Short-wavelength Fundus Autofluorescence Imaging Research Articles

Genotypic and Phenotypic Characterization of a Cohort of Patients Affected by Rod Cyclic Nucleotide Channel-Associated Retinitis Pigmentosa.

Retinitis pigmentosa (RP), a heterogeneous inherited retinal disorder causing gradual vision loss, affects over 1 million people worldwide. Pathogenic variants in CNGA1 and CNGB1 genes, respectively, accounting for 1% and 4% of cases, impact the cyclic nucleotide-gated channel in rod photoreceptor cells. The aim of this study was to describe and compare genotypic and clinical characteristics of a cohort of patients with CNGA1- or CNGB1-related RP and to explore potential genotype-phenotype correlations. The following data from patients with CNGA1- or CNGB1-related RP, followed in five Italian inherited retinal degenerations services, were retrospectively collected: genetic variants in CNGA1 and CNGB1, best-corrected visual acuity (BCVA), ellipsoid zone (EZ) width, fundus photographs, and short-wavelength fundus autofluorescence (SW-AF) images. Comparisons and correlation analyses were performed by first dividing the cohort in two groups according to the gene responsible for the disease (CNGA1 and CNGB1 groups). In parallel, the whole cohort of RP patients was divided into two other groups, according to the expected impact of the variants at protein level (low and high group). In total, 29 patients were recruited, 11 with CNGA1- and 18 with CNGB1-related RP. In both CNGA1 and CNGB1, 5 novel variants in CNGA1 and 5 in CNGB1 were found. BCVA was comparable between CNGA1 and CNGB1 groups, as well as between low and high groups. CNGA1 group had a larger mean EZ width compared to CNGB1 group, albeit not statistically significant, while EZ width did not differ between low and high groups A statistically significant correlation between EZ width and BCVA as well as between EZ width and age were observed in the whole cohort of RP patients. Fundus photographs of all patients in the cohort showed classic RP pattern, and in SW-AF images an hyperautofluorescent ring was observed in 14/21 patients. Rod CNG channel-associated RP was demonstrated to be a slowly progressive disease in both CNGA1- and CNGB1-related forms, making it an ideal candidate for gene augmentation therapies.

Pentosan polysulfate maculopathy: a brief primer for general practitioners, ophthalmologists, optometrists and urologists.

Pentosan polysulfate maculopathy: a brief primer for general practitioners, ophthalmologists, optometrists and urologists.

CLINICAL ASSOCIATIONS AND PROGNOSTIC IMPLICATIONS OF REPAIR TISSUE PROLIFERATION IN EYES WITH RETINAL PIGMENT EPITHELIUM TEARS.

To investigate demographic and clinical factors influencing the longitudinal changes of retinal pigment epithelium (RPE) dehiscence area after RPE tears, including the presence of RPE tear-associated repair proliferation (TARP), and identify factors associated with TARP development over follow-up. Retrospective, single-center, observational cohort study of patients with a history of macular neovascularization and RPE tear. The area of RPE dehiscence was measured on repeated short-wavelength fundus autofluorescence imaging. Associations between covariates and RPE dehiscence areas were tested with multivariable linear mixed models. Associations between TARP development and clinical variables were investigated with Cox regression models. Factors associated with visual acuity changing rates were explored with linear mixed models. Thirty-seven eyes of 36 patients were included in this study and followed for a median time of 18 months. Tear-associated repair proliferation was identified in 27 eyes (73%). The median time for TARP detection was 112 days; none of the investigated factors was significantly associated with TARP occurrence. The presence of TARP (estimate: -0.042 mm2/month; P = 0.001) and female gender (estimate: -0.035 mm2/month; P = 0.006) were associated with slower rates of RPE dehiscence enlargement over time. Faster rates of visual improvement were observed in eyes with TARP compared with those without TARP (estimate = -0.010 logarithm of the minimum angle of resolution/month if TARP was present; P = 0.008). Retinal pigment epithelium tear repair with TARP and female gender were associated with slower RPE degeneration after RPE tears. The presence of TARP was associated better visual prognosis. Additional research on factors promoting TARP development may have therapeutic and prognostic implications.

