Search

The molecular signaling cascades that regulate angiogenesis and microvascular remodeling are fundamental to normal development, healthy physiology, and pathologies such as inflammation and cancer. Yet quantifying such complex, fractally branching vascular patterns remains difficult. We review application of NASA’s globally available, freely downloadable VESsel GENeration (VESGEN) Analysis software to numerous examples of 2D vascular trees, networks, and tree-network composites. Upon input of a binary vascular image, automated output includes informative vascular maps and quantification of parameters such as tortuosity, fractal dimension, vessel diameter, area, length, number, and branch point. Previous research has demonstrated that cytokines and therapeutics such as vascular endothelial growth factor, basic fibroblast growth factor (fibroblast growth factor-2), transforming growth factor-beta-1, and steroid triamcinolone acetonide specify unique “fingerprint” or “biomarker” vascular patterns that integrate dominant signaling with physiological response. In vivo experimental examples described here include vascular response to keratinocyte growth factor, a novel vessel tortuosity factor; angiogenic inhibition in humanized tumor xenografts by the anti-angiogenesis drug leronlimab; intestinal vascular inflammation with probiotic protection by Saccharomyces boulardii, and a workflow programming of vascular architecture for 3D bioprinting of regenerative tissues from 2D images. Microvascular remodeling in the human retina is described for astronaut risks in microgravity, vessel tortuosity in diabetic retinopathy, and venous occlusive disease.

Purpose Lafora disease is a genetic neurodegenerative metabolic disorder caused by insoluble polyglucosan aggregate accumulation throughout the central nervous system and body. The retina is an accessible neural tissue, which may offer alternative methods to assess neurological diseases quickly and noninvasively. In this way, noninvasive imaging may provide a means to characterize neurodegenerative disease, which enables earlier identification and diagnosis of disease and the ability to monitor disease progression. In this study, we sought to characterize the retina of individuals with Lafora disease using non-invasive retinal imaging. Methods One eye of three individuals with genetically confirmed Lafora disease were imaged with optical coherence tomography (OCT) and adaptive optics scanning light ophthalmoscopy (AOSLO). When possible, OCT volume and line scans were acquired to assess total retinal thickness, ganglion cell-inner plexiform layer thickness, and outer nuclear layer + Henle fiber layer thickness. OCT angiography (OCTA) scans were acquired in one subject at the macula and optic nerve head (ONH). AOSLO was used to characterize the photoreceptor mosaic and examine the retinal nerve fiber layer (RNFL). Results Two subjects with previous seizure activity demonstrated reduced retinal thickness, while one subject with no apparent symptoms had normal retinal thickness. All other clinical measures, as well as parafoveal cone density, were within normal range. Nummular reflectivity at the level of the RNFL was observed using AOSLO in the macula and near the ONH in all three subjects. Conclusions This multimodal retinal imaging approach allowed us to observe a number of retinal structural features in all three individuals. Most notably, AOSLO revealed nummular reflectivity within the inner retina of each subject. This phenotype has not been reported previously and may represent a characteristic change produced by the neurodegenerative process.

Purpose Fundus albipunctatus is an inherited cause of congenital stationary night blindness. The objective of this report is to describe structural changes occurring in a macular phenotype of a novel RDH5 mutation producing fundus albipunctatus using high-resolution in vivo imaging. A 62-year-old male with longstanding night blindness underwent imaging and genetic evaluation. High-resolution images of the photoreceptor mosaic were compared to those of a healthy subject. Results of a comprehensive ophthalmic evaluation and genetic testing with imaging including fundus photography, spectral-domain optical coherence tomography (OCT), fluorescein angiography (FA), OCT angiography (OCT-A), and adaptive optics scanning light ophthalmoscopy (AOSLO) are described. Observations The patient presented with visual acuity of 20/25 in both eyes and longstanding poor dark adaptation. Anterior segment examination was unremarkable. Fundoscopy revealed well circumscribed bilateral perifoveal mottling and atrophy in both eyes. Discrete white-yellow flecks were present beyond the vascular arcades extending to the far periphery. Genetic testing revealed a novel compound heterozygous RDH5 mutation (c.388C > T, p.Gln130*; c.665T > C, p.Leu222Pro). OCT demonstrated perifoveal photoreceptor and outer retinal irregularities, which corresponded to a window defect with late staining on FA. OCT-A demonstrated normal retinal vasculature with patchy areas of non-perfusion in the choriocapillaris. Macular abnormalities in both eyes were imaged using AOSLO to assess cone and rod photoreceptor architecture. While clinical features are consistent with a primary rod disorder, confocal AOSLO showed a paucity of normal cones with a small spared central island in both eyes. Rods appeared larger and more irregular throughout the macula. Non-confocal split detection AOSLO imaging revealed the presence of cone inner segments in dark regions of confocal imaging, indicating some degree of photoreceptor preservation. Conclusions and Importance The AOSLO imaging of this particular macular phenotype of fundus albipunctatus demonstrates some of the structural photoreceptor abnormalities that occur in this condition, adding insight to the variable presentation of RDH5 retinopathy. The presence of preserved inner segment architecture suggests the possibility that gene therapy could play a future role in treating this condition.

