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Updated: Aug 6, 2026
Primary Culture of Porcine Retinal Pigment Epithelial Cells
Published on: September 23, 2022
Lack of antioxidative properties of vitamin C and pyruvate in cultured retinal pigment epithelial cells
Oliver Zeitz1, Lars Schlichting, Gisbert Richard
1Universitätsklinikum Hamburg-Eppendorf, Klinik und Poliklinik ffir Augenheilkunde, Martinistr. 52, 20246 Hamburg, Germany. zeitz@uke.uni-hamburg.de
Background:
Oxidative damage to the retinal pigment epithelium might be involved in the pathogenesis of age related macular degeneration. Thus antioxidative protection represents a rationale for a causative therapy or prophylaxis. The aim of the present study is to evaluate antioxidative properties of vitamin C and pyruvate at retinal pigment epithelial (RPE) cells exposed to oxidative stress.
Methods:
The ability of vitamin C and pyruvate to quench hydroxyl radicals was tested using the di-hydro-rhodamine (DHR) assay. Cells of the human RPE cell line ARPE-19 were exposed for 8 min to hydroxyl radicals generated by the Fenton reaction from 2.25 mM H2O2 and 30 microM Fe3+ -nitrilo-tri-acetate. This was done in the absence and presence of 0.3-3.0 mM pyruvate and vitamin C, respectively. Cell survival was analysed by vitality staining (life-dead-assay) and expressed as cell survival ratio. A survival ratio <1.0 indicates cell loss.
Results:
At concentrations from 0.1 to 1.0 mM vitamin C and pyruvate quench hydroxyl radicals in the DHR assay in absence of living matter. In the presence of 0.1- 0.3 mM vitamin C and pyruvate, ARPE-19 showed a reduced survival ratio (0.87 +/- 0.01 to 0.89 +/- 0.02 after 6 h) which was not the case at the higher concentrations between 1 and 3 mM. The exposure of ARPE-19 cells to hydroxyl radicals reduced the survival ratio to 0.92 +/- 0.02. At concentrations at which vitamin C and pyruvate exert toxic effects, a potentiation of radical induced cell death can be observed (survival ratio 0.79 +/- 0.02 and 0.82 +/- 0.03, respectively). Higher concentrations of vitamin C or pyruvate had no explicit protective effect to the hydroxyl radical induced damage.
Discussion:
Although vitamin C and pyruvate are potent hydroxyl radical quenchers in vitro they failed to protect cultured ARPE-19 cells from oxidative stress induced cell death. In contrast, when applying the scavengers at low concentrations a potentiation of cell damage was observed.