Stage-dependent choriocapillaris impairment in Best vitelliform macular dystrophy characterized by optical coherence tomography angiography

Characterization of vascular impairment in Best vitelliform macular dystrophy (BVMD) is essential for the development of treatment modalities and therapy trials. As such, we seek to characterize the choriocapillaris (CC) at each stage of the disease process in 22 patients (44 eyes) with a diagnosis of BVMD confirmed by genetic sequencing. We utilize optical coherence tomography angiography (OCTA) images to characterize the CC and correlate our findings to the status of the retinal pigment epithelium (RPE) as observed on short-wavelength fundus autofluorescence (SW-AF) images. We observed that in the vitelliruptive stage, the CC appeared as bright and granular in the area where the vitelliform lesion was present. In the atrophic stage, varying degrees of CC atrophy were observed within the lesion area, with the regions of CC atrophy appearing as hypoautofluorescent on SW-AF images. Our results suggest that the CC impairment observed in the vitelliruptive stage of BVMD progressively culminates in the CC atrophy observed at the atrophic stage. As such, OCTA imaging can be used to characterize CC impairment in BVMD patients as part of diagnosis and tracking of disease progression. Our findings suggest that the best window of opportunity for therapeutic approaches is before the atrophic stage, as it is during this stage that CC atrophy is observed.

Quantitative Fundus Autofluorescence in HCQ Retinopathy.

PurposeTo increase our understanding of the mechanisms underlying hydroxychloroquine (HCQ) retinopathy, analyses by quantitative fundus autofluorescence (qAF) and near-infrared fundus autofluorescence (NIR-AF) were compared to results obtained by recommended screening tests.MethodsThirty-one patients (28 females, 3 males) were evaluated with standard automated perimetry and spectral domain optical coherence tomography (SD-OCT); 28 also had multifocal electroretinography (mfERG). Measurement of short-wavelength fundus autofluorescence (SW-AF) by qAF involved the use of an internal fluorescent reference and intensity measurements in eight concentric segments at 7° to 9° eccentricity. For semiquantitative analysis of NIR-AF, intensities were acquired along a vertical axis through the fovea.ResultsFour of 15 high-dose (total dose >1000 g, daily dose >5.0 mg/kg) patients and one of 16 low-dose (total dose <1000 g, daily dose 4.4 mg/kg) patients were diagnosed with HCQ-associated retinopathy based on abnormal 10-2 visual fields, SD-OCT, and SW-AF imaging. Three of the high-dose patients also had abnormal mfERG results. Of the five patients exhibiting retinopathy, two had qAF color-coded images revealing higher intensities inferior, nasal, and lateral to the fovea. The abnormal visual fields also exhibited superior-inferior differences. Mean NIR-AF gray-level intensities were increased in four high-dose patients with no evidence of retinopathy. In two patients with retinopathy, NIR-AF intensity within the parafovea was below the normal range. One high-dose patient (6.25 mg/kg) had only abnormal mfERG results.ConclusionsThese findings indicate that screening for HCQ retinopathy should take into consideration superior-inferior differences in susceptibility to HCQ retinopathy.

Factors Influencing Retinal Pigment Epithelium-Atrophy Progression Rate in Stargardt Disease.