Introduction. We present a case of serpiginous choroidopathy (SC) with novel OCTA and en face OCT reflectance findings which help identify subclinical disease progression. Case Presentation. En face OCT reflectance images demonstrated outer retinal tubules (ORT) at the serpiginous lesion margins of affected and unaffected retina on multimodal imaging. OCTA findings demonstrate variable dropout of choriocapillaris in “normal” retina beyond lesion borders which was not visible on standard imaging and which demonstrated a clear transition zone beyond the ORT. Discussion. This is the first report of choriocapillaris atrophy identified on OCTA not identified on traditional multimodal imaging in serpiginous choroidopathy. Damage to vasculature only visible with OCTA may help characterize the distribution of inflammation, aiding in monitoring of suppression not illustrated by traditional imaging and which may threaten the central macula. ORT in SC suggest death and reorganization of outer segments from dysfunction of the choriocapillaris and RPE, as well as serve to demarcate the area of chronic or old inflammation, supporting the hypothesis that the choriocapillaris is the primary site of inflammation in SC. Based on these findings, we recommend OCTA on all patients with serpiginous choroidopathy to monitor underlying state of inflammation and help determine immunosuppressive threshold.

Purpose To image retinal macrophages at the vitreoretinal interface in the living human retina using a clinical optical coherence tomography (OCT) device. Methods Eighteen healthy controls and three patients with retinopathies were imaged using a clinical spectral-domain OCT. In controls, 10 sequential scans were collected at three different locations: (1) ∼9 degrees temporal to the fovea, (2) the macula, and (3) the optic nerve head (ONH). Intervisit repeatability was evaluated by imaging the temporal retina twice on the same day and 3 days later. Only 10 scans at the temporal retina were obtained from each patient. A 3-µm OCT reflectance (OCT-R) slab located above the inner limiting membrane (ILM) surface was averaged. Results In controls, ramified macrophage-like cells with regular spatial separation were visualized in the temporal and ONH OCT-R images; however, cell structures were not resolvable at the macula. Interim changes in cell position suggestive of cell translocation were observed between images collected on the same day and those collected 3 days later. There was considerable variation in cell density and nearest-neighbor distance (NND) across controls. Mean ± SD cell densities measured at the temporal and ONH were 78 ± 23 cells/mm2 and 57 ± 16 cells/mm2, respectively. Similarly, mean ± SD NNDs measured at the temporal and ONH were 74.3 ± 13.3 µm and 93.3 ± 20.0 µm, respectively. Nonuniform spatial distribution and altered morphology of the cells were identified in patients with retinopathies. Conclusions Our findings showed regular spatial separation and ramified morphology of macrophage-like cells on the ILM surface with cell translocation over time in controls. Their distribution and morphology suggest an origin of macrophage-like cells such as microglia or hyalocytes.

ObjectivesTo examine interocular asymmetry of foveal avascular zone (FAZ) and parafoveal capillary density metrics in sickle cell retinopathy (SCR) using optical coherence tomography angiography (OCT-A).MethodsThis cross-sectional, retrospective study evaluated SCR patients and unaffected controls who underwent 3x3mm macular OCT-A imaging using a spectral domain-OCT system. FAZ (area, perimeter, and acircularity index) and parafoveal capillary density metrics were computed for both eyes of each participant. In unaffected controls, interocular difference in FAZ and parafoveal capillary density metrics were evaluated using Bland-Altman plots. SCR patients with interocular difference outside the upper 97.5% and lower 2.5% limits of agreement from controls were defined as having interocular asymmetry. Area under receiver operating characteristic curve (AROC) was also performed to determine the ability of the absolute interocular difference to differentiate between subjects with SCR-including non-proliferative SCR (NP-SCR) and proliferative SCR (P-SCR)-and unaffected controls.ResultsThirty-one patients with SCR (21 NP-SCR and 10 P-SCR) and 14 race-matched and age-matched controls were included for analysis. Interocular asymmetry was seen for all FAZ and parafoveal capillary density metrics in NP-SCR and P-SCR subjects. SCR subjects showed greater disease severity in the left-eye for FAZ and parafoveal capillary density metrics.ConclusionsNP-SCR and P-SCR patients demonstrated quantifiable interocular asymmetry in FAZ and parafoveal capillary density metrics compared to unaffected subjects, with left-eye predominance in disease severity.

