Abstract
Oxidative stress affects all the structures of the human eye, particularly the retina and its retinal pigment epithelium (RPE). The RPE limits oxidative damage by several protective mechanisms, including the non-enzymatic antioxidant system zinc-metallothionein (Zn-MT). This work aimed to investigate the role of Zn-MT in the protection of RPE from the oxidative damage of reactive oxygen intermediates by analytical and biochemical-based techniques. The Zn-MT system was induced in an in vitro model of RPE cells and determined by elemental mass spectrometry with enriched isotopes and mathematical calculations. Induced-oxidative stress was quantified using fluorescent probes. We observed that 25, 50 or 100 μM of zinc induced Zn-MT synthesis (1.6-, 3.6- and 11.9-fold, respectively), while pre-treated cells with zinc (25, 50, and 100 μM) and subsequent 2,2′-Azobis(2-methylpropionamidine) dihydrochloride (AAPH) treatment increased Zn-MT levels in a lesser extent (0.8-, 2.1-, 6.1-fold, respectively), exerting a stoichiometric transition in the Zn-MT complex. Moreover, AAPH treatment decreased MT levels (0.4-fold), while the stoichiometry remained constant or slightly higher when compared to non-treated cells. Convincingly, induction of Zn-MT significantly attenuated oxidative stress produced by free radicals’ generators. We conclude that the stoichiometry of Zn-MT plays an important role in oxidative stress response, related with cellular metal homeostasis.
Highlights
The human eye is a highly specialized organ of vision in which the light energy interacts with the neurosensory cells of the retina, where the photoreception process occurs [1]
We studied how zinc supplementation affects the Zn-MT redox system, and the further effects of oxidative stress-induced by AAPH on the levels of zinc, MTs, and the Zn-MTs stoichiometry, by isotope pattern deconvolution (IPD), IDA-HPLC-inductively coupled plasma-mass spectrometer (ICP-MS) following previous procedures [28]
We used an in vitro model of retinal pigment epithelium (RPE) cells to study the protective role of the Zn-MT system against oxidative stress
Summary
The human eye is a highly specialized organ of vision in which the light energy interacts with the neurosensory cells of the retina, where the photoreception process occurs [1]. The retina consists of an inner multilayer of neurosensory cells and an outer single neuroepithelium, the retinal pigment epithelium (RPE). RPE oxidative damage and inflammatory processes contribute to the pathogenesis of age-related eye diseases, including glaucoma, age-related macular degeneration (AMD), diabetic retinopathy and retinitis pigmentosa [7,8,9,10]. The continuous exposure of the RPE to light energy, the oxygen-rich environment and the high metabolic activity provide an ideal framework for the formation of reactive oxygen species (ROS) with the potential to damage proteins, deoxyribonucleic acid (DNA) and lipids [11]
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