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K. M. Bertram, C. J. Baglole, R. P. Phipps, R. T. Libby; Cigarette Smoke Induces Oxidative Stress and Apoptosis in Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1838.
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The single most important environmental risk factor for age-related macular degeneration (AMD) is cigarette smoke. This study examined the damage that cigarette smoke causes to retinal pigment epithelial (RPE) cells in vitro.
After exposure to cigarette smoke extract (CSE) or hydroquinone (HQ), ARPE-19 cells or primary human RPE cells were assessed for viability, mitochondrial membrane potential (ΔΨm), mitochondrial superoxide production, intracellular glutathione (GSH) content, cell size, and 4-hydroxy-2-nonenal (4-HNE). Furthermore, expression of heme oxygenase-1 (HO-1) and vascular endothelial growth factor (VEGF) was also assessed. To establish whether antioxidants could protect RPE cells from CSE-induced damage, cell viability was quantified after GSH and N-acetyl-cysteine (NAC) were added to control and CSE-exposed cultures.
CSE induced oxidative damage and apoptosis in both ARPE-19 and primary human RPE cells. Viability decreased in a dose-dependant manner, where the LD50 for both ARPE-19 cells and human primary RPE cells occurred at a dose of 10% CSE. Morphological changes indicative of apoptosis in ARPE-19 cells, including a reduction in cell size and nuclear condensation, were observed after CSE exposure. Significant increases in lipid peroxidation (4-HNE) and mitochondrial superoxide production were observed. Significant decreases in intracellular glutathione (GSH) levels were observed after 5% CSE exposure compared to control (15 ± 3.3 nmoles/mg protein vs. 35 ± 8.4 nmoles/mg protein, respectively; p <0.05) . Furthermore, HO-1 and VEGF expression was induced in CSE-exposed cells. Exogenous administration of antioxidants (GSH and NAC) prevented oxidative damage to the RPE cells caused by CSE, as measured by ΔΨm, 4-HNE and mitochondrial superoxide production.
These results demonstrate that cigarette smoke is a potent inducer of oxidative cell death in human RPE cells. These data support the hypothesis that exposure to cigarette smoke severely damages RPE cells and may trigger important events in AMD pathogenesis including RPE cell death and VEGF expression.
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