Background/Purpose:To report a case of persistent retinal toxicity associated with a high dose of sildenafil citrate intake.Methods:Single retrospective case report.Results:A 31-year-old white man with no medical history presented with complaints of bilateral multicolored photopsias and erythropsia

Aims To assess peripapillary perfused capillary density (PCD) in primary open-angle glaucoma (POAG) across stage of disease. Methods In this observational, cross-sectional study, 60 eyes with varying stages of POAG and 24 control eyes were imaged on a spectral domain optical coherence tomography angiography system (AngioVue, Optovue, Fremont, California, USA) generating images centred on the optic nerve head. Major blood vessels were removed using custom automated software. PCD was calculated as a percentage as the ratio of pixels associated with perfused capillaries to the total number of pixels in the corresponding region-of-interest (ROI). Analysis of covariance was used to compare PCD among the subject groups and control for possible covariates. Area under the receiver operating characteristic curve (AROC) and sensitivity at 95% specificity were calculated to assess the capability of PCD to distinguish mild glaucoma from control. The Pearson9s product-moment correlation coefficient was used to assess correlations between PCD and circumpapillary retinal nerve fibre layer thickness (cpRNFLT) and visual field mean deviation (MD). Results PCD demonstrated a progressive stepwise decrease from control eyes throughout worsening POAG stage at all ROIs. PCD demonstrated AROC and sensitivity values comparable to cpRNFLT and visual field parameters and exhibited significant correlations with cpRNFLT and MD at all corresponding ROIs. Conclusions PCD displayed significant correlations with morphological and functional indices and exhibited diagnostic capabilities comparable to currently employed clinical variables. Our preliminary results suggest that PCD analysis may prove to be a useful tool in monitoring POAG across stage and identifying early POAG.

PURPOSE: To assess the frequency and impact of abnormal foveal avascular zone (FAZ) topography (i.e., a fragmented FAZ) on visual acuity and foveal anatomic features. DESIGN: Prospective, cross-sectional study from March 2018 through July 2019. PARTICIPANTS: Two-hundred fifty participants were screened from a normative OCT angiography database. Of those, 12 participants were found to have at least 1 eye with a fragmented FAZ. Eight returned for follow-up imaging, along with an additional 3 participants with ocular disease (amblyopia, autosomal recessive bestrophinopathy, premature birth) having a similar FAZ phenotype. METHODS: Follow-up OCT imaging and monocular best-corrected visual acuity (BCVA) were performed for these 11 participants. Twenty-four participants with a clearly defined FAZ were recruited for comparison. A normative database was created measuring parafoveal intercapillary area (PICA) to determine of an FAZ was fragmented. MAIN OUTCOME MEASURES: Monocular BCVA, foveal pit depth, foveal pit area, PICA, outer nuclear layer thickness, foveal inner retinal area, and peak cone density. RESULTS: The frequency of a fragmented FAZ was 4.8% of individuals (12 of 250) or 3.6% of eyes (18 of 500 eyes). A significant difference was found between the control eyes and eyes with fragmented FAZs for foveal pit depth, pit area, and total PICA (P < 0.001, P = 0.002, and P < 0.001, respectively). The presence of a fragmented FAZ did not affect visual acuity. CONCLUSIONS: The presence of a fragmented FAZ seems not to be a rare phenotype in individuals with normal vision. The presence of altered FAZ topography in patients with retinal or systemic disease could impact negatively the accuracy and sensitivity of biomarkers dependent on FAZ identification.

Background/aimsTo assess foveal avascular zone (FAZ) morphology and parafoveal capillary perfusion in patients with various stages of sickle cell retinopathy (SCR) using optical coherence tomography angiography (OCT-A).MethodsThis is a multi-institutional retrospective study of patients with various stages of SCR compared with healthy controls. Parafoveal OCT-A images obtained using a commercial spectral domain-OCT system were reviewed. Foveal-centred 3×3 mm full vascular slab OCT-As were used for image processing and data analysis. FAZ area, perimeter, and acircularity index were determined on the OCT-A image after manual delineation of the FAZ border. Quadrant-based parafoveal capillary density and per cent area deviating from normal distribution were also measured.ResultsFifty-two patients with SCR (33 non-proliferative and 19 proliferative) and 20 age and race-matched healthy controls were included. One randomly selected eye per study participant was analysed. FAZ perimeter and acircularity index were significantly greater in SCR eyes when compared with the controls. While parafoveal capillary density was significantly lower, per cent area deviated from normal distribution was significantly higher in SCR eyes than that of the control. However, no statistically significant difference between the two SCR stages was observed. In quadrant-based analysis, the temporal quadrant showed greater parafoveal capillary dropout due to SCR, with the most profound effect in patients with proliferative SCR.ConclusionsAbnormal FAZ morphology and altered parafoveal capillary perfusion were found in patients with SCR. Our customised OCT-A image analysis method uniquely highlights significant quantitative alterations in perfusion density mapping in a qualitative display, with minimal obscuration of OCT-A image detail.

