Author: Leon von der Emde (Germany)
Co-authors: Marc Vaisband, Jan Hasenauer, Christine A. Curcio, Kenneth R Sloan, Thomas Ach
Purpose
Altered retinal pigment epithelium (RPE) is a hallmark of age-related macular degeneration (AMD) [PMID: 31173079]. Morphology and function are closely intertwined and pathological RPE phenotypes may not be able to preserve important functions for outer retinal health [PMID: 31714897]. In this study, we quantified differences in RPE morphology between donors with unaffected macula and AMD. Further, we characterized differences in RPE morphology dependent on donor´s age and the respective retinal location.
Setting/Venue
Globes were collected from the Advancing Sight Network Birmingham, AL, USA. Microscopic imaging was achieved at the Leibniz Institute of Photonic Technology, Jena. Cell analyses were carried out at the University Hospital of Bonn. Data analyses were performed at the Department of Ophthalmology, Birmingham, AL, and statistical analyses were performed at the Haussdorff Center for Mathematics, Bonn.
Methods
Seven AMD-affected human RPE flatmounts (early: 3, late dry: 1; neovascular: 3) and 15 human RPE flatmounts from all two age-groups (≤51 years, n = 8; >80 years, n = 7) were imaged at three locations (fovea, perifovea, near periphery) using a laser scanning confocal fluorescence microscope (excitation 488 nm; emission: 500-750 nm; z-stack step size 390 nm). Using FIJI, RPE cell bodies were manually marked and extracted. Shape descriptors for each cell (e.g., cell area, length of the major and minor axes, eccentricity, solidity, perimeter, aspect ratio, form factor) were calculated with the help of customized software and a principal component analyses (PCA) was calculated.
Results
Of 4404 cells analysed (fovea 927; perifovea: 1430; near-periphery 2047), 626 were from AMD affected and 3778 were from healthy donors. The first principal component was able to explain 47% of variability (first two: 79%). AMD RPE cells were greatly enlarged with altered morphology and PCA identified area as the most important followed by the perimeter and the form factor{ratio between area of the block and the area of the circumscribed hull} as the shape descriptors that were strongly discriminatory. Differences between AMD affected and unremarkable maculae were most pronounced at the fovea.
Conlusions
We were able to accurately quantify RPE morphology with the presence of AMD. Enlarged and deformed cells in AMD are indicative of a dysfunctional RPE. Interestingly, RPE alterations were most pronounced at the fovea, an area typically affected by accumulation of soft drusen in AMD as well as preservation of cone-mediated visual acuity. The results of this study may help guide the interpretation of RPE morphology in in vivo studies utilizing adaptive optics-assisted imaging. Further, the results of this study could be applied to characterize stem cell derived RPE cell cultures.
Financial Disclosure
Von der Emde, Vaisband, Hasenauer, Sloan: none. Curcio: Genentech (f), Heidelberg Engineering (F), Rgeneron (F), MacRegen Inc (I) Ach: Roche (C), Novartis (F,R), MacRegen Inc (I) Funding: NIH/NEI 1R01EY027948-01 (TA, CAC), R01EY029595 (CAC), R01EY024738 (CAC)
Comments
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