione; D003676:Deferoxamine; D000241:Adenosine; D011887:Raffinose; D002118:Calcium