PURPOSE Intravitreal injections acutely and temporarily increase intraocular pressure (IOP), and this may have cumulative long-term effects including an increased risk for glaucoma surgery. This study was designed to measure retinal perfusion density changes on optical coherence tomography (OCT) angiography and OCT thickness alterations associated with acutely increased IOP after intravitreal injections. METHODS Retrospective observational clinical study of 40 eyes (39 patients) with various retinopathies from October 2016 to June 2017 at a tertiary care retina clinic in NYC. Patients were older than 18 years, with vision >20/100, able to fixate and without media opacities precluding OCT angiography, receiving intravitreal bevacizumab or aflibercept for diabetic retinopathy, retinal vein occlusion, macular degeneration, retinal neovascularization, or radiation retinopathy. The 3-mm × 3-mm macular and 4.5-mm × 4.5-mm peripapillary OCT angiography perfusion density, macular OCT thickness, and IOP were measured before and immediately after intravitreal injections. Paired t-test was used to compare preinjection and postinjection values for perfusion density and OCT thickness. Regression analysis was performed for potential effects of baseline IOP, IOP change, and age. RESULTS Statistically significant decreases in angiographic perfusion density (P < 0.05) were found in most areas of the superficial and deep layer macular OCT angiography, and the overall optic nerve head and the radial peripapillary capillary layer, preferentially temporal. Macular OCT thickness was significantly decreased in the temporal region and increased in the nasal region. Regression analysis showed relationships between age and decreased superficial macular perfusion. Preinjection IOP was only related to OCT thickness in the fovea. Intraocular pressure change was related only to decreased superficial macular perfusion density. CONCLUSION Intravitreal injections produce acute IOP changes that are associated with reduced macular and peripapillary perfusion density. Therefore, it is possible that patients receiving regular intravitreal injections may be sustaining perfusion-related injury to ocular structures that may produce glaucomatous damage to the macula and optic nerve.

Purpose To compare perfused capillary density (PCD) in diabetic patients and healthy controls using optical coherence tomography angiography (OCTA). Methods Forty controls, 36 diabetic subjects without clinical retinopathy (NoDR), 38 with nonproliferative retinopathy (NPDR), and 38 with proliferative retinopathy (PDR) were imaged using spectral-domain optical coherence tomography. A 3 × 3-mm full-thickness parafoveal OCTA scan was obtained from each participant. Following manual delineation of the foveal avascular zone (FAZ), FAZ area, perimeter, and acircularity index were determined. Seven consecutive equidistant 200-μm-wide annular segments were drawn at increasing eccentricities from the FAZ margin. Annular PCD (%) was defined as perfused capillary area divided by the corresponding annulus area after subtraction of noncapillary blood vessel areas. Nonparametric Kruskal-Wallis testing with Bonferroni correction was performed in pairwise comparisons of group PCD values. Results The NoDR group demonstrated consistently higher PCD compared to the control group in all 7 annuli, reaching statistical significance (36.6% ± 3.30% vs 33.6% ± 3.98%, P = .034) at the innermost annulus (FAZ margin to 200 μm out). The NPDR and PDR groups demonstrated progressively decreasing PCD. Differences in FAZ metrics between the NoDR and control groups did not reach statistical significance. Conclusions Relative to healthy controls, increased PCD values in the NoDR group likely represent an autoregulatory response to increased metabolic demand, while the decrease in PCD that follows in NPDR and PDR results largely from an incremental loss of capillary segments. These findings, consistent with previous studies, demonstrate the potential of OCTA as a clinical tool for earlier objective detection of preclinical diabetic retinopathy. NOTE: Publication of this article is sponsored by the American Ophthalmological Society.

Pathophysiology of sickle cell disease (SCD) features intermittent vaso-occlusion of microcirculatory networks that facilitate ischemic damage. Past research has, however, relied on static images to characterize this active disease state. This study develops imaging metrics to more fully capture dynamic vascular changes, quantifying intermittent retinal capillary perfusion in unaffected controls and SCD patients using sequential optical coherence tomography angiography (OCT-A) scans. The results reveal significant dynamic variation of capillary perfusion in SCD patients compared to controls. This measurement of vaso-occlusive burden in patients would provide utility in monitoring of the disease state and in evaluating treatment efficacy.

Purpose The purpose of this study was to investigate the effect of device and scan size on quantitative optical coherence tomography angiography (OCT-A) metrics. Methods The 3 × 3 mm scans from Optovue AngioVue and Zeiss AngioPlex systems were included for 18 eyes of 18 subjects without ocular pathology. The foveal avascular zone (FAZ) was segmented manually by two observers, from which estimates of FAZ area (using both the nominal image scale and the axial length corrected image scale) and acircularity were derived. Three scan sizes (3 mm, 6 mm HD, and 8 mm) from the AngioVue system were included for 15 eyes of 15 subjects without ocular pathology. For each subject, larger image sizes were resized to the same resolution as 3 × 3 mm scans, aligned, then cropped to a common area. FAZ area, FAZ acircularity, average and total parafoveal intercapillary area, vessel density, and vessel end points were computed. Results Between the devices used here, there were no significant differences in FAZ acircularity (P = 0.88) or FAZ area using scaled (P = 0.11) or unscaled images (P = 0.069). Although there was no significant difference in FAZ area across scan sizes (P = 0.30), vessel morphometry metrics were all significantly influenced by scan size. Conclusions The scan devices and sizes used here do not affect FAZ area measures derived from manual segmentations. In contrast, vessel morphometry metrics are affected by scan size. As individual differences in axial length induce differences in absolute scan size, extreme care should be taken when interpreting metrics of vessel morphometry, both between and within OCT-A devices. Translational relevance A better characterization of the confounds surrounding OCT-A retinal vasculature metrics can lead to improved application of these metrics as biomarkers for retinal and systemic diseases.

