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B. Rozanowski, M.B. Rozanowska, M.E. Boulton; Toxicity of All-Trans Retinal to the Retinal Pigment Epithelium . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1643.
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Purpose: To determine whether, due to the proximity of the retinal pigment epithelium (RPE) to photoreceptor outer segments (POS), all-trans retinal (RAL) accumulated in POS as a result of photobleaching of rhodopsin, imposes a risk of oxidative and photooxidative damage to RPE cells. Methods: To simplify the system a confluent monolayer of a human RPE cell line (ARPE-19) was exposed to liposomes containing RAL (0-2.6 mM). Liposomes were made of phosphatidylcholine (PC) or a mixture of PC and phosphatidylethanolamine (PE; 65:35 mol/mol), which is known to form condensation products with RAL within POS. Cells were incubated in dark or exposed to visible light (2.4 mW/cm2) for up to 3 hours. Cell viability was determined using the MTT assay for mitochondrial activity and a live/dead double-staining assay for plasma membrane permeability. Products formed due to interaction of RAL and PE in liposomes were identified by HPLC separation and absorption detection. Results: It was observed that RAL at concentrations above 0.5 mM induced a statistically significant toxic effect even in the dark after 1 h incubation time but only if incorporated in PC liposomes. In the presence of both RAL and visible light, the toxicity of RAL/PC liposomes was observed even for 0.1 mM RAL and three hours incubation time. Dark toxicity was partially ameliorated upon addition of catalase. Phototoxicity of RAL was significantly reduced by a-tocopherol and zeaxanthin present either in liposomes or in RPE cells. The toxicity and phototoxicity of RAL was dramatically inhibited when PE was incorporated in liposomes. Even for 1 hour incubation in the presence of 2.6 mM RAL, which induced death of almost all cells in the absence of PE, in the presence of PE above 90% of the cells remained viable. In liposomes containing PE, RAL formed condensation products with PE, i.e. N-retinylidene phosphatidylethanolamine and its protonized form. Conclusions: Exposure of RPE cells in culture to liposomes containing RAL leads to toxic and phototoxic effects, which are dramatically ameliorated if PE is a liposomal component due to formation of products with RAL, which are substantially less toxic than free RAL.
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