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K. L. Rhoades, Y. Patel, R. J. Collier, C. Romano; Agonism of the Nicotinic Acetylcholine Receptors Protects ARPE-19 Cells From Oxidative Damage. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5913.
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Oxidative damage to the RPE cells is thought to play a major role in the development of the progression of Age-related Macular Degeneration (AMD). Oxysterols are oxygenated derivatives of cholesterol that act as key regulatory molecules in a variety of cellular processes. Recently, there have been several reports implicating oxysterols, in particular 7-ketocholesterol (7kCh), as causative in the oxidative damage to retinal pigment epithelial (RPE) cells observed in AMD Increasing evidence demonstrates that activation of the nicotinic acetylcholine receptors (nAChRs) provides neuroprotection from several different insults including β amyloid, hypoxia, and glutamate toxicity. Several groups have demonstrated that the α7 and α4β2 nAChRs specifically are responsible for the neuroprotective effects of nicotine. We wished to evaluate the ability of nicotine, a potent nAChR agonist, to prevent RPE cell death when exposed to oxidative damage.
Using the spontaneously arising human RPE cell line, ARPE-19, as a model system for cultured RPE cells, we have developed an in vitro survival assay using 7kCh as a cell death induction signal. Oxidative damage and apoptosis is induced by the addition of 7kCh, in the absence or presence of neurotrophic factors. WST-1 is used to measure cell viability following 24 hours of treatment with 7kCh. Nicotine was evaluated in this model at concentrations from 100 nM to 100 µM.
ARPE-19 cells exposed to 7kCh in the presence of nicotine at concentrations of 30 µM and 100 µM exhibited increased survival compared cells exposed to 7kCh alone. ARPE-19 cells need to be pre-treated with nicotine to allow protection with this compound. Adding nicotine only at the time of the oxidative stress does not protect the cells from damage, indicating that nicotine is not acting exclusively as an antioxidant. Furthermore, specific antagonists of α7 and α4β2 nAChRs (metheyllycaconitine and di-hydro-beta-ethroidine, respectively) block the protection caused by nicotine. We did not observe any change in cell morphology or proliferation in response to treatment with nicotine alone at the concentrations found to be effective for protection.
Nicotine treatment causes a profound, receptor mediated protection of ARPE-19 cells exposed to severe oxidative injury cause by 7kCh. This class of compounds, in particular, agonists of the α7 and α4β2 nicotinic acetyl choline receptors, may provide novel therapeutic strategies for the treatment of AMD.
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