SLO Imaging Research Articles

Comparison of Widefield Laser Ophthalmoscopy and ETDRS Retinal Area for Diabetic Retinopathy

To evaluate agreement of nonmydriatic confocal scanning laser ophthalmoscopy (SLO; EIDON [CenterVue]) and the 7-standard field ETDRS area on ultrawide-field (UWF) SLO imaging for identification of diabetic retinopathy (DR) severity. Single-site, prospective, comparative, instrument validation study. One hundred ten eyes of 55 patients with diabetes mellitus were evaluated. Each patient underwent nonmydriatic, nonsimultaneous stereoscopic imaging using the EIDON camera and 4 fields of 60°× 55° were acquired (macula centered, disc centered, temporal macula, superotemporal). Mydriatic UWF retinal images were acquired using a nonsimultaneous stereographic protocol with UWF imaging (California; Optos plc). Before grading, a standardized ETDRS 7-field image mask was applied to all UWF retinal images. Images from each device were graded independently by 2 masked graders using the ETDRS clinical DR classification. Any discrepancy in DR grading between the devices was adjudicated by a third grader. κ Levels of agreement, sensitivity, and specificity for DR thresholds. Severity by ETDRS grading was as follows: no DR, 10.9%; mild nonproliferative DR (NPDR), 45.5%; moderate NPDR, 16.5%; severe NPDR, 11.8%; proliferative DR, 12.7%; high-risk proliferative DR, 2.7%; and ungradable, 0%. After adjudication, the level of DR identified on EIDON images agreed exactly with that of UWF ETDRS imaging in 87% of eyes (n= 96) and was within 1 step in 99.1% of eyes (n= 109) with a simple κ value of 0.8244 ± 0.0439 (95% confidence interval [CI], 0.7385-0.9104) and weighted (linear) κ value of 0.9041 ± 0.0257 (95% CI, 0.8537-0.9545). Sensitivity and specificity compared with ETDRS field grading for any DR were 0.96 and 0.75, for moderate NPDR or worse were 0.96 and 0.97, and for severe NPDR or worse were 0.91 and 1.00, respectively. Nonmydriatic 4-field stereoscopic widefield imaging using the EIDON device was comparable with the DR severity identified within the ETDRS 7-standard field area of UWF images. Future studies will need to evaluate the applicability of this device as a clinical and research tool and the impact of different widefield coverage areas.

Histopathological assessments reveal retinal vascular changes, inflammation, and gliosis in patients with lethal COVID-19

PurposeThe purpose of this study is to assess for histopathological changes within the retina and the choroid and determine the long-term sequelae of the SARS-CoV-2 infection.MethodsEyes from seven COVID-19-positive and six similar age-matched control donors with a negative test for SARS-CoV-2 were assessed. Globes were evaluated ex vivo with macroscopic, SLO and OCT imaging. Macula and peripheral regions were processed for Epon embedding and immunocytochemistry.ResultsFundus analysis shows hemorrhagic spots and increased vitreous debris in several of the COVID-19 eyes compared to the controls. OCT-based measurements indicated an increased trend in retinal thickness in the COVID-19 eyes; however, the difference was not statistically significant. Histology of the retina showed presence of hemorrhages and central cystoid degeneration in several of the donors. Whole mount analysis of the retina labeled with markers showed changes in retinal microvasculature, increased inflammation, and gliosis in the COVID-19 eyes compared to the controls. The choroidal vasculature displayed localized changes in density and signs of increased inflammation in the COVID-19 samples.ConclusionsIn situ analysis of the retinal tissue suggests that there are severe subclinical abnormalities that could be detected in the COVID-19 eyes. This study provides a rationale for evaluating the ocular physiology of patients that have recovered from COVID-19 infections to further understand the long-term effects caused by this virus.

Subretinal Drusenoid Deposits Revealed by Color SLO and Retro-Mode Imaging

Subretinal Drusenoid Deposits Revealed by Color SLO and Retro-Mode Imaging

Determining the Location of the Fovea Centralis Via En-Face SLO and Cross-Sectional OCT Imaging in Patients Without Retinal Pathology.

