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R. K. Crouch, P. Goletz, Y. Koutalos, K. L. Schey, A. Hanneken, Z. Ablonczy; Human A2E Levels Are Higher in the Peripheral (Extramacular) RPE Than in the Macular Region of the RPE. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1300.
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To quantitate the levels of A2E and other bis-retinoids in the extramacular and macular regions of the human RPE.
Human RPE was removed from fresh autopsy eyes and dissected into two portions: a 1 cm central circle and the remaining tissue outside that section. The RPE was extracted with 1:1 chloroform/methanol and analyzed by HPLC. For mass spectral imaging, the RPE/choroid was carefully dissected and flattened onto ITO coated slides, dried, and sprayed with 15 mg/mL sinnapinic acid in 70% ethanol. Tissues were analyzed in a Bruker Autoflex III TOF-TOF mass spectrometer in 250 micron steps across the tissue surface in the 520-2600 m/z range.
Quantitation of A2E levels by HPLC indicated that the levels are 2 to 5 fold higher in the peripheral RPE (extramacular) than in the macular region. The chromatographic profiles of the retinoid extracts were similar in autopsy eyes ranging from ages 59 to 89 years although there was an overall decrease in total A2E with advancing age. Extracts of the macular RPE contain an unidentified major component with an absorbance maximum of 430 nm; however this is only a minor component in the peripheral RPE extract. Mass spectral imaging showed the oxidized (plus one and two oxygens) forms of A2E follow the same pattern as that for the nonoxidized A2E, namely strong in the peripheral RPE and weak in the macula. Mass spectral imaging of a fetal human eye showed no A2E, indicating that the compound is a product of a functional visual cycle.
The distribution of A2E in the human RPE does not follow the fluorescence pattern of lipofuscin and the levels are higher in the extramacular RPE than in the macular region.
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