As techniques of retinal imaging have evolved, anatomic features that were only assessable in the laboratory have become available in the clinic for patient care. The retinal capillaries were initially described on microscope sections in the pathology laboratory. As optical methods have advanced these features have become part of the routine clinical landscape inspected daily by physicians. This paper briefly traces the evolution of these techniques and shows how they fit into the modern diagnostic armamentarium of ophthalmic retinal care.

Purpose To present a method for age-matched deviation mapping in the assessment of disease-related changes to the radial peripapillary capillaries (RPCs). Methods We reviewed 4.5x4.5mm en face peripapillary OCT-A scans of 133 healthy control eyes (133 subjects, mean 41.5 yrs, range 11–82 yrs) and 4 eyes with distinct retinal pathologies, obtained using spectral-domain optical coherence tomography angiography. Statistical analysis was performed to evaluate the impact of age on RPC perfusion densities. RPC density group mean and standard deviation maps were generated for each decade of life. Deviation maps were created for the diseased eyes based on these maps. Large peripapillary vessel (LPV; noncapillary vessel) perfusion density was also studied for impact of age. Results Average healthy RPC density was 42.5±1.47%. ANOVA and pairwise Tukey-Kramer tests showed that RPC density in the ≥60yr group was significantly lower compared to RPC density in all younger decades of life (p

Importance Solar retinopathy is a rare form of retinal injury that occurs after direct sungazing. Objective To enhance understanding of the structural changes that occur in solar retinopathy by obtaining high-resolution in vivo en face images. Design, Setting, and Participant Case report of a young adult woman who presented to the New York Eye and Ear Infirmary with symptoms of acute solar retinopathy after viewing the solar eclipse on August 21, 2017. Main Outcomes and Measures Results of comprehensive ophthalmic examination and images obtained by fundus photography, microperimetry, spectral-domain optical coherence tomography (OCT), adaptive optics scanning light ophthalmoscopy, OCT angiography, and en face OCT. Results The patient was examined after viewing the solar eclipse. Visual acuity was 20/20 OD and 20/25 OS. The patient was left-eye dominant. Spectral-domain OCT images were consistent with mild and severe acute solar retinopathy in the right and left eye, respectively. Microperimetry was normal in the right eye but showed paracentral decreased retinal sensitivity in the left eye with a central absolute scotoma. Adaptive optics images of the right eye showed a small region of nonwaveguiding photoreceptors, while images of the left eye showed a large area of abnormal and nonwaveguiding photoreceptors. Optical coherence tomography angiography images were normal in both eyes. En face OCT images of the right eye showed a small circular hyperreflective area, with central hyporeflectivity in the outer retina of the right eye. The left eye showed a hyperreflective lesion that intensified in area from inner to middle retina and became mostly hyporeflective in the outer retina. The shape of the lesion on adaptive optics and en face OCT images of the left eye corresponded to the shape of the scotoma drawn by the patient on Amsler grid. Conclusions and Relevance Acute solar retinopathy can present with foveal cone photoreceptor mosaic disturbances on adaptive optics scanning light ophthalmoscopy imaging. Corresponding reflectivity changes can be seen on en face OCT, especially in the middle and outer retina. Young adults may be especially vulnerable and need to be better informed of the risks of viewing the sun with inadequate protective eyewear.

To describe a new method of retinal vascular perfusion density mapping using optical coherence tomography angiography and to compare current staging of diabetic retinopathy based on clinical features with a new grading scale based on perifoveal perfusion densities.A retrospective review was performed on subjects with diabetic retinopathy and age-matched controls imaged with a spectral domain optical coherence tomography system (Optovue XR Avanti, Fremont, CA). Split-spectrum amplitude-decorrelation angiography (SSADA) generated optical coherence tomography angiograms of the superficial retinal capillaries, deep retinal capillaries, and choriocapillaris. Skeletonized optical coherence tomography angiograms were used to create color-coded perfusion maps and capillary perfusion density values for each image. Capillary perfusion density values were compared with clinical staging, and groups were compared using analysis of variance and Kruskal-Wallis analyses.Twenty-one control and 56 diabetic retinopathy eyes were imaged. Diabetic eyes were grouped according to clinical stage. Capillary perfusion density values from each microvascular layer were compared across all groups. Capillary perfusion density values were significantly lower in nearly all layers of all study groups compared with controls. Trend analysis showed a significant decrease in capillary perfusion density values as retinopathy progresses for most layers.Quantitative retinal vascular perfusion density mapping agreed closely with grading based on clinical features and may offer an objective method for monitoring disease progression in diabetic retinopathy.