PurposeThe purpose was to establish the position of the fovea centralis to the optic nerve via en-face, near-infrared spectral domain optical coherence tomography (NIR-OCT) in healthy patients. This may shed light on physiological variability and be used for studying subtle cases of foveal ectopia in macular pathology and after retinal detachment.MethodsSD-OCT data of 890 healthy eyes were retrospectively analyzed. Exclusion criteria included axial myopia causing tilting of the optic disc, peripapillary atrophy >1/3 the width of the disc, macular images excluding greater than half of the optic disc, and patients unable to maintain vertical head positioning. Two independent reviewers measured the horizontal and vertical distance from the fovea to the optic disc center and optic disc diameter via cross-sectional and en-face scanning laser ophthalmoloscopy OCT imaging.Results890 eyes were included in the study. The right and left eyes differed in the horizontal distance from the fovea to the disc center (4359 vs. 4248 µm, P < 0.001) and vertical distance from the fovea to the disc center (464 µm vs. 647, P < 0.001). This corresponded to a smaller angle between the right and left eyes (6.07° vs. 8.67°, P < 0.001). Older age was associated with a larger horizontal (P = 0.008) and vertical distance (0.025). These differences persisted after correcting for axial length in the 487 patients with axial-length data.ConclusionsThis study compares the position of the fovea centralis among individuals without macular pathology on a micron level basis. The significant variability between right and left eyes indicates that contralateral eye evaluation cannot be reliably used. Rather, true foveal ectopia requires assessments of preoperative and postoperative NIR-OCT scans. This finding is relevant to retinal detachment cases and evaluation of subtle foveal ectopia.Translational RelevanceThis finding is relevant to retinal detachment cases and evaluation of subtle foveal ectopia.

Genetic Rescue Reverses Microglial Activation in Preclinical Models of Retinitis Pigmentosa.

Microglia cells (MGCs) play a key role in scavenging pathogens and phagocytosing cellular debris in the central nervous system and retina. Their activation, however, contributes to the progression of multiple degenerative diseases. Given the potential damage created by MGCs, it is important to better understand their mechanism of activation. Here, we explored the role of MGCs in the context of retinitis pigmentosa (RP) by using four independent preclinical mouse models. For therapeutic modeling, tamoxifen-inducible CreER was introduced to explore changes in MGCs when RP progression halted. The phenotypes of the MGCs were observed using live optical coherence tomography, live autofluorescence, and immunohistochemistry. We found that, regardless of genetic background, MGCs were activated in neurodegenerative conditions and migrated beyond the layers where they are typically found to the inner and outer segments, where degeneration was ongoing. Genetic rescue not only halted degeneration but also deactivated MGCs, regardless of whether the intervention occurred at the early, middle, or late stage of the disease. These findings suggest that halting long-term disease progression may be more successful by downregulating MGC activity while co-administering the therapeutic intervention.

Ultra-wide-field scanning laser ophthalmoscopy assists in the clinical detection and evaluation of asymptomatic early-stage familial exudative vitreoretinopathy

This study aims to investigate the ability of the ultra-wide-field scanning laser ophthalmoscope (UWF SLO) in clinically detecting and evaluating asymptomatic early-stage familial exudative vitreoretinopathy (FEVR). We retrospectively reviewed 163 eyes of 83 asymptomatic family members of 48 patients with FEVR. UWF SLO imaging (Optos® PLC, Scotland, UK) was performed on asymptomatic family members as a preliminary screening test for fundus anomalies, and the findings were compared with subsequent examinations using indirect fundus ophthalmoscopy in full mydriasis, fluorescein angiography (FA), fundus autoflourescence, and genetic sequencing. A total of 86 eyes of 43 asymptomatic family members were clinically diagnosed with early-stage FEVR, and 17 of the affected 43 family members were also genetically diagnosed. Compared with FA as a standard, the UWF SLO was highly effective in diagnosing FEVR with a sensitivity and specificity of 93.0% and 97.5%, respectively. The UWF SLO was able to diagnose early-stage FEVR in 93.0% of eyes, and guided the selection of therapies in 46.5% of the eyes studied. UWF SLO is a valuable imaging tool for detecting fundus anomalies related to early-stage FEVR, and this tool can assist in the clinical diagnosis and evaluation of early-stage FEVR in asymptomatic family members of patients with FEVR.

Adaptive-optics SLO imaging combined with widefield OCT and SLO enables precise 3D localization of fluorescent cells in the mouse retina.

Adaptive optics scanning laser ophthalmoscopy (AO-SLO) has recently been used to achieve exquisite subcellular resolution imaging of the mouse retina. Wavefront sensing-based AO typically restricts the field of view to a few degrees of visual angle. As a consequence the relationship between AO-SLO data and larger scale retinal structures and cellular patterns can be difficult to assess. The retinal vasculature affords a large-scale 3D map on which cells and structures can be located during in vivo imaging. Phase-variance OCT (pv-OCT) can efficiently image the vasculature with near-infrared light in a label-free manner, allowing 3D vascular reconstruction with high precision. We combined widefield pv-OCT and SLO imaging with AO-SLO reflection and fluorescence imaging to localize two types of fluorescent cells within the retinal layers: GFP-expressing microglia, the resident macrophages of the retina, and GFP-expressing cone photoreceptor cells. We describe in detail a reflective afocal AO-SLO retinal imaging system designed for high resolution retinal imaging in mice. The optical performance of this instrument is compared to other state-of-the-art AO-based mouse retinal imaging systems. The spatial and temporal resolution of the new AO instrumentation was characterized with angiography of retinal capillaries, including blood-flow velocity analysis. Depth-resolved AO-SLO fluorescent images of microglia and cone photoreceptors are visualized in parallel with 469 nm and 663 nm reflectance images of the microvasculature and other structures. Additional applications of the new instrumentation are discussed.

Fundus autofluorescence, OCT thickness evaluation, angiography and immunohistochemistry correlation in albino P23H rats

AbstractPurpose To evaluate the correlation between OCT changes and fundus autofluorescence (FAF) with immunohistochemistry (ICC) findings in an animal model of RP, the P23H rat and to investigate retinal and choroidal vascularization using fluorescein and indocianin green angiographyMethods Twenty albino homozigous P23H line 1 rats aging from P18 to 27 months and wild‐type albino Sprague‐Dawley (SD rats) (2 and 11 months old) were used for this study. Normal pigmented Long Evans (LE) 2 months old were used to compare FAF findings. SLO imaging and OCT were acquired using an Spectralis system (Heidelberg Engineering, Germany). For checking FAF, fluorescence was excited using diode laser at 488 nm. Thickness measurements were evaluated avoiding the ones close to the optic nerve. ICC was performed to correlate with the findings of OCT and AF changesResults During the course of P23H degeneration, the FAF pattern varied from not findings in young animals, some spotting at 2 months old to a mosaic of hyperfluorescent dots in the rats of 6 months or older. Retinal thicknesses diminished during the time. P23H rats showed great changes in morphology in advanced ages. Mean retinal thickness values varied from 189.88 μm at P60 to 58.15 μm at 27 months old. Retinal vascular plexus were diminished with time, and vessels exhibiting an abnormal, tortuous morphology could be observed.Conclusion FAF is a non‐invasive procedure that can detect changes in metabolic activity at the RPE in animal models of retinal degeneration in vivo. OCT and ICC show a good correlation. Retinal vascular plexus changes with aging.

High-Resolution Imaging of Retinal Nerve Fiber Bundles in Glaucoma Using Adaptive Optics Scanning Laser Ophthalmoscopy

To detect pathologic changes in retinal nerve fiber bundles in glaucomatous eyes seen on images obtained by adaptive optics (AO) scanning laser ophthalmoscopy (AO SLO). Prospective cross-sectional study. Twenty-eight eyes of 28 patients with open-angle glaucoma and 21 normal eyes of 21 volunteer subjects underwent a full ophthalmologic examination, visual field testing using a Humphrey Field Analyzer, fundus photography, red-free SLO imaging, spectral-domain optical coherence tomography, and imaging with an original prototype AO SLO system. The AO SLO images showed many hyperreflective bundles suggesting nerve fiber bundles. In glaucomatous eyes, the nerve fiber bundles were narrower than in normal eyes, and the nerve fiber layer thickness was correlated with the nerve fiber bundle widths on AO SLO (P < .001). In the nerve fiber layer defect area on fundus photography, the nerve fiber bundles on AO SLO were narrower compared with those in normal eyes (P < .001). At 60 degrees on the inferior temporal side of the optic disc, the nerve fiber bundle width was significantly lower, even in areas without nerve fiber layer defect, in eyes with glaucomatous eyes compared with normal eyes (P = .026). The mean deviations of each cluster in visual field testing were correlated with the corresponding nerve fiber bundle widths (P = .017). AO SLO images showed reduced nerve fiber bundle widths both in clinically normal and abnormal areas of glaucomatous eyes, and these abnormalities were associated with visual field defects, suggesting that AO SLO may be useful for detecting early nerve fiber bundle abnormalities associated with loss of visual function.

A clinical planning module for adaptive optics SLO imaging.

To develop a clinical planning module (CPM) to improve the efficiency of imaging subjects with a steerable wide-field adaptive optics scanning laser ophthalmoscope (AOSLO) and to evaluate the performance of this module by imaging the retina in healthy and diseased eyes. We developed a software-based CPM with two submodules: a navigation module and a montage acquisition module. The navigation module guides the AOSLO to image identified retinal regions from a clinical imaging platform using a matrix-based mapping between the two. The montage acquisition module systematically moves the AOSLO steering mirrors across the retina in predefined patterns. The CPM was calibrated using a model eye and tested on five normal subjects and one patient with a retinal nerve fiber layer defect. Within the central ±7° from the fixation target, the CPM can direct the AOSLO beam to the desired regions with localization errors of <0.3°. The navigation error increases with eccentricity, and larger errors (up to 0.8°) were evident for regions beyond 7°. The repeatability of CPM navigation was tested on the same locations from two subjects. The localization errors between trials on different days did not differ significantly (p > 0.05). The region with a size of approximately 13° × 10° can be imaged in about 30 min. An approximately 12° × 4.5° montage of the diseased region from a patient was imaged in 18 min. We have implemented a clinical planning module to accurately guide the AOSLO imaging beam to desired locations and to quickly acquire high-resolution AOSLO montages. The approach is not only friendly for patients and clinicians but also convenient to relate the imaging data between different imaging platforms.

Photoreceptor Damage and Foveal Sensitivity in Surgically Closed Macular Holes: An Adaptive Optics Scanning Laser Ophthalmoscopy Study

To assess photoreceptor structure using adaptive optics scanning laser ophthalmoscopy (AO SLO) and spectral-domain optical coherence tomography (SD OCT) and to evaluate the relationship between structural abnormalities and foveal sensitivity in eyes with surgically closed macular hole (MH). Prospective, interventional case series. Twenty-one eyes of 19 patients with idiopathic MH underwent a full ophthalmologic examination, including SD OCT at baseline. Imaging with SD OCT, an original prototype AO SLO system, and microperimetry were performed at 6 months after surgery. All patients underwent anatomically successful MH closure. On AO SLO, dark areas (0.004 to 0.754 mm(2)) were seen in all eyes after MH repair. Lower cone density correlated with poorer postoperative visual acuity and lower mean foveal sensitivity (both P < .001). Larger dark areas on AO SLO correlated with poorer postoperative visual acuity (P = .003) and lower mean foveal sensitivity (P = .006). Cone density was significantly lower and dark areas were significantly larger in eyes that had defects of the outer segments in the fluid cuff before surgery (P = .018 and P = .001, respectively) and moderately reflective foveal lesions after surgery (P < .001 and P < .001, respectively). Larger dark areas correlated with longer symptom duration before surgery (P < .001). Structural damage to the photoreceptor layer correlated with greater decreases in visual function in eyes with surgically closed MH. AO SLO imaging is a useful and quantitative tool for detecting photoreceptor abnormalities and their association with visual acuity and retinal sensitivity in eyes with closed MH.

OCT-Guided Photodynamic Therapy for Angiographic Occult CNV in Pathologic Myopia

Pathologic myopia (PM) is frequently complicated by choroidal neovascularization (CNV). Diagnosis is mainly clinical and angiographic but in recent years optical coherence tomography (OCT) has been noted to add important information. The authors report on the successfully OCT-guided photodynamic therapy (PDT) of an angiographic occult CNV complicating PM. Observational case report. Fluorescein angiography with a confocal SLO (HRA, Heidelberg Engineering, Germany) and OCT Stratus (Carl Zeiss Meditec, Inc.) imaging were used for diagnosis and monitoring of the CNV. Standard PDT was performed. A highly myopic 17-year-old girl complained of a drop in visual acuity (VA) in left eye (LE), dating back a few weeks. Her best-corrected (BC) VA was 20/40 in the LE, with some metamorphopsia. No hemorrhage or evident signs of CNV were visible either at fundus or at dynamic fluorescein angiography. OCT scans indicated a slight elevation of the RPE-choriocapillary complex with rarefaction of neuroretinal tissue that has been interpreted as CNV. PDT was then performed. LE BCVA had improved to 20/25, metamorphopsias disappeared, and at OCT examination no retinal morphologic modification was evident. Nine months later, BCVA and ophthalmoscopy are still stable. In this case, OCT was the fundamental tool for the correct diagnosis and posttherapymonitoring of CNV-complicated PM. The CNV, not clearly detectable using angiographic imaging, was treated with PDT, and results in terms of VA and anatomic resolution were good.

High-resolution imaging of retinal cells in the living eye

During diseases such as age-related macular degeneration, glaucoma, or retinal degeneration, visualizing and counting retinal cells would be of interest to diagnose early stages and to determine the short-term progression rate of these conditions. This is of major interest to optimize neuroprotective/regenerative therapies in these slowly progressive diseases. Such cellular imaging is already feasible in the eye of laboratory animals. In the near future, it is likely that visualization of individual neuronal cells in humans will become a routine clinical procedure, thanks to continuous technological improvements in optical imaging technologies. In mice, commercially available confocal scanning laser ophthalmoscope (cSLO) allows imaging of axons, capillaries, as well as dye-labelled cells (such as gfp-expressing cells). In humans, commercially available optical coherence tomography (OCT) allows routine imaging of retinal layering, but not of individual cells. Several research groups (including ours) are working in the highly competitive area of retinal cellular-imaging in humans. Different systems of adaptive optics, SLO, and OCT (in spectral domain a-mode or en face mode) and second harmonic generation imaging are under evaluation. These techniques have demonstrated cellular-level imaging capabilities of photoreceptors, of the nerve fibre layer and of capillaries. The main challenges that these systems face are biological (determination of the origin of the signal), medical (the clinical pertinence of the information) and technical (optimization of hardware, development of user interface).

Spontaneous resolution of a full thickness idiopathic macular hole: fundus autofluorescence and OCT imaging

To report on the spontaneous closure of a full thickness juxtafoveolar idiopathic macular hole (IMH) monitored with fundus autofluorescence (AF) as well as optical coherence tomography (OCT) imaging. Observational case report. Fundus Autofluorescence with a confocal SLO (HRA, Heidelberg Engineering, Germany) and OCT imaging were used to monitor the spontaneous evolution of a stage II IMH. A 70 year-old woman with unremarkable ocular history received a diagnosis of idiopathic macular hole in the left eye. Bright autofluorescence corresponding to the IMH was documented with the confocal SLO and OCT imaging could confirm the presence of an hour glass shaped full thickness juxtafoveolar IMH. Biomicroscopy revealed no posterior vitreous detachment (PVD). Few months later clinical examination demonstrated the presence of typical symptoms and signs of PVD (miodesopsias and Weiss ring). The bright autofluorescence corresponding to the IMH disappeared and OCT imaging documented a normal fovea in morphology and thickness. Spontaneous closure of full thickness juxtafoveolar IMH can occur and may be properly monitored with fundus AF as well as OCT imaging.

Oximetry with a multiple wavelength SLO.

Oximetry may be useful for understanding pathologies of the eye. We use a prototype scanning laser ophthalmoscope capable of simultaneous multiple wavelength imaging to record fundus images. The system is run under non-confocal conditions in using slit apertures with a width of up to 600 microm. The laser lines launched into the SLO were the 633 nm line of a HeNe-laser, and the 815 nm line from a tunable (700 to 900 nm) cw Ti:Sapphire-laser. These wavelengths were selected because of their availability and absorption characteristics. The difference in absorption at these two wavelengths is used to assess blood oxygen content. Images were averaged to improve the signal to noise ratio. This simultaneous method of measuring oxygen content may be preferred to other techniques such as sequential SLO imaging at different wavelengths, or spot analysis using a modified fundus camera. The advantages of this technique are that image registration is not required, and a large area of the retina can be assessed concurrently.