Automated segmentation of retinal pigment epithelium cells in fluorescence adaptive optics images

Rangel-Fonseca, Piero; Gómez-Vieyra, Armando; Malacara-Hernández, Daniel; Wilson, Mario; Williams, David R.; Rossi, Ethan A.

doi:10.1364/josaa.30.002595

Cited by 22 publications

(16 citation statements)

References 32 publications

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“…Semiautomated custom segmentation software 44 and photoreceptor marking software (based on a previously published algorithm 45 ) produced binary images of RPE cell boundaries and photoreceptor coordinates for each ROI. Automated RPE segmentation errors were manually corrected as previously described, 22 where segmentation within ROIs was limited to cell boundaries that could be discerned confidently by a grader.…”

Section: Methodsmentioning

confidence: 99%

Human Retinal Pigment Epithelium: In Vivo Cell Morphometry, Multispectral Autofluorescence, and Relationship to Cone Mosaic

Granger

Yang

Song

et al. 2018

Invest. Ophthalmol. Vis. Sci.

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PurposeTo characterize in vivo morphometry and multispectral autofluorescence of the retinal pigment epithelial (RPE) cell mosaic and its relationship to cone cell topography across the macula.MethodsRPE cell morphometrics were computed in regularly spaced regions of interest (ROIs) from contiguous short-wavelength autofluorescence (SWAF) and photoreceptor reflectance images collected across the macula in one eye of 10 normal participants (23–65 years) by using adaptive optics scanning light ophthalmoscopy (AOSLO). Infrared autofluorescence (IRAF) images of the RPE were collected with AOSLO in seven normal participants (22–65 years), with participant overlap, and compared to SWAF quantitatively and qualitatively.ResultsRPE cell statistics could be analyzed in 84% of SWAF ROIs. RPE cell density consistently decreased with eccentricity from the fovea (participant mean ± SD: 6026 ± 1590 cells/mm2 at fovea; 4552 ± 1370 cells/mm2 and 3757 ± 1290 cells/mm2 at 3.5 mm temporally and nasally, respectively). Mean cone-to-RPE cell ratio decreased rapidly from 16.6 at the foveal center to <5 by 1 mm. IRAF revealed cells in six of seven participants, in agreement with SWAF RPE cell size and location. Differences in cell fluorescent structure, contrast, and visibility beneath vasculature were observed between modalities.ConclusionsImprovements in AOSLO autofluorescence imaging permit efficient visualization of RPE cells with safe light exposures, allowing individual characterization of RPE cell morphometry that is variable between participants. The normative dataset and analysis of RPE cell IRAF and SWAF herein are essential for understanding microscopic characteristics of cell fluorescence and may assist in interpreting disease progression in RPE cells.

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Section: Methodsmentioning

confidence: 99%

Human Retinal Pigment Epithelium: In Vivo Cell Morphometry, Multispectral Autofluorescence, and Relationship to Cone Mosaic

Granger

Yang

Song

et al. 2018

Invest. Ophthalmol. Vis. Sci.

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“…An early version of a cell segmentation algorithm [38], developed in-house and implemented in MATLAB (MathWorks, Natick, MA, USA), was used to extract the boundaries of individual RPE cells. Briefly, the algorithm used several image processing steps, including smoothing, edge detection, edge correction and binarization to produce a binary image of the borders of RPE cells.…”