PURPOSE To better understand the nature of glaucomatous damage of the macula, especially the structural changes seen between relatively healthy and clearly abnormal (AB) retinal regions, using an adaptive optics scanning light ophthalmoscope (AO-SLO). METHODS Adaptive optics SLO images and optical coherence tomography (OCT) vertical line scans were obtained on one eye of seven glaucoma patients, with relatively deep local arcuate defects on the 10-2 visual field test in one (six eyes) or both hemifields (one eye). Based on the OCT images, the retinal nerve fiber (RNF) layer was divided into two regions: (1) within normal limits (WNL), relative RNF layer thickness within mean control values ±2 SD; and (2) AB, relative thickness less than -2 SD value. RESULTS As seen on AO-SLO, the pattern of AB RNF bundles near the border of the WNL and AB regions differed across eyes. There were normal-appearing bundles in the WNL region of all eyes and AB-appearing bundles near the border with the AB region. This region with AB bundles ranged in extent from a few bundles to the entire AB region in the case of one eye. All other eyes had a large AB region without bundles. However, in two of these eyes, a few bundles were seen within this region of otherwise missing bundles. CONCLUSIONS The AO-SLO images revealed details of glaucomatous damage that are difficult, if not impossible, to see with current OCT technology. Adaptive optics SLO may prove useful in following progression in clinical trials, or in disease management, if AO-SLO becomes widely available and easy to use.

PURPOSE To compare the peripapillary perfused capillary density (PCD) among eyes with exfoliation syndrome (XFS), exfoliation glaucoma (XFG), primary open-angle glaucoma (POAG), and controls. DESIGN Observational, cross-sectional study. METHODS Optical coherence tomography angiography (OCTA) images (4.5×4.5 mm) of the optic nerve head were obtained using a commercial spectral domain OCTA system (AngioVue Avanti RTVue-XR, OptoVue, Fremont, CA). Two concentric circles with 1.95-mm (inner) and 3.45-mm (outer) diameters were placed manually, producing an annulus of width 0.75 mm centered at the optic disc. PCD was calculated as the ratio of pixels associated with capillaries to the pixels in the annulus after large blood vessel removal. Continuous variables were assessed by analysis of variance and Tukey honest significant difference (HSD) test. Multiple linear regression analysis was performed to adjust for confounding factors. RESULTS Forty-three XFG eyes, 31 POAG eyes matched for visual field mean deviation, 33 XFS eyes, and 45 control eyes were analyzed. PCD demonstrated a progressive decrease from controls to XFS to POAG to XFG. Pairwise comparisons revealed significant differences in PCD between each pair of groups (all P

PURPOSE Both non-arteritic anterior ischemic optic neuropathy (NAION) and primary open-angle glaucoma (POAG) damage retinal ganglion cell axons, which are perfused by the radial peripapillary capillaries. To evaluate the pattern of ischemia, we compared peripapillary capillary density (PCD) in NAION eyes to POAG eyes matched for visual field mean deviation and retinal nerve fiber layer thickness. METHODS 31 chronic NAION (>6 months after the acute event) and unaffected fellow eyes (31 subjects), 42 moderate and severe POAG eyes (27 subjects), and 77 control eyes (46 healthy subjects) were imaged with a commercial optical coherence tomography angiography system (AngioVue, Avanti RTVue-XR, Optovue, CA) at two academic institutions. Two concentric circles of diameters 1.95mm (inner) and 3.45mm (outer) were manually placed on images centered on the optic nerve head, producing an annular region-of-interest. Image analysis with major vessel removal was performed using a custom program. Whole-image, whole-annulus, and sectoral PCDs were measured. RESULTS Whole-image and whole-annulus PCDs in NAION and moderate and severe POAG eyes were significantly decreased compared to unaffected fellow eyes and control eyes (all P

PURPOSE Deep glaucomatous defects near fixation were followed over time with an adaptive optics-scanning light ophthalmoscope (AO-SLO) to better understand the progression of these defects and to explore the use of AO-SLO in detecting them. METHODS Six eyes of 5 patients were imaged with an AO-SLO from 2 to 4 times for a range of 14.6 to 33.6 months. All eyes had open-angle glaucoma with deep defects in the superior visual field (VF) near fixation as defined by 10-2 VFs with 5 or more points less than -15 dB; two of the eyes had deep defects in the inferior VF as well. AO-SLO images were obtained around the temporal edge of the disc. RESULTS In 4 of the 6 eyes, the edge of the inferior-temporal disc region of the retinal nerve fiber (RNF) defect seen on AO-SLO moved closer to fixation within 10.6 to 14.7 months. In 4 eyes, RNF bundles in the affected region appeared to lose contrast and/or disappear. CONCLUSIONS Progressive changes in RNF bundles associated with deep defects on 10-2 VFs can be seen within about 1 year with AO-SLO imaging. These changes are well below the spatial resolution of the 10-2 VF. On the other hand, subtle thinning of regions with RNF bundles is not easy to see with current AO-SLO technology, and may be better followed with OCT. TRANSLATIONAL RELEVANCE AO-SLO imaging may be useful in clinical trials designed to see very small changes in deep defects.

