Histologic Basis of Variations in Retinal Pigment Epithelium Autofluorescence in Eyes with Geographic Atrophy

Abstract

Purpose Lipofuscin contained in the retinal pigment epithelium (RPE) is the main source of fundus auto-fluorescence (FAF), the target of an imaging method useful for estimating the progression of geographic atrophy (GA) in clinical trials. To establish a cellular basis for hyperfluorescent GA border zones, histologic autofluorescence (HAF) was measured at defined stages of RPE pathologic progression. Design Experimental study. Participants and Controls Ten GA donor eyes (mean age ± standard deviation, 87.1±4.0 years) and 3 age-matched control eyes (mean age ± standard deviation, 84.0±7.2 years) without GA. Methods Ten–micrometer-thick sections were divided into zones of RPE morphologic features according to an 8-point scale. Any HAF excited by 488 nm light was imaged by laser confocal microscopy. The HAF intensity summed along vertical lines perpendicular to Bruch’s membrane at 0.2-μm intervals served as a surrogate for FAF. Intensity profiles in 151 zones were normalized to grade 0 at a standard reference location in each eye. Cross-sectional area, mean, and sum autofluorescence for individual RPE cells were measured (cellular autofluorescence [CAF]). Main Outcome Measures Statistically significant differences in intensity and localization of HAF and CAF at defined stages of RPE morphologic progression for GA and control eyes. Results The RPE morphologic features were most abnormal (cell rounding, sloughing, and layering; grade 2) and HAF intensity profiles were highest and most variable immediately adjacent to atrophic areas. Peaks in HAF intensity frequently were associated with vertically superimposed cells. The HAF value that optimally separated reactive RPE was 0.66 standard deviations more than the mean for uninvolved RPE and was associated with a sensitivity of 75.8% and a specificity of 76.3%. When variable cell area was accounted for, neither mean nor sum CAF differed significantly among the RPE pathologic grades. Conclusions Areas with advanced RPE alterations are most likely to exhibit clinically recognizable patterns of elevated FAF around GA, but may not predict cells about to die, because of vertically superimposed cells and cellular fragments. These data do not support a role for lipofuscin-related cell death and call into question the rationale of treatments targeting lipofuscin.

In vivo imaging of retinal pigment epithelium cells in age related macular degeneration

“…This is a familiar problem for those using in vivo imaging methods. Many of the RPE cell morphologies seen in the later stages of AMD, such as multilayered cells and heaping and sloughing of cells could disrupt our ability to see cellular structure [5]. The AF imaging technique in FAOSLO capitalizes on the fact that the normal RPE is arranged in a flat monolayer.…”

“…Histological studies of postmortem eyes have shown that substantial changes occur in RPE cell mosaic morphology in AMD [4,5]. These changes may precede and/or accompany RPE cell death and the degeneration of overlying photoreceptors [4,6].…”

“…Histological studies have shown that RPE mosaic morphology is drastically altered in AMD [4,5], but these studies of post mortem eyes are limited in that they only reveal a single time point and cannot compare changes in RPE morphology to other measures, such as cSLO FAF imaging, SD-OCT, or AO reflectance imaging of the photoreceptor cell mosaic. In vivo cellular resolution imaging has the potential to identify early disease changes in the RPE cell mosaic, before the heterogeneous patterns of hyper AF and hypo AF seen in conventional FAF imaging arise.…”

Hyperreflective Pyramidal Structures on Optical Coherence Tomography in Geographic Atrophy Areas

“…However, it is notable that no hyperautofluorescence was noted, suggesting that these HPS are not mainly made of lipofuscin. 38 Differently form reticular pseudodrusen, which are characterized by progressive reabsorption and migration of the hyperreflective material in the inner retinal layers before the development of GA, HPS appear as heterogeneous hyperreflective structure accumulating in the context of GA. 32,33 Alternatively, HPS could be compared with soft drusen. Soft drusen are yellowish with indistinct margins, bigger than hard drusen, confined to the macula, precursor to choroidal neovascularization, and consisting in focal accentuations of basal linear deposits, specific for AMD.…”

Increased Fundus Autofluorescence and Progression of Geographic Atrophy Secondary to Age-Related Macular Degeneration: The GAIN Study

“…Inquiries to Marc Biarnés, C/ Vilana 12, Office 90, Institut de la màcula i de la retina, Centro Médico Teknon, Barcelona, Spain; e-mail: biarnes@ oo.upc.edu Nonetheless, the role of lipofuscin in GA pathogenesis remains controversial. Most studies 6,[8][9][10] suggest that it has deleterious effects on RPE that may lead to cell death and GA progression, while other clinical 11,12 and basic research findings 13,14 do not support this hypothesis. Indeed, a visual cycle modulator did not meet the primary efficacy endpoint in a phase II clinical trial.…”