Section: Rpe Cell Segmentation and Analysismentioning

confidence: 99%

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

Rossi

Rangel-Fonseca

Parkins

et al. 2013

Biomed. Opt. Express

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Morgan and colleagues demonstrated that the RPE cell mosaic can be resolved in the living human eye non-invasively by imaging the short-wavelength autofluorescence using an adaptive optics (AO) ophthalmoscope. This method, based on the assumption that all subjects have the same longitudinal chromatic aberration (LCA) correction, has proved difficult to use in diseased eyes, and in particular those affected by age-related macular degeneration (AMD). In this work, we improve Morgan's method by accounting for chromatic aberration variations by optimizing the confocal aperture axial and transverse placement through an automated iterative maximization of image intensity. The increase in image intensity after algorithmic aperture placement varied depending upon patient and aperture position prior to optimization but increases as large as a factor of 10 were observed. When using a confocal aperture of 3.4 Airy disks in diameter, images were obtained using retinal radiant exposures of less than 2.44 J/cm 2 , which is ~22 times below the current ANSI maximum permissible exposure. RPE cell morphologies that were strikingly similar to those seen in postmortem histological studies were observed in AMD eyes, even in areas where the pattern of fluorescence appeared normal in commercial fundus autofluorescence (FAF) images. This new method can be used to study RPE morphology in AMD and other diseases, providing a powerful tool for understanding disease pathogenesis and progression, and offering a new means to assess the efficacy of treatments designed to restore RPE health. L. Dunaief, J. Z. Baffi, and J. Ambati, "DICER1 deficit induces Alu RNA toxicity in age-related macular degeneration," Nature 471(7338), 325-330 (2011).

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“…Deployment of adaptive optics (AO) in fundus cameras, 14 cSLO, 15 and OCT 16 instruments has further improved optical resolution, thus permitting imaging of single cells, such as rods, foveal cones, 17 , 18 RPE cells, 19 and blood cells in small retinal vessels. 20 , 21 This is of interest for monitoring longitudinal changes in diseases, such as retinitis pigmentosa, 22 , 23 retinal dystrophy, 24 – 26 age-related macular degeneration, 27 , 28 and diabetic retinopathies, 29 – 31 especially since cellular scale imaging can permit semiautomated quantification of the density and spacing of these cells 32 – 37 as well as AO-assisted microperimetry, 38 , 39 …”

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confidence: 99%

Two-Photon Autofluorescence Imaging Reveals Cellular Structures Throughout the Retina of the Living Primate Eye

Sharma

Williams²,

Palczewska

et al. 2016

Invest. Ophthalmol. Vis. Sci.

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PurposeAlthough extrinsic fluorophores can be introduced to label specific cell types in the retina, endogenous fluorophores, such as NAD(P)H, FAD, collagen, and others, are present in all retinal layers. These molecules are a potential source of optical contrast and can enable noninvasive visualization of all cellular layers. We used a two-photon fluorescence adaptive optics scanning light ophthalmoscope (TPF-AOSLO) to explore the native autofluorescence of various cell classes spanning several layers in the unlabeled retina of a living primate eye.MethodsThree macaques were imaged on separate occasions using a custom TPF-AOSLO. Two-photon fluorescence was evoked by pulsed light at 730 and 920 nm excitation wavelengths, while fluorescence emission was collected in the visible range from several retinal layers and different locations. Backscattered light was recorded simultaneously in confocal modality and images were postprocessed to remove eye motion.ResultsAll retinal layers yielded two-photon signals and the heterogeneous distribution of fluorophores provided optical contrast. Several structural features were observed, such as autofluorescence from vessel walls, Müller cell processes in the nerve fibers, mosaics of cells in the ganglion cell and other nuclear layers of the inner retina, as well as photoreceptor and RPE layers in the outer retina.ConclusionsThis in vivo survey of two-photon autofluorescence throughout the primate retina demonstrates a wider variety of structural detail in the living eye than is available through conventional imaging methods, and broadens the use of two-photon imaging of normal and diseased eyes.

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Automated segmentation of retinal pigment epithelium cells in fluorescence adaptive optics images

Cited by 22 publications

References 32 publications

Human Retinal Pigment Epithelium: In Vivo Cell Morphometry, Multispectral Autofluorescence, and Relationship to Cone Mosaic

Human Retinal Pigment Epithelium: In Vivo Cell Morphometry, Multispectral Autofluorescence, and Relationship to Cone Mosaic

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

Two-Photon Autofluorescence Imaging Reveals Cellular Structures Throughout the Retina of the Living Primate Eye

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