Selective removal of zinc from cell culture media

J. Clair, R. Talwalkar, C. J. McClain, B. Hennig

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Little is known about the requirements and functions of zinc (Zn) in maintaining endothelial cell integrity. A Zn-deficient medium is needed to study this variable under culture conditions. To achieve this goal, several methods were employed to remove Zn selectively from media used for the culture of endothelial cells. Medium 199 (M199) supplemented with 1%, 5%, or 10% fetal bovine serum (FBS) was dialyzed against 0.1% 1,10-orthophenanthroline for 10 days or against 4% chelating resin (chelex) for 10 days at 4°C using trace metal-free dialysis tubing. Alternately, M199 plus FBS was incubated directly with 4% chelex for 10 days at 4°C with constant stirring. Subsequently, changes in medium minerals were determined. Significant decreases in Zn concentration occurred only in M199 plus FBS, when incubated directly with chelex for 10 days at 4°C. There was little or no consistent decrease in Zn concentration with the other methods. Magnesium (Mg) and calcium (Ca) concentrations were not altered significantly by any of the treatments. To determine if the removal of zinc from cell culture media with chelex can affect cellular metabolism, albumin transfer across endothelial monolayers and angiotensin-converting enzyme (ACE) activity were measured. Cells grown in chelex-treated M199 plus 1 or 5% FBS had significantly higher rates of albumin transfer, indicating greater disruption in endothelial barrier function due to Zn deficiency. ACE, a zinc-dependent enzyme, also showed a significant decrease in activity in cells grown in chelex-treated media as compared with controls. We suggest that incubation of FBS-supplemented M199 directly with 4% chelex for 10 days at 4°C can be used as a method for selectively removing Zn from cell culture media. Furthermore, chelex-mediated extraction of Zn from cell culture media produced significant effects on cellular metabolism, indicating that Zn is important for endothelial cell integrity.

Original languageEnglish
Pages (from-to)143-151
Number of pages9
JournalJournal of Trace Elements in Experimental Medicine
Volume7
Issue number4
StatePublished - 1994

Keywords

  • Endothelial cell barrier function
  • Endothelial cells
  • Minerals
  • Nutrition
  • Zinc deficiency

ASJC Scopus subject areas

  • Biochemistry

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