Objectives To assess the effect of image registration and averaging on the visualization and quantification of the radial peripapillary capillary (RPC) network on optical coherence tomography angiography (OCTA). Methods Twenty-two healthy controls were imaged with a commercial OCTA system (AngioVue, Optovue, Inc.). Ten 10x10° scans of the optic disc were obtained, and the most superficial layer (50-μm slab extending from the inner limiting membrane) was extracted for analysis. Rigid registration was achieved using ImageJ, and averaging of each 2 to 10 frames was performed in five ~2x2° regions of interest (ROI) located 1° from the optic disc margin. The ROI were automatically skeletonized. Signal-to-noise ratio (SNR), number of endpoints and mean capillary length from the skeleton, capillary density, and mean intercapillary distance (ICD) were measured for the reference and each averaged ROI. Repeated measures analysis of variance was used to assess statistical significance. Three patients with primary open angle glaucoma were also imaged to compare RPC density to controls. Results Qualitatively, vessels appeared smoother and closer to histologic descriptions with increasing number of averaged frames. Quantitatively, number of endpoints decreased by 51%, and SNR, mean capillary length, capillary density, and ICD increased by 44%, 91%, 11%, and 4.5% from single frame to 10-frame averaged, respectively. The 10-frame averaged images from the glaucomatous eyes revealed decreased density correlating to visual field defects and retinal nerve fiber layer thinning. Conclusions OCTA image registration and averaging is a viable and accessible method to enhance the visualization of RPCs, with significant improvements in image quality and RPC quantitative parameters. With this technique, we will be able to non-invasively and reliably study RPC involvement in diseases such as glaucoma.

PURPOSE To analyze the foveal microvasculature of young healthy eyes and older vasculopathic eyes, imaged using in vivo adaptive optics scanning light ophthalmoscope fluorescein angiography (AOSLO FA). METHODS AOSLO FA imaging of the superficial retinal microvasculature within an 800-μm radius from the foveal center was performed using simultaneous confocal infrared (IR) reflectance (790 nm) and fluorescence (488 nm) channels. Corresponding IR structural and FA perfusion maps were compared with each other to identify nonperfused capillaries adjacent to the foveal avascular zone. Microvascular densities were calculated from skeletonized FA perfusion maps. RESULTS Sixteen healthy adults (26 eyes; mean age 25 years, range, 21-29) and six patients with a retinal vasculopathy (six eyes; mean age 55 years, range, 44-70) were imaged. At least one nonperfused capillary was observed in five of the 16 healthy nonfellow eyes and in four of the six vasculopathic eyes. Compared with healthy eyes, capillary nonperfusion in the vasculopathic eyes was more extensive. Microvascular density of the 16 healthy nonfellow eyes was 42.0 ± 4.2 mm(-1) (range, 33-50 mm(-1)). All six vasculopathic eyes had decreased microvascular densities. CONCLUSIONS AOSLO FA provides an in vivo method for estimating foveal microvascular density and reveals occult nonperfused retinal capillaries. Nonperfused capillaries in healthy young adults may represent a normal variation and/or an early sign of pathology. Although limited, the normative data presented here is a step toward developing clinically useful microvascular parameters for ocular and/or systemic diseases.

PURPOSE To investigate the association between the size and shape of the foveal avascular zone and retinal thickness in healthy subjects. METHODS In vivo imaging of the foveal microvasculature was performed on 32 subjects by using an adaptive optics scanning laser ophthalmoscope (AOSLO). Motion contrast maps of the AOSLO images were used to generate a montage revealing the foveal capillary network. Foveal avascular zone (FAZ) diameters along the horizontal (FAZH) and vertical (FAZV) meridians were measured on the montages. An asymmetry index (AI) of the FAZ was then computed as the ratio of the FAZH to FAZV. Retinal thickness was investigated by using spectral-domain optical coherence tomography (SDOCT). Inner retinal layer (INLFAZ) thickness and outer nuclear layer (ONLFAZ) thickness were measured at the edges of the FAZ on the horizontal and vertical SDOCT scans on the same eye. RESULTS The foveal capillary network was readily visualized in all subjects. As expected there was individual variation in the size and shape of the FAZ. Along the horizontal and vertical meridians, the mean±SD (μm) of the FAZ diameter was 607±217 and 574±155, respectively. The INLFAZ thickness was 68±9 and 66±9, and the ONLFAZ thickness was 103±13 and 105±11, respectively. The mean±SD of the AI was 1.03±0.27. The difference between FAZH and FAZV decreases with increasing FAZ area (P=0.004). Mean ONLFAZ was negatively correlated with FAZ effective diameter (P

