Fluorescence Lifetime Imaging Ophthalmoscopy (FLIO) of Macular Pigment

Abstract

PurposeTo describe different patterns of macular pigment (MP) seen in fluorescence lifetime imaging ophthalmoscopy (FLIO) and to analyze ex vivo fluorescence characteristics of carotenoids.MethodsA total of 31 eyes of young healthy subjects, 4 eyes from patients with albinism, 36 eyes with macular telangiectasia type 2 (MacTel), 24 eyes with retinitis pigmentosa, and 1 eye with a macular hole were included in this clinic-based, cross-sectional study. All subjects underwent Heidelberg Engineering FLIO and MP measurements (dual-wavelength autofluorescence). Fundus autofluorescence (FAF) lifetimes of a 30° retinal field were detected in two spectral channels (SSC: 498–560 nm; LSC: 560–720 nm), and amplitude-weighted mean fluorescence lifetimes (τm) were calculated. Additionally, autofluorescence lifetimes of known dilutions of lutein and zeaxanthin were measured in a cuvette in free- and protein-associated states.ResultsMP shows a significant inverse correlation to foveal FAF lifetimes measured with FLIO (SSC: r = −0.608; P < 0.001). Different distribution patterns can be assigned to specific disease-related changes. Two patients with albinism, who did not have MP, were found to be missing short FAF lifetimes. In solvent, lutein and zeaxanthin show very short autofluorescence lifetimes (∼50–60 ps; SSC), as do their respective binding proteins (∼40–50 ps; SSC). When combining carotenoids with their specific binding proteins, the decay times shift to longer means (∼70–90 ps; SSC).ConclusionsThis study expands upon previous findings of an impact of MP on short FAF lifetimes by describing ex vivo autofluorescence lifetimes of carotenoids and different in vivo autofluorescence patterns that can be associated with certain diseases.

Bleaching effects and fluorescence lifetime imaging ophthalmoscopy

“…One may argue that the macular pigment, retinal carotenoids accumulating in the fovea, may have an impact. However, these pigments are distributed in an area of 1 mm in diameter at the fovea due to the presence of macular pigment binding proteins, and their fluorescence emission is mostly found in the SSC [24,25,39]. The SSC shows no significant changes in the fluorescence lifetime with bleaching.…”

“…More than 10 known endogenous fluorophores exist in the human retina [18,24,25] but the FLIO technique is currently limited to the simultaneous detection of at most three spatially resolved fluorescence lifetimes. The approximated fluorescence lifetime in FLIO typically does not correspond to a single endogenous fluorophore, as it comprises fluorescence information from multiple retinal fluorophores.…”

Fluorescence Lifetime Imaging Ophthalmoscopy (FLIO)

“…A different FLIO study investigated fundus autofluorescence lifetimes in patients with albinism [50]. These patients usually do not have a foveal depression, which is often described as foveal hypoplasia.…”

“…During the last 6 years, the diagnostic potential of this device referred as "Spectralis FLIO" was systematically explored in clinical studies in Bern, Jena and Salt Lake City focusing on different pathologies. The results can be found in a series of interesting and seminal publications [35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52][53]. Some selected highlights of the clinical results are summarized in the Sects.…”

“…Although the macula appears dark in standard intensity autofluorescence images due to the strong absorption of the blue light by the macular pigment (MP), it has been shown with FLIO [37,50] that the MP emits a weak but measurable fluorescence with very short lifetimes. The autofluorescence lifetimes of the carotenoids lutein and zeaxanthin were determined using ex vivo FLIO imaging.…”