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Danielle J. Harper, Marco Augustin, Kornelia Schutzenberger, Martin Glösmann, Thomas Konegger, Pablo Eugui, Antonia Lichtenegger, Conrad W. Merkle, Christoph K Hitzenberger, Bernhard Baumann; White light hyperspectral optical coherence tomography reveals melanin distribution in the retinal pigment epithelium. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4742. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To demonstrate the ability of white light hyperspectral optical coherence tomography (OCT) to perform in vivo localization of melanin in the retinal pigment epithelium (RPE) of rats and mice.
The retinas of non-pigmented Oncins France Strain A (OFA) rats (N = 3, 13 weeks old), Long Evans (LE) rats (N = 3, 13 weeks old), Brown Norway (BN) rats (N = 3, 18 weeks old) and mice from a C57BL/6 background (N = 3, 66 weeks old) were imaged with a white light spectral domain OCT system which was designed in-house. All animals were anesthetized using isoflurane and pupils were dilated using tropicamide. Artificial tear drops were applied throughout the experiments to keep the eyes moist. The backscattered signal at all wavelengths ranging from 400 – 700 nm was simultaneously acquired. By sub-sampling the wavelength spectra in post-processing, a hyperspectral stack of 11 wavelength ranges was created for each image. In addition to the standard intensity-based images, spectral variance maps were also created. Segmentation of the RPE was performed in all 6 eyes of each strain (5 eyes for the mouse due to motion in one dataset) and the mean of the spectral variance was calculated. A line profile of the variance of the RPE was also created to reveal the spatial distribution of the melanin.
The spectral variance maps highlighted the nerve fiber layer, the end tips of some photoreceptors, the RPE and the choroid. Due to the small particulate nature of melanin, it is expected from Mie theory that the melanin will be responsible for this variation in the RPE and choroid. Segmentation of the RPE indicated that the relationship of the melanin concentration in the strains is: mouse>LE>BN>OFA (Fig. 1F). These results were found to be statistically significant (one-way ANOVA test, F (3, 1000) = 107.5167, p < 0.05) followed by a post hoc Tukey HSD test which revealed statistical significance between all groups (p < 0.05 for LE vs mouse, all others p < 0.01). The RPE line profile variance can also indicate where the RPE cell nuclei lie due to their uniform backscattering coefficient as a function of wavelength, in contrast to that of the melanin (Fig. 2).
By analyzing wavelength dependent variance in the visible light range with OCT, melanin can be localized in vivo in the retinal pigment epithelium of mice and rats.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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