Bilberry (Vaccinium myrtillus) anthocyanins modulate heme oxygenase-1 and glutathione S-transferase-pi expression in ARPE-19 cells.
OBJECTIVE
To determine whether anthocyanin-enriched bilberry extracts modulate pre- or posttranslational 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 preincubated with anthocyanin and nonanthocyanin phenolic fractions of a 25% enriched extract of bilberry (10(-6)-1.0 mg/mL) and, after phenolic removal, cells were oxidatively challenged with H(2)O(2). 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
Preincubation with bilberry extract ameliorated the intracellular increase of H(2)O(2)-induced free radicals in RPE, though H(2)O(2) cytotoxicity was not affected. By 4 hours, the extract had upregulated 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 nonanthocyanin phenolic fractions contributed similarly to mRNA upregulation.


CONCLUSIONS
Anthocyanins and other phenolics from bilberry upregulate 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.
Milbury
Paul E
PE
Antioxidants Research Laboratory, Jean Mayer USDA Human Nutrition Research Center on Aging, Tufts University, Boston, Massachusetts 02111, USA. paul.milbury@tufts.edu
Graf
Brigitte
B
Curran-Celentano
Joanne M
JM
Blumberg
Jeffrey B
JB
D000872:Anthocyanins / Q000494:pharmacology*; D015153:Blotting, Western; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D005609:Free Radicals / Q000378:metabolism; D005960:Glucosides / Q000494:pharmacology*; D051549:Glutathione S-Transferase pi / Q000235:genetics / Q000378:metabolism*; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D018384:Oxidative Stress; D010857:Pigment Epithelium of Eye / Q000187:drug effects* / Q000378:metabolism; D010936:Plant Extracts / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D015854:Up-Regulation; D029800:Vaccinium myrtillus / Q000737:chemistry*
D000872:Anthocyanins; D005609:Free Radicals; D005960:Glucosides; D010936:Plant Extracts; D012333:RNA, Messenger; C462279:cyanidin-3-O-beta-glucopyranoside; D006861:Hydrogen Peroxide; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D051549:Glutathione S-Transferase pi