July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Elevated levels of A2E and lipofuscin fluorescence in the retinal pigment epithelium of mice that lack retinol dehydrogenase RDH8 are decreased by dark rearing
Author Affiliations & Notes
  • Yiannis Koutalos
    Ophthalmology, Medical Univ of South Carolina, Charleston, South Carolina, United States
  • Nicholas P Boyer
    Ophthalmology, Medical Univ of South Carolina, Charleston, South Carolina, United States
  • Debra A Thompson
    Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, Michigan, United States
    Biological Chemistry, University of Michigan School of Medicine, Ann Arbor, Michigan, United States
  • Footnotes
    Commercial Relationships   Yiannis Koutalos, None; Nicholas Boyer, None; Debra Thompson, None
  • Footnotes
    Support  NIH grants R01 EY014850 (YK) and P30 EY07003 (DAT), and Research to Prevent Blindness (DAT)
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 2334. doi:
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      Yiannis Koutalos, Nicholas P Boyer, Debra A Thompson; Elevated levels of A2E and lipofuscin fluorescence in the retinal pigment epithelium of mice that lack retinol dehydrogenase RDH8 are decreased by dark rearing. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2334.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : To examine the magnitude of the contribution of all-trans retinal to lipofuscin formation in the RPE. All-trans retinal released by bleached photopigments is reduced to all-trans retinol by RDH8, whereas 11-cis retinal is not a RDH8 substrate. Both 11-cis and all-trans retinal can form bis-retinoids, which comprise a major component of RPE lipofuscin. Mice that lack RDH8 are unable to reduce all-trans retinal to all-trans retinol, maximizing the contribution of all-trans retinal to bis-retinoid and lipofuscin formation.

Methods : Rdh8-/- mice were reared in cyclic light or in darkness for up to 6 months. Some animals were switched from light to dark-rearing for periods ranging from 1 to 8 weeks. The bis-retinoid A2E was measured from chloroform-methanol extracts of RPE-choroid samples with HPLC-UV/VIS spectroscopy. Lipofuscin fluorescence was measured from whole flattened eyecups after removal of the retina on a SP2 Leica Laser Scanning Confocal microscope (excitation, 488 nm; emission, 565-725 nm). Samples were prepared under dim red light.

Results : Both lipofuscin fluorescence and A2E levels increased with age in both cyclic-light- and dark-reared mice. Fluorescence emission spectra of RPE lipofuscin granules were similar in dark- and cyclic-light-reared animals, with a broad peak at ~610 nm. A2E accumulated at rates of ~1.7 and ~1.1 pmol/eye/month in cyclic-light- and dark-reared animals, respectively. RPE lipofuscin fluorescence increased at a rate ~2× higher in light- compared to dark-reared animals. Moving mice from cyclic-light to dark-rearing drastically reduced the rate of A2E accumulation. After initially being reared in cyclic light, mice at 6 months of age that had spent the previous 8 weeks in darkness had the same levels of A2E as mice that were dark-reared for the entire 6 month period.

Conclusions : In Rdh8-/- mice reared in cyclic-light, all-trans retinal makes a sizeable contribution to RPE lipofuscin formation. In wild-type mice, more than 80% of all-trans retinal generated by light is rapidly reduced to all-trans retinol, thus we expect the contribution of all-trans retinal to RPE lipofuscin formation be much less than that of 11-cis retinal. The decrease in the levels of A2E in cyclic-light-reared mice that were moved to darkness for different periods of time is consistent with processing of A2E within RPE cells.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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