PurposeTo evaluate demographic, clinical, imaging, and genetic factors associated with retinal pigment epithelium enlargement in Stargardt disease (STGD1) and to measure the agreement between short-wavelength fundus autofluorescence (SW-FAF) and near-infrared fundus autofluorescence (NIR-FAF).MethodsRetrospective cohort study of patients with STGD1 with ≥2 gradable SW-FAF images. RPE-atrophy areas were measured on SW-FAF and NIR-FAF at each visit and regressed against time to obtain the rate of RPE-atrophy enlargement. Agreement between SW-FAF and NIR-FAF with regards to baseline atrophic areas and rates of enlargement was evaluated. Baseline factors predictive of faster SW-FAF RPE-atrophy enlargement were investigated with linear mixed models.ResultsFifty-four eyes of 28 patients (median age: 45 years; 13 males) were included. SW-FAF and NIR-FAF agreed well for slow rates of RPE-atrophy progression, but agreement decreased as the rate increased. Median (interquartile range [IQR]) rate of RPE-atrophy expansion was 0.18 (0.10–0.85) mm2/year on SW-FAF and 0.24 (0.08–0.33) mm2/year on NIR-FAF. Larger baseline RPE-atrophy area (estimate: 0.057 mm2/year, P < 0.001), worse visual acuity (0.305 mm2/year, P = 0.005), multifocal disease (0.401 mm2/year, P = 0.02), and SW-FAF pattern (0.534 mm2/year, P =0 .03) were associated with a faster rate of progression (predictive R2: 0.65).ConclusionsSW-FAF and NIR-FAF are not interchangeable in the evaluation of RPE-atrophy enlargement, and both imaging modalities may be required for optimal detection of disease progression. A multivariable model based on baseline clinical and imaging information may identify patients at higher risk of fast disease progression.Translational RelevanceThe knowledge of the agreement of different FAF modalities, the estimated rates of RPE-atrophy enlargement, and factors predictive of faster anatomic decay in STGD1 may allow tailored clinical management and better clinical trials design.

Disease asymmetry and hyperautofluorescent ring shape in retinitis pigmentosa patients

Retinitis pigmentosa (RP) is described as a bilateral disease with inter-eye symmetry that presents on short-wavelength fundus autofluorescence (SW-AF) imaging with hyperautofluorescent (hyperAF) rings with an ellipsoid shape and regular borders. Nevertheless, both asymmetry and irregular ring morphologies are also observed. In this retrospective study of 168 RP patients, we characterize the degree of inter-eye asymmetry and frequency of irregular hyperAF ring morphologies according to mode of inheritance and disease-causing gene by using SW-AF imaging and spectral-domain optical coherence tomography (SD-OCT) scans. We observed that from 336 eyes, 290 (86%) presented with regular hyperAF rings and 46 (14%) presented with irregular shapes. From the 168 patients, 23 (14%) presented with asymmetric disease, with 16 (70%) of these patients also presenting with irregular ring shapes. Patients with autosomal dominant RP (adRP) had the highest proportion of irregular ring shapes (21%) and disease asymmetry (23%) in comparison to other modes of inheritance. Furthermore, both RP1 and RHO-adRP had the highest proportions of both disease asymmetry and irregular ring morphology. Our results suggest that in patients presenting with either irregular ring shapes or asymmetric disease, emphasis should be placed in targeted gene sequencing of genes known to cause adRP, such as RHO and RP1.

Functional Relevance and Structural Correlates of Near Infrared and Short Wavelength Fundus Autofluorescence Imaging in ABCA4-Related Retinopathy.

PurposeTo evaluate the functional relevance and structural correlates of autofluorescence (AF) alterations under short-wavelength (SW) and near-infrared (NIR) excitation light in ABCA4-related retinopathy.MethodsIn this prospective, cross-sectional case series, 88 eyes of 44 patients with ABCA4-related retinopathy (mean age, 37.6 years; range, 9–77 years) underwent SW-AF and NIR-AF imaging. The AF images were graded for disease characteristic patterns by two independent readers and correlated with alterations in optical coherence tomography (OCT) and impairment of retinal sensitivity along a foveo-papillary line assessed by fundus-controlled microperimetry.ResultsA centrifugal sequence of AF patterns from atrophic lesions to homogeneous background was found for both AF modalities. The eccentricity of each AF pattern in NIR-AF was larger compared to those in SW-AF (P < 0.001). Increasing eccentricity of each pattern correlated with increasing retinal sensitivity. The distant border of the zone of hyperfluorescent flecks in SW-AF and hypoautofluorescent flecks in NIR-AF correlated with the margins of the ellipsoid zone loss in OCT (r = 0.979 and r = 0.971, P < 0.001). The expansion of hypofluorescent flecks in SW-AF was associated with the boundaries of external limiting membrane loss (r = 0.933, P < 0.001).ConclusionsSW-AF and NIR-AF revealed a characteristic sequence of AF patterns that correlated with functional and structural alterations, suggesting different stages in disease progression.Translational RelevanceAlterations in NIR-AF exceeded those in SW-AF images, substantiating the hypothesis of different AF origins and suggesting NIR-AF as surrogate marker for early disease-related changes.

