July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
The hyperspectral autofluorescence (AF) differentiation of lipofuscin in the RPE and vitelliform debris of canine bestrophinopathy
Author Affiliations & Notes
  • Yuehong Tong
    Ophthalmology, Icahn School of Medicine at Mount Sinai, New York, New York, United States
  • Taariq Mohammed
    Ophthalmology, New York University School of Medicine, New York, New York, United States
  • Neel Dey
    Computer Science & Engineering, New York University, New York, New York, United States
  • Thomas Ach
    Ophthalmology, University Hospital Würzburg, Würzburg, Germany
  • Martin Hammer
    Ophthalmology, University of Jena, Jena, Germany
  • R Theodore Smith
    Ophthalmology, Icahn School of Medicine at Mount Sinai, New York, New York, United States
  • Karina E Guziewicz
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 5858. doi:
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    • Get Citation

      Yuehong Tong, Taariq Mohammed, Neel Dey, Thomas Ach, Martin Hammer, R Theodore Smith, Karina E Guziewicz; The hyperspectral autofluorescence (AF) differentiation of lipofuscin in the RPE and vitelliform debris of canine bestrophinopathy. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5858.

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

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Abstract

Purpose :
Mutations of bestrophin 1 (BEST1) cause retinal disorders such as bestrophinopathies in human and canine BEST1 disease (cBest1) in dogs. The spectrum of lipofuscin (LF) in cBest1 and normal dogs has been shown to be the same in PMID:28111324. The further characterization of fluorophores in the RPE and in the vitelliform material of cBest1 may extend our understanding of pathology of cBest1 and more information on the human disease as well.

Methods :
We prepared cross section slides from a canine retina with cBest1. We acquired images using a Nuance hyperspectral camera connected to a Zeiss microscope with 20X objective and 10nm emission intervals from 420 nm to 720 nm at excitations 436 nm, 480 nm and 505 nm. We applied non-negative matrix factorization for image processing to extract 5 abundances and their corresponding spectra.

Results :
Fig. 1 is a combined RGB AF image of a cBest1 retina. Recovered spectra C1 and C3 had peaks at 540 nm and 570 nm, respectively, with a notch at 620 nm in C3, at excitation 436 nm (Fig. 2), strikingly similar to normal human spectra S1 and S2 (PMID:27226929). The abundance images 1 and 3 were localized similarly both to the RPE and granules in vitelliform space. The granules, which appear more orange in total AF, show a greater preponderance of the longer wavelength C3 in the vitelliform debris. The spectrum C2 peaking at 510 nm localized to all tissue including collagen in the choroid, and is a background artifact. The short wavelength spectrum C5 (purple line, peak 430 nm) localized to a single vitelliform granule, abundance 5 in Fig. 2 and is consistent with fluorophores from the neural retina, such as NADP and FAD, not LF. The low intensity spectrum C4 appears to be noise.

Conclusions :
We report 2 individual AF spectra appearing in both LF and vitelliform material in the cBest1 retina that are quite similar to human RPE spectra. The longer wavelength source predominates in the vitelliform material. These findings will enhance our understanding of the mechanism of cBest1 as we move closer to treatment of bestrophinopathies. Further research with additional tissues is warranted.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

 

Figure 1: RGB total AF image of cBest dog’s retina. Star represents vitelliform space; solid arrow shows the granules in the vitelliform space and hollow arrow shows RPE layer.

Figure 1: RGB total AF image of cBest dog’s retina. Star represents vitelliform space; solid arrow shows the granules in the vitelliform space and hollow arrow shows RPE layer.

 


Figure 2. Individual spectra recovered and corresponding abundance images.


Figure 2. Individual spectra recovered and corresponding abundance images.

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