Though one prominent theory of atherogenesis involves free-radical oxidation of low-density lipoprotein (LDL) within the vessel wall by one of the vascular cell types, the mechanism for cell-mediated LDL oxidation remains unclear[sn1]. In these studies we examined the effects of media phenols, thiols, and metals on endothelial cell–mediated oxidation. We found that cell culture media such as Dulbecco modified Eagle medium and minimal essential medium are unable to support cell-mediated oxidation of LDL because they contain high concentrations of phenol red (PR) and tyrosine, both of which strongly inhibit cell-mediated oxidation. Ham's F-10, a commonly used medium for cell-mediated oxidation experiments, is also not entirely appropriate, as it contains both PR and cysteine. Cysteine is not critical for endothelial cell–mediated oxidation, but does increase oxidation of LDL in the absence of cells. Finally, of utmost importance to cell-mediated oxidation was the presence of either micromolar concentrations of Fe(II) or physiological concentrations of holo-ceruloplasmin, the protein which carries copper in plasma. An appropriate culture medium for use in cell-mediated oxidation experiments should thus contain either micromolar concentrations of Fe(II) or physiological concentrations of holo-ceruloplasmin, and should be prepared without PR, cysteine, or large concentrations of tyrosine, all of which are shown here to inhibit endothelial cell–mediated LDL oxidation. These results are consistent with a mechanism of cell-mediated oxidation involving Fenton-type chemistry and redox cycling of the metal.
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In Vitro Cellular & Developmental Biology - Animal
Vol. 36 • No. 9
October 2000
Vol. 36 • No. 9
October 2000
atherosclerosis
copper
free radical
iron
lipoprotein