Multimodal Imaging in Best Vitelliform Macular Dystrophy.

PurposeIn patients diagnosed with Best vitelliform macular dystrophy (BVMD), quantitative fundus autofluorescence (qAF), near-infrared fundus autofluorescence (NIR-AF), and spectral-domain optical coherence tomography (SD-OCT) were used to elucidate pathogenic mechanisms.MethodsFourteen patients heterozygous for BEST1 mutations were recruited. qAF was analyzed using short-wavelength fundus autofluorescence (SW-AF) images. Mean gray levels (GL) were determined in nonlesion areas (7 to 9° eccentricity) and adjusted by GL measured in an internal fluorescent reference. NIR-AF images (787 nm; sensitivity of 96) were captured and saved in non-normalized mode. Horizontal SD-OCT images also were acquired and BVMD was staged according to the OCT findings.ResultsIn the pre-vitelliform stage, NIR-AF imaging revealed an area of reduced fluorescence, whereas in the vitelliruptive stage, puncta of elevated NIR-AF signal were present. In both SW-AF and NIR-AF images, the vitelliform lesion in the atrophic stage was marked by reduced signal. At all stages of BVMD, nonlesion qAF was within the 95% confidence intervals for healthy eyes. Similarly, the NIR-AF intensity measurements outside the vitelliform lesion were comparable to the healthy control eye. SD-OCT scans revealed a fluid-filled detachment between the ellipsoid zone and the hyperreflectivity band attributable to RPE/Bruch's membrane.ConclusionsNIR-AF imaging can identify the pre-vitelliform stage of BVMD. Mutations in BEST1 are not associated with increased levels of SW-AF outside the vitelliform lesion. Elevated SW-AF within the fluid-filled lesion likely reflects the inability of RPE to phagocytose outer segments due to separation of RPE from photoreceptor cells, together with progressive photoreceptor cell impairment.

Longitudinal Clinical Follow-up and Genetic Spectrum of Patients With Rod-Cone Dystrophy Associated With Mutations in PDE6A and PDE6B

A precise phenotypic characterization of retinal dystrophies is needed for disease modeling as a basis for future therapeutic interventions. To compare genotype, phenotype, and structural changes in patients with rod-cone dystrophy (RCD) associated with mutations in PDE6A or PDE6B. In a retrospective cohort study conducted in Paris, France, from January 2007 to September 2017, 54 patients from a cohort of 1095 index patients with RCD underwent clinical examination, including personal and familial history, best-corrected visual acuity (BCVA), color vision, slitlamp examination, full-field electroretinography, kinetic visual fields (VFs), retinophotography, optical coherence tomography, near-infrared fundus autofluorescence, and short-wavelength fundus autofluorescence imaging. Genotyping was performed using microarray analysis, targeted next-generation sequencing, and Sanger sequencing validation with familial segregation when possible. Data were analyzed from September 1, 2017, to February 1, 2018. Clinical variables were subsequently analyzed in 2018. Phenotype and genotype comparison of patients carrying mutations in PDE6A or PDE6B. Of the 54 patients included in the study, 19 patients of 17 families (11 women [58%]; mean [SD] age at diagnosis, 14.83 [10.63] years) carried pathogenic mutations in PDE6A, and 35 patients of 26 families (17 women [49%]; mean [SD] age at diagnosis, 21.10 [11.56] years) had mutations in PDE6B, accounting for prevalences of 1.6% and 2.4%, respectively. Among 49 identified genetic variants, 14 in PDE6A and 15 in PDE6B were novel. Overall, phenotypic analysis revealed no substantial differences between the 2 groups except for night blindness as a presenting symptom that was noted to be more prevalent in the PDE6A than PDE6B group (80% vs 37%, respectively; P = .005). The mean binocular BCVA and VF decrease over time (measured as mean individual slopes coefficients) was comparable between patients with PDE6A and PDE6B mutations: 0.04 (0.12) vs 0.02 (0.05) for BCVA (P = .89) and 14.33 (7.12) vs 13.27 (6.77) for VF (P = .48). Mutations in PDE6A and PDE6B accounted for 1.6% and 2.4%, respectively, in a cohort of French patients with RCD. The functional and structural findings reported may constitute the basis of disease modeling that might be used for better prognostic estimation and candidate selection for photoreceptor therapeutic rescue.

