Abstract
Purpose::
Although the Age-Related Eye Disease Study (AREDS) has shown that supplemental dietary zinc decreases the progression of age-related macular degeneration (AMD), little is understood about the molecular mechanisms of this effect. The purpose of this study is to explore how zinc protects by characterizing the zinc transporter proteins in human retinal pigment epithelial cells (RPE).
Methods::
Human RPE cells were isolated from postmortem donor eyes. Pure cultures were obtained and RPE cells were incubated for 6 hours with various concentrations of zinc (5,10, 50 and 100µmole) and 2 and 4µmole N,N,N,N-Tetrakis(2-Pyridylmethyl)ethylenediamine (TPEN) to induce zinc deficiency . The expression of transporter proteins that mediate the uptake of zinc were analyzed by RT-PCR using gene-specific primers. Flow cytometry with FluoZn TM-3 was utilized to assess the intra-cellular zinc concentration.
Results::
RT-PCR studies suggested that human RPE cells expressed Zip 2 as the predominant transporter. When the extracellular concentration of zinc was increased, the intracellular concentration increased as well, and was accompanied by an increase in the expression of Zip2. When TPEN was added to the culture medium, the intracellular zinc concentration diminished, and was accompanied by a decrease in Zip2 expression.
Conclusions::
Zinc is an important trace element which is present in RPE cells and has been shown to protect the RPE against oxidative stress. The zinc transporter protein, Zip2, was identified as a predominantly expressed transporter in RPE cells and its expression appears to be regulated by the concentration of zinc. This mechanism of uptake should play a critical role in zinc protection of RPE cells from oxidative stress
Keywords: age-related macular degeneration • antioxidants • gene/expression