Analysis of Autofluorescent retinal images and measurement of atrophic lesion growth in Stargardt disease
Introduction
The accumulation of toxic lipofuscin fluorophores in the retinal pigment epithelium (RPE) represents a common pathological pathway leading to photoreceptor cell loss in a number of disease entities (Bui et al., 2006, Dorey et al., 1989, Eldred and Lasky, 1993, Feeney-Burns et al., 1980, Okubo et al., 1999, Sparrow and Boulton, 2005, Weiter et al., 1986) (for a review, see Travis et al., 2007). For example, the lipofuscin fluorophores, A2E and A2PE-H2, are dramatically elevated in the RPE of postmortem samples taken from patients with recessive Stargardt Disease (STGD) and in the abcr−/− mouse model of STGD (Mata et al., 2000, Weng et al., 1999). The cytotoxicity of A2E in RPE cells is well known and is assumed to occur through a series of well-defined biochemical events that ultimately lead to RPE and overlying photoreceptor cell death (De and Sakmar, 2002, Finnemann et al., 2002, Sparrow et al., 1999, Sparrow et al., 2003, Suter et al., 2000, Travis et al., 2007).
Scanning laser ophthalmoscopy (SLO) provides an effective means for characterizing the topographical distribution of lipofuscin fluorophores noninvasively in human subjects. This imaging technique capitalizes on a fundamental property of retinal fluorophores in that they emit light in the spectral range 500–700 nm when excited by short-wavelength light (<490 nm). The topographical distribution and density of the emitted light define regions where the lipofuscin fluorophores have accumulated above background levels (hyperfluorescence, HF). More importantly, intact RPE and overlying photoreceptor cells are required for the production of lipofuscin fluorophores. Thus, in the absence of other factors that might reduce fundus autofluorescence (e.g. hemorrhages), regions with fluorescent signal well below background levels (hypofluorescence) identify regions where RPE and photoreceptor cells are presumed to have been irretrievably lost. Thus, AF imagery has become a powerful tool to identify regions of retinal atrophy. Further, the abnormal patterns of increased fundus fluorescence that surround atrophic lesions (ALs) may serve as a prognostic marker for the future progression and expansion of the AL (Holz et al., 2001, Holz et al., 2007, Schmitz-Valckenberg et al., 2006, Schmitz-Valckenberg et al., 2009). However, the validity of this sequence of events has recently been challenged (Hwang et al., 2006, Smith et al., 2009).
In this study, a novel Edge-Flow-Driven image segmentation algorithm (UCSB Vision Research Lab, vision.ece.ucsb.edu) is described and used to measure AL area in STGD. The algorithm is embedded in a series of automated image processing steps that allow rapid analysis of serial images. We test the hypothesis that this methodology is sufficiently sensitive to monitor progressive change of AL size in STGD so that it might be used as an outcome measure in a clinical trial. In addition, we evaluated the association of regional loss of RPE and overlying photoreceptors, particularly in areas where rod and cone photoreceptor densities are highest, with measures of rod- and cone-mediated retinal function as determined by standardized electroretinography (ERG). We test the hypothesis that large central ALs that are accompanied by subnormal ERG responses may be interpreted incorrectly as indicating panretinal dysfunction.
Section snippets
Subjects
Fifty-two STGD subjects (mean age = 41.0 ± 16.6 years, range 9–78 yrs) at varying stages of disease participated in a prospective study. All affected individuals were confirmed as having recessive (or sporadic) STGD or STGD-like disease based on a clinical exam by at least two retina specialists based on the presence of bilateral macular or posterior pole disease consisting of subretinal flecks, RPE disturbances and/or geographic atrophy and a careful family history. Based on fundus appearance
Measurement variability
A series of 5–7 AF images were obtained from each of five STGD patients, all recorded on the same day but with different camera sensitivities to vary edge gradients as described in the Methods section. In addition, the patient’s head was repositioned between trials to produce changes in image orientation and magnification. Across all tested eyes, the median variation (data not shown) for the series of images was 2.9% (range = 2.2%–7.7%). (A t-test comparing the variability between eyes was not
Discussion
The main goal of this study was to evaluate progressive change in AL size in patients with recessive STGD. To this end, a novel automated methodology was developed to process AF retinal images obtained with scanning laser ophthalmoscopy. The methodology required linking together several image analysis programs that ran sequentially but which automatically accomplish the task of registering serial images, maximizing signal-to-noise ratios, and segmenting the AL using an Edge-Flow-Driven
Commercial relationships
None.