Hyperautofluorescent Dots are Characteristic in Ceramide Kinase Like-associated Retinal Degeneration

There is a lack of studies which seek to discern disease expression in patients with mutations that alter retinal ceramide metabolism, specifically in the ceramide kinase like (CERKL) gene. This cross-sectional case series reports a novel phenotypic manifestation of CERKL-associated retinopathy. Four unrelated patients with homozygous CERKL mutations underwent a complete ocular exam, spectral-domain optical coherence tomography, short-wavelength fundus autofluorescence (SW-AF), quantitative autofluorescence (qAF), and full-field electroretinogram (ffERG). Decreased visual acuity and early-onset maculopathy were present in all patients. All four patients had extensive hyperautofluorescent foci surrounding an area of central atrophy on SW-AF imaging, which has not been previously characterized. An abnormal spatial distribution of qAF signal was seen in one patient, and abnormally elevated qAF8 signal in another patient. FfERG recordings showed markedly attenuated rod and cone response in all patients. We conclude that these patients exhibit several features that, collectively, may warrant screening of CERKL as a first candidate: early-onset maculopathy, severe generalized retinal dysfunction, peripheral lacunae, intraretinal pigment migration, and hyperautofluorescent foci on SW-AF.

Mutations in GPR143/OA1 and ABCA4 Inform Interpretations of Short-Wavelength and Near-Infrared Fundus Autofluorescence.

PurposeWe sought to advance interpretations and quantification of short-wavelength fundus autofluorescence (SW-AF) emitted from bisretinoid lipofuscin and near-infrared autofluoresence (NIR-AF) originating from melanin.MethodsCarriers of mutations in X-linked GPR143/OA1, a common form of ocular albinism; patients with confirmed mutations in ABCA4 conferring increased SW-AF; and subjects with healthy eyes were studied. SW-AF (488 nm excitation, 500–680 nm emission) and NIR-AF (excitation 787 nm, emission >830 nm) images were acquired with a confocal scanning laser ophthalmoscope. SW-AF images were analyzed for quantitative autofluoresence (qAF). Analogous methods of image acquisition and analysis were performed in albino and pigmented Abca4−/− mice and wild-type mice.ResultsQuantitation of SW-AF (qAF), construction of qAF color-coded maps, and examination of NIR-AF images from GPR143/OA1 carriers revealed mosaics in which patches of fundus exhibiting NIR-AF signal had qAF levels within normal limits whereas the hypopigmented areas in the NIR-AF image corresponded to foci of elevated qAF. qAF also was increased in albino versus pigmented mice. Although melanin contributes to fundus infrared reflectance, the latter appeared to be uniform in en face reflectance images of GPR143/OA1-carriers. In patients diagnosed with ABCA4-associated disease, NIR-AF increased in tandem with increased qAF originating in bisretinoid lipofuscin. Similarly in Abca4−/− mice having increased SW-AF, NIR-AF was more pronounced than in wild-type mice.ConclusionsThese studies corroborate RPE melanin as the major source of NIR-AF but also indicate that bisretinoid lipofuscin, when present at sufficient concentrations, contributes to the NIR-AF signal. Ocular melanin attenuates the SW-AF signal.