Purpose We evaluate the impact of age and signal strength index (SSI) on foveal avascular zone (FAZ) metrics and parafoveal capillary density measured using optical coherence tomography angiography (OCT-A), and propose a deviation mapping approach that accounts for age-group, SSI, eccentricity, and variation in FAZ size. Methods Parafoveal OCT-A with full vascular layer was obtained for 261 controls and four patients with retinal abnormalities. Parafoveal capillary densities were measured within eight consecutive 200-μm wide annuli from the FAZ border. In controls, the impacts of age and SSI on FAZ metrics and parafoveal capillary density were evaluated. Deviation maps highlighting regions with density at the lower and upper tails of the age-group and eccentricity matched distribution were generated. Results Linear regressions showed significant correlations between age, SSI, and mean parafoveal capillary density. There was a significant difference in FAZ metrics and parafoveal capillary densities with different age groups after controlling for SSI using univariate analysis. However, the effect of age on parafoveal capillary density disappeared after controlling for SSI using multivariate linear regression analysis. Our deviation mapping approach was able to identify regions with abnormal density in four patients. Conclusions Our findings suggest that the relationship between parafoveal capillary density and age is confounded by SSI. Parafoveal capillary density is SSI- and eccentricity-dependent. An age-group and eccentricity matched normative database was used as the basis for a parafoveal capillary density deviation mapping technique, providing an intuitive way to assess the status of parafoveal capillary density in individual eyes. Translational relevance Understanding the impact of age and SSI on parafoveal capillary density is critical for providing accurate interpretation of OCT-A. We demonstrate an age-group and eccentricity matched deviation mapping technique for an intuitive assessment of retinal regions with abnormal density.

Purpose The purpose of this study was to explore macula vessel density and foveal avascular zone (FAZ) parameters in exfoliation glaucoma (XFG) compared to primary open-angle glaucoma (POAG) using optical coherence tomography angiography (OCTA). Methods This was a cross-sectional observational study. Twenty-six XFG and 28 POAG patients with comparable visual field defects on Humphrey 24-2 and 10-2 perimetries were recruited. OCTA scans (3 × 3 mm) centered on the fovea were obtained. Built-in software was used to measure superficial capillary plexus (SCP) vessel density at different quadrants of the macula. Custom software was then used to create a full-thickness image. The FAZ was manually delineated, and large vessels were removed. Vessel density in eight concentric rings with increments of 200-μm diameters from the delineated FAZ was measured. FAZ parameters were calculated using the custom software. Results SCP density was significantly lower in the superior (mean difference [MD] = -4.32, 95% confidence interval [CI] = -7.02, -1.61, P = 0.003) and nasal (MD = -3.00, 95% CI = 05.22, -0.77, P = 0.010) quadrants in XFG versus POAG. SCP vessel density using the concentric ring approach revealed significantly decreased values at all eight rings in XFG versus POAG. In the full-thickness analysis, density was significantly less in the XFG group in all rings except the initial 200 μm. No significant differences existed in FAZ parameters between the groups. Conclusions Despite the presence of comparable central visual field defects, the macula vessel density was predominately lower in XFG compared with POAG in our sample of patients. Further studies are warranted to investigate the consistency of our results.

Purpose This study proposes an optical coherence tomography angiography (OCTA) frame-averaging method and investigates the effects of the number of frames acquired and averaged on metrics quantifying the foveal avascular zone (FAZ), vessel morphology, and parafoveal intercapillary area (PICA). Methods Ten OCTA frames were acquired for each of the 19 subjects without known retinal disease using the AngioVue OCTA system. For each subject, acquired frames were ranked by an image quality metric. A subset of frames was then registered and averaged. The effects of the number of frames acquired and averaged on FAZ segmentation and metrics of FAZ geometry, vessel morphology, and PICA were analyzed. Results Frame averaging increased the accuracy of the automatically segmented FAZ region; for example, the absolute error in FAZ area decreased from 0.026 mm2 (1 frame) to 0.005 mm2 (5 frames). Averaging multiple frames exponentially decreased the estimated number of vessel endpoints and increased the average vessel length with a 32% decrease in number of endpoints and 14% increase in average vessel length when averaging five frames compared with one. Frame averaging also improved the precision of PICA estimates. Conclusions Averaging multiple OCTA frames using the Optovue AngioVue system reduced error in FAZ segmentation and improved the robustness of OCTA vessel morphology and perfusion metrics. The study demonstrated limited benefit in acquiring and averaging more than five frames. Translational relevance Averaging multiple OCTA frames improved the robustness of OCTA foveal biomarkers with limited benefit when averaging more than five frames.

Given the complexity of the current system used to stage diabetic retinopathy (DR) and the risks and limitations associated with intravenous fluorescein angiography (IVFA), noninvasive quantification of DR severity is desirable. We examined the utility of acircularity index and axis ratio of the foveal avascular zone (FAZ), metrics that can noninvasively quantify the severity of diabetic retinopathy without the need for axial length to correct for individual retinal magnification. A retrospective review was performed of type 2 diabetics and age-matched controls imaged with optical coherence tomography angiography (OCTA). Diabetic eyes were divