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A Pawlak, M Rózanowska, JM Burke, T Sarna, JD Simon; Role of A2E in Photoreactivity of Lipofuscin Granules . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4566.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To determine the role of A2E in photoreactivity of lipofuscin (LF) granules. Methods:LF granules were isolated from human RPE from donors above 40 years old. A2E was synthesized according to the procedure described by Parish et al [Proc. Natl. Acad. Sci. USA, (1998) 95: 14609-13]. Concentrations of LF granules were measured by counting in a hemocytometer. Lipophilic components of LF, including A2E, were extracted by Folch's method. Photoreactivity of LF and A2E was compared during irradiation with narrow or broad band light by 1) monitoring of photo-induced oxygen uptake measured by electron spin resonance (ESR) oximetry, 2) photogeneration of free radicals measured by ESR spin trapping, and 3) photogeneration of singlet oxygen measured by time-resolved detection of singlet oxygen phosphorescence. Results:The values of the rates of blue light-induced oxygen uptake mediated by 6x109 LF granules/ml, extracted lipophilic components of LF and A2E, are 40, 7.2 and 0.007 mM/min, respectively, when normalized to the corresponding content per lipofuscin granule. Even when the concentration of A2E was increased 267 times, to 0.12 mM, so the absorption at the excitation wavelength (434 nm) matched the absorption of LF extract, still the rates of photo-induced oxygen uptake were more than 4 times lower than that for LF extract. For LF extract, the rates of photo-induced oxygen uptake were further increased by in the presence of polyunsaturated lipids, while the additional substrates of oxidation had very little effect on the rates of oxygen uptake mediated by A2E. The action spectrum of photo-induced oxygen uptake mediated by LF extract matched absorption spectrum of LF extract, but not A2E. The rate of photo-induced accumulation of spin adducts with superoxide mediated by LF extract was higher than that mediated by corresponding amount of A2E. The quantum yield of singlet oxygen generation by photoexcited A2E was 4 times lower than that mediated by LF extract. Conclusion:A2E plays only a minor role in aerobic photoreactivity of LF and the main photosensitizer(s) responsible for LF photoreactivity remain to be identified.
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