Quantitative Autofluorescence Intensities in Acute Zonal Occult Outer Retinopathy vs Healthy Eyes

Acute zonal occult outer retinopathy (AZOOR) remains a challenging diagnosis. Early recognition of the disease depends on advances in imaging modalities that can improve phenotyping and contribute to the understanding of the underlying pathogenesis. To expand the range of approaches available to assist in the identification of AZOOR by multimodal imaging and to analyze the fundus lesions by quantifying short-wavelength fundus autofluorescence (quantitative fundus autofluorescence [qAF]) and spectral-domain optical coherence tomography. In this observational study, patients underwent imaging at Columbia University Medical Center between November 2010 and March 2016 and were analyzed between September 2015 and August 2016. Six patients diagnosed as having AZOOR were studied by qAF and spectral-domain optical coherence tomography and were compared with 30 age and race/ethnicity–matched controls from a database of 277 healthy control eyes. In unaffected regions of the macula, qAF was calculated within predetermined circularly arranged segments (qAF8). In addition, qAF was measured within specified regions of interest positioned at the autofluorescent lesion border (AZOOR line). Electroretinograms and electro-oculograms were recorded in 5 of 6 patients. Among 6 patients (age range, 26-61 years; 4 female; 4 of white race/ethnicity, 1 Asian, and 1 Hispanic), 5 exhibited an autofluorescent AZOOR line in short-wavelength fundus autofluorescence images, delineating the peripapillary lesion. The mean (SD) region-of-interest qAF measured on the AZOOR line was 60 (26) times higher than in healthy control eyes (P = .03) at equivalent fundus locations. The qAF8 within nondiseased macular regions were within the normal range. At the lesion border, spectral-domain optical coherence tomography revealed a loss of outer retinal integrity in all patients. Single-flash cone b-wave latency and 30-Hz flicker latency responses were significantly delayed bilaterally. Lesions with smooth, homogeneous borders exhibited only minimal expansion in size over time, while the lesion in a patient with a heterogeneous border progressed more rapidly. The finding that qAF is elevated at the border between diseased and nondiseased retina in patients with AZOOR contributes to the understanding of the natural history of the disease.

Intraretinal Correlates of Reticular Pseudodrusen Revealed by Autofluorescence and En Face OCT.

PurposeWe sought to determine whether information revealed from the reflectance, autofluorescence, and absorption properties of RPE cells situated posterior to reticular pseudodrusen (RPD) could provide insight into the origins and structure of RPD.MethodsRPD were studied qualitatively by near-infrared fundus autofluorescence (NIR-AF), short-wavelength fundus autofluorescence (SW-AF), and infrared reflectance (IR-R) images, and the presentation was compared to horizontal and en face spectral domain optical coherence tomographic (SD-OCT) images. Images were acquired from 23 patients (39 eyes) diagnosed with RPD (mean age 80.7 ± 7.1 [SD]; 16 female; 4 Hispanics, 19 non-Hispanic whites).ResultsIn SW-AF, NIR-AF, and IR-R images, fundus RPD were recognized as interlacing networks of small scale variations in IR-R and fluorescence (SW-AF, NIR-AF) intensities. Darkened foci of RPD colocalized in SW-AF and NIR-AF images, and in SD-OCT images corresponded to disturbances of the interdigitation (IZ) and ellipsoid (EZ) zones and to more pronounced hyperreflective lesions traversing photoreceptor-attributable bands in SD-OCT images. Qualitative assessment of the outer nuclear layer (ONL) revealed thinning as RPD extended radially from the outer to inner retina. In en face OCT, hyperreflective areas in the EZ band correlated topographically with hyporeflective foci at the level of the RPE.ConclusionsThe hyperreflective lesions corresponding to RPD in SD-OCT scans are likely indicative of degenerating photoreceptor cells. The darkened foci at positions of RPD in NIR-AF and en face OCT images indicate changes in the RPE monolayer with the reduced NIR-AF and en face OCT signal suggesting a reduction in melanin that could be accounted for by RPE thinning.

Vascular abnormalities in patients with Stargardt disease assessed with optical coherence tomography angiography

AimsTo describe the vascular abnormalities in patients affected by Stargardt disease (STGD1) by means of optical coherence tomography angiography (OCT-A).MethodsCross-sectional case series, with the following inclusion criteria: diagnosis of STGD1,...

