Three-dimensional properties of diseased and normal RPE cell sheets

Abstract

Purpose: Human retinal pigmented epithelium (RPE) is known to exhibit differences in spatial density, specifically, a greater cellular density in macula relative to peripheral RPE. Compromised RPE status is believed to contribute to declining photoreceptor function and numbers in age-related macular degenerative conditions (AMD). This study aims to identify patterns of morphometric parameters in normal and aged healthy and diseased human and model mouse RPE.

Methods: RPE sheets from rd10 and C57Bl/6J mice, as well as both normal and AMD human cadaveric eyes were stained with phalloidin or ZO-1 and propidium iodide to identify cell membranes and nuclei respectively. Analysis of cellular spatial point patterns were analyzed using R’s ‘spatstat’ package; ACME was used to examine three dimensional arrangement and relationship between nuclei and their containing cells; CellProfiler was used to analyze cell packing, segmentation and texture characteristics.

Results: As previously described, normal human and mouse RPE displayed regularity of structure and increased cellular density in central retina when compared to peripheral. Due to significantly different parameters, young versus old and normal versus diseased RPE sheets could be readily distinguished. Further, AMD human RPE and rd10 mouse RPE exhibited greater variability of size, shape, and other parameters, and increased spatial disorder relative to normal RPE.<br />

Conclusions: There are clear morphometric distinctions between young and old, and normal and diseased human and mouse RPE sheets. Marked spatial irregularity was observed in diseased cells, in contrast to the more regular arrangement in healthy RPE. The findings described here expand understanding of the RPE’s response to both genetic disease and the aging process.<br />