Abstract
Purpose: :
Oxidative stress is believed to contribute significantly to the pathogenesis of many diseases including age-related macular degeneration, diabetic retinopathy, and glaucoma. Hence, development of methods for antioxidant protection is of vital importance and may be broadly relevant in disease prevention. Selenomethionine (SeMet) is the main dietary form of selenium, an essential trace mineral whose antioxidant properties are well recognized. It is postulated also that dietary supplementation of SeMet may be of benefit in preventing degenerative retinal diseases. However, the mechanism(s) of SeMet action is not totally clear. In mammalian cells, the cystine/glutamate exchanger xc- is the major supplier of cysteine, the limiting amino acid in the synthesis of glutathione (GSH) and thus is an important determinant of GSH status. The retinal pigment epithelium (RPE) normally contains high levels of GSH. The purpose of this study was to investigate the influence of SeMet on the activity of xc- in ARPE-19 cells, a human RPE cell line.
Methods: :
ARPE-19 cells were cultured in the presence or absence of SeMet. Xc- activity was monitored by the uptake of glutamate in the absence of Na+. RT-PCR, Northern blot, immunohistochemistry and Western blot techniques were used to analyze changes in mRNA and protein levels of xCT and of 4f2hc, the light and heavy chains of xc-, respectively. Similar techniques were used to analyze also expression of Nrf2, a key regulator of the antioxidant response. GSH levels were measured using a commercial kit.
Results: :
Xc- activity was 3- to 4-fold higher in SeMet-treated ARPE-19 cells than in control cells. Na+-dependent uptake of glutamate was not affected by SeMet treatment, demonstrating specificity of the SeMet effect on xc-. Nrf2 mRNA and protein expression was upregulated in SeMet treated cells. GSH levels were also significantly increased.
Conclusions: :
SeMet upregulates the cystine/glutamate transporter xc-, increases Nrf2 expression and raises intracellular GSH levels in ARPE-19 cells. This influence of SeMet on xc- may be beneficial in protecting RPE cells against oxidant-induced damage.
Keywords: retina • retinal pigment epithelium • antioxidants