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Patrick Pallitto, Zsolt Ablonczy, John E Donello, Yiannis Koutalos, Julia E Herrmann, Rosalie K Crouch; Fluorescence and mass spectrometric imaging of monkey (Cynomolgus) retinal pigment epithelium confirms the mismatching of lipofuscin and A2E distributions observed in human tissue. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):2352.
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Lipofuscin and its bisretinoid components have been implicated in the development of age-related macular degeneration (AMD). In cadaveric human eyes we have found a complete mismatch between lipofuscin fluorescence and the bisretinoid distributions. The goal of this project was to examine this relationship in a non-human primate model with a retinal organization similar to that of humans.
Ophthalmologically naive young (<10 yrs, N=4) and old (>10 yrs, N=4) cynomolgus monkey eyes were enucleated, dissected to yield RPE/choroid tissue layers, which were then flat-mounted on indium-tin-oxide-coated conductive slides. To compare the distributions of lipofuscin and bisretinoids, fluorescence and mass spectrometric (MALDI) imaging were carried out sequentially on the same tissues. First, fluorescence measurements were carried out in a Maestro2 imaging system (Caliper Life Sciences), the slides were then coated with 40mL of sinapinic acid in 70% ethanol for MALDI imaging. Mass spectra over the entire tissue were collected using an Autoflex II spectrometer (Bruker) in positive linear acquisition mode at 350 nm spatial resolution in the range of m/z 510-1500 with gating suppression set to m/z 490.
Lipofuscin fluorescence intensity in the cynomolgus RPE was highest in the posterior pole, tapering off toward the periphery. In the posterior pole it exhibited an emission maximum of ~600nm (λex=488nm). On the other hand, bisretinoid levels (including A2E) were highest in the periphery, showing a lack of correlation between lipofuscin fluorescence and bisretinoid distributions.
Multimodal imaging in the RPE of the cynomolgus primate model reproduced the lipofuscin and bisretinoid distributions previously obtained for human cadaveric eyes, with A2E distribution highest in the periphery and completely lacking correlation with fluorescence. We conclude that the mismatch between lipofuscin and bisretinoids, which has not been observed in mice, is due to specific anatomical features of the primate eye, such as the macula.
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