ARS | Publication request: Bilberry (Vaccinium myrtillus) Anthocyanins Modulate Heme Oxygenase-1 and Glutathione S-Transferase-pi Expression in the ARPE-19 Cells

Submitted to: Investigative Opthalmology and Visual Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 9, 2007
Publication Date: April 26, 2007
Publisher's URL: http://iovs.org/cgi/content/abstract/48/5/2343
Citation: Milbury, P., Graf, B., Curran-Celentano, J.M., Blumberg, J. 2007. Bilberry (Vaccinium myrtillus) Anthocyanins Modulate Heme Oxygenase-1 and Glutathione S-Transferase-pi Expression in the ARPE-19 Cells. Investigative Opthalmology and Visual Science. 48(5):2343-2349.

Interpretive Summary: Bilberry, a close relative to the commercial blueberry, are marketed as a supplement to promote eye health. The content of anthocyanins, a class of flavonoids responsible for the color in berries, is high in bilberries. The health properties of anthocyanins, and flavonoids in general, is often attributed to their antioxidant qualities. Yet concentrations of these phytochemicals achieved via dietary means are very low when compared to other endogenous or exogenous antioxidants. Such concentrations are insufficient to attribute direct radical quenching as the mechanism responsible for the health beneficial effects of berry consumption. The research results presented show that, while anthocyanins can quench intracellular radicals they cannot prevent or diminish cell death in retinal pigment epithelial in culture exposed to high levels of oxidant or radical damage. However, both anthocyanin and other phenolics in bilberries can increase levels of messenger RNA for the oxidative stress protective enzymes heme oxygenase-1 and glutathione S-transferase in retinal pigment epithelial cells. These results suggest that, in animals, consumption of berries may promote increases in protective proteins that may aid in increasing endogenous defenses against oxidative damage. Under in vivo conditions of aging, this nutrient/gene interaction may be more important as a protective mechanism than direct antioxidant mechanisms to promoting eye health.

Technical Abstract: PURPOSE. To determine whether anthocyanin-enriched bilberry extracts modulate pre- or post-translational levels of oxidative stress defense enzymes heme-oxygenase (HO)-1 and glutathione S-transferase-pi (GST-pi) in cultured human retinal pigment epithelial (RPE) cells. METHODS. Confluent ARPE-19 cells were pre-incubated with anthocyanin and non-anthocyanin phenolic fractions of a 25%-enriched extract of bilberry (10x-6 to 1.0 mg/mL) and, after phenolic removal, cells were oxidatively challenged with H2O2. The concentration of intracellular glutathione was measured by HPLC and free radical production determined by the dichlorofluorescin diacetate assay. HO-1 and GST-pi protein and mRNA levels were determined by Western blot and RT-PCR, respectively. RESULTS. Pre-incubation with bilberry extract ameliorated the intracellular increase of H2O2 -induced free radicals in RPE, though H2O2 cytotoxicity was not affected. By 4 h, the extract had up-regulated HO-1 and GST-pi protein by 2.8- and 2.5-fold, respectively, and mRNA by 5.5- and 7.1-fold, respectively, in a dose-dependent manner. Anthocyanin and non-anthocyanin phenolic fractions contributed similarly to mRNA up-regulation. CONCLUSIONS. Anthocyanins and other phenolics from bilberry up-regulate the oxidative stress defense enzymes HO-1 and GST-pi in RPE, suggesting that they stimulate signal transduction pathways influencing genes controlled by the antioxidant response element.