September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Extracellular Presence of a Non-Specific Retinal Carrier Prevents the Formation of Lipofuscin Precursors from 11-Cis Retinal in Human Rod Photoreceptor Outer Segments
Author Affiliations & Notes
  • Leopold Adler
    Ophthalmology, Medical University of South Carolina, Charleston, South Carolina, United States
  • Chunhe Chen
    Ophthalmology, Medical University of South Carolina, Charleston, South Carolina, United States
  • Federico Gonzalez-Fernandez
    Departments of Ophthalmology & Pathology, University of Mississippi School of Medicine, Jackson, Mississippi, United States
    R&D Division, Veterans Affairs Medical Center, Jackson, Mississippi, United States
  • Yiannis Koutalos
    Ophthalmology, Medical University of South Carolina, Charleston, South Carolina, United States
  • Footnotes
    Commercial Relationships   Leopold Adler, None; Chunhe Chen, None; Federico Gonzalez-Fernandez, None; Yiannis Koutalos, None
  • Footnotes
    Support  Supported by: NEI Grant EY014850 (YK); an unrestricted award to the Department of Ophthalmology at Medical University of South Carolina from Research to Prevent Blindness; Start-up Award, Research! Mississippi; Activation award, VA Office of Research and Development (ORD); Merit Review Award I01BX007080, ORD (F.G.-F.); NEI grant EY09412 (F.G.-F.)
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 1765. doi:
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      Leopold Adler, Chunhe Chen, Federico Gonzalez-Fernandez, Yiannis Koutalos; Extracellular Presence of a Non-Specific Retinal Carrier Prevents the Formation of Lipofuscin Precursors from 11-Cis Retinal in Human Rod Photoreceptor Outer Segments. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1765.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : The 11-cis retinal chromophore of rhodopsin, the rod visual pigment, is supplied to rod photoreceptor outer segments extracellularly via the interphotoreceptor matrix. 11-cis retinal can react with outer segments to generate precursors of lipofuscin that eventually accumulate in lysosomes of the retinal pigment epithelium . We tested whether extracellular bovine serum albumin (BSA), a non-specific retinal carrier, can prevent the formation of lipofuscin precursors from 11-cis retinal in isolated human rod photoreceptors.

Methods : Fresh human donor eyes were procured through the National Disease Resource Interchange. Eyes were dissected and the retinas isolated within 48 hrs after death. Isolated retinas were bleached to destroy residual rhodopsin, and stored in a physiological saline solution at 4 °C. Metabolically active rod photoreceptors were isolated from the peripheral region of human retinas. Interphotoreceptor retinoid binding protein (IRBP) was extracted from bovine retinas and purified to homogeneity by combination of Con-A affinity, ion exchange, and size-exclusion chromatography. Its concentration was determined by amino acid analysis and absorption spectrophotometry. Rod photoreceptors were isolated from the bleached retinas and incubated in the presence of either IRBP and 11-cis retinal to regenerate rhodopsin, or IRBP alone. After incubation, cells were washed with a physiological saline solution and placed on an epifluorescence microscope stage at 37 °C. Cells were then exposed to different concentrations of 11-cis retinal with different concentrations of BSA as carrier. The levels of rod outer segment lipofuscin precursors were determined from their fluorescence (ex, 490 nm; em >520 nm).

Results : Exposure to 11-cis retinal resulted in a significant increase in lipofuscin precursor levels, which was higher for cells that contained rhodopsin. Higher extracellular concentrations of BSA resulted in lower levels of lipofuscin precursors formed by the exogenous 11-cis retinal.

Conclusions : 11-cis retinal can generate lipofuscin precursors in the outer segment of human rod photoreceptors. The extracellular presence of a retinal carrier limits the formation of lipofuscin precursors, likely by buffering 11-cis retinal.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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