Characteristics of Hyperautofluorescent Choroidal Vessels within the Macular Atrophic Area Using Spectral-Domain Optical Coherence Tomography

Purpose: The aim of this study was to clarify the characteristic findings in patients with geographic atrophy with or without hyperautofluorescent choroidal vessels within macular atrophic areas on short-wavelength fundus autofluorescence imaging. Procedures: Sixty-seven eyes of 43 consecutive patients with macular atrophic areas were divided into groups with (group 1) and without (group 2) hyperautofluorescent choroidal vessels on fundus autofluorescence imaging and then retrospectively studied using spectral-domain optical coherence tomography. Results: In group 1 (n = 21), the average subfoveal choroidal thickness was 61.5 ± 20.1 μm, and the average foveal retinal thickness was 93.0 ± 51.3 μm. On the other hand, in group 2 (n = 46), the average subfoveal choroidal thickness was 200.7 ± 83.1 μm, and the average foveal retinal thickness was 109.2 ± 58.5 μm. Although retinal thickness did not differ significantly between the two groups (p = 0.28), the difference in choroidal thickness was statistically significant (p < 0.001). Conclusions: Choroidal thinning might contribute to the hyperautofluorescence of choroidal vessels.

THE VALUE OF RETINAL IMAGING WITH INFRARED SCANNING LASER OPHTHALMOSCOPY IN PATIENTS WITH STARGARDT DISEASE

To demonstrate the value of infrared scanning laser ophthalmoscopy (SLO) for determining structural retinal and choroidal changes in patients with Stargardt disease and its comparison to findings on short-wavelength fundus autofluorescence (SW-AF) imaging, spectral-domain optical coherence tomography, and microperimetry measurements. Forty-four eyes of 22 patients with Stargardt disease were studied using infrared-SLO, spectral-domain optical coherence tomography, macular microperimetry, SW-AF, electroretinography, and fundus photography. Although SW-AF imaging outlined the regions of retinal pigment epithelial (RPE) atrophy (hypofluorescence) and enhanced the visibility of more funduscopically apparent flecks (hyperfluorescence), infrared-SLO imaging outlined the regions of choroidal, and RPE, atrophic changes. Degenerative changes in photoreceptor and RPE cell layers, evident on spectral-domain optical coherence tomography imaging, were associated with either hyporeflective or hyperreflective images on infrared-SLO imaging, depending on whether both RPE and choroidal atrophy (hyperreflectance) or solely RPE atrophy (hyporeflectance) was present. Threshold elevations on microperimetry testing corresponded to both RPE and choroidal atrophy on infrared-SLO imaging and RPE atrophy on SW-AF. Although SW-AF identifies regions of RPE atrophy, infrared-SLO also identifies the involvement of the choroid that has important implications for the potential improvement in visual function from treatment. Thus, infrared-SLO imaging offers an additional advantage beyond that obtained with SW-AF.

Analysis of visual pigment by fundus autofluorescence

This study investigated changes of short-wavelength fundus autofluorescence (SW-AF) by retinal bleaching effects. All measurements were performed with the Heidelberg Retina Angiograph 2 (HRA 2). Initially, experimental imaging was done on a healthy eye after dark adaptation. Photopigment was bleached within the central 30° of the fundus by HRA 2 excitation light. Then SW-AF imaging of this region was performed, and SW-AF of the surrounding, unbleached 25° fundus region was subsequently studied by a wide-field lens. Next, another 30° SW-AF image of the posterior pole was obtained after complete dark adaptation. Then an extra SW-AF examination was performed with 15° temporal eccentricity, overlapping the original examination area. Finally, a successive image series was carried out on the dark-adapted eye to test for bleaching kinetics. The second and third experiments were also performed on eyes with macular dystrophies. Distinct regions of increased SW-AF were observed after strong illumination with the blue excitation light in all eyes studied. During light adaptation mean gray levels showed a saturation plateau after an initial steep increase. The resulting gray-value maps showed significant differences of pixel intensities between bleached and unbleached parts of the fundus. Two-dimensional density difference maps allowed analysis of visual pigment distribution and density in both healthy eyes and eyes with macular dystrophies. Our observations highlight the viability of objective, non-invasive evaluation of visual pigment in the healthy and diseased human retina by means of confocal fundus autofluorescence.