Acknowledgements
We are grateful to the Foundation Fighting Blindness and the Sarkaria Family Fund for their generous support of this research. We also thank Kristin Lipka and David Le Beck for their assistance with recording the Autofluorescent images, and Dr. Carolina M. Ortube, Ariadna Martinez, and Arturo Garcia for their help with recruiting patients.
Grant Support: Foundation Fighting Blindness and the Sarkaria Family Fund for Macular Dystrophy Research.
References (43)
- et al.
Characterization of native retinal fluorophores involved in biosynthesis of A2E and lipofuscin-associated retinopathies
J. Biol. Chem.
(2006) - et al.
Lipofuscin of human retinal pigment epithelium
Am. J. Ophthalmol
(1980) - et al.
Electroretinographic evidence for altered phototransduction gain and slowed recovery from photobleaches in albino mice with a MET450 variant in RPE65
Exp. Eye Res.
(2003) - et al.
RPE lipofuscin and its role in retinal pathobiology
Exp. Eye Res.
(2005) - et al.
A2E-epoxides damage DNA in retinal pigment epithelial cells. Vitamin E and other antioxidants inhibit A2E-epoxide formation
J. Biol. Chem.
(2003) - et al.
Age-related macular degeneration. The lipofusion component N-retinyl-N-retinylidene ethanolamine detaches proapoptotic proteins from mitochondria and induces apoptosis in mammalian retinal pigment epithelial cells
J. Biol. Chem.
(2000) - et al.
Insights into the function of Rim protein in photoreceptors and etiology of Stargardt’s disease from the phenotype in abcr knockout mice
Cell
(1999) - et al.
Interaction of A2E with model membranes. Implications to the pathogenesis of age-related macular degeneration
J. Gen. Physiol.
(2002) - et al.
Automated analysis of digital fundus autofluorescence images of geographic atrophy in advanced age-related macular degeneration using confocal scanning laser ophthalmoscopy (cSLO)
BMC. Ophthalmol.
(2005) - et al.
Cell loss in the aging retina. Relationship to lipofuscin accumulation and macular degeneration
Invest. Ophthalmol. Vis. Sci.
(1989)
Pattern Classification and Scene Analysis. Pattern Classification and Scene Analysis
Retinal age pigments generated by self-assembling lysosomotropic detergents
Nature
The lipofuscin component A2E selectively inhibits phagolysosomal degradation of photoreceptor phospholipid by the retinal pigment epithelium
Proc. Natl. Acad. Sci. U.S.A.
Multiscale vessel enhancement filtering
Variational image segmentation using boundary functions
IEEE Trans. Image Process
Fundus autofluorescence and development of geographic atrophy in age-related macular degeneration
Invest. Ophthalmol. Vis. Sci.
Progression of geographic atrophy and impact of fundus autofluorescence patterns in age-related macular degeneration
Am. J. Ophthalmol.
Predictive value of fundus autofluorescence for development of geographic atrophy in age-related macular degeneration
Invest. Ophthalmol. Vis. Sci.
Anatomical pathways for color vision in the human retina
Vis. Neurosci.
Correction of the disease phenotype in the mouse model of Stargardt disease by lentiviral gene therapy
Gene Ther.
Cited by (51)
Atrophy Expansion Rates in Stargardt Disease Using Ultra-Widefield Fundus Autofluorescence
2021, Ophthalmology ScienceClinical spectrum, genetic complexity and therapeutic approaches for retinal disease caused by ABCA4 mutations
2020, Progress in Retinal and Eye ResearchAutomatic image analyser to assess retinal vessel calibre (ALTAIR). A new tool to evaluate the thickness, area and length of the vessels of the retina
2020, International Journal of Medical InformaticsQuantifying the Rate of Ellipsoid Zone Loss in Stargardt Disease
2018, American Journal of OphthalmologyCitation Excerpt :The high intragrader and intergrader ICC suggests that highly precise measurements can be obtained using the area of EZ loss, and indicates that this method can provide a high degree of confidence in defining disease progression. Prior studies of STGD progression based on area of RPE atrophy range from 0.28 mm2/year to 1.58 mm2/year.27,28,30,31,44 Testa and associates estimated the rate of RPE loss as seen on OCT in a heterogeneous group of STGD patients using an automated segmentation program to be around 0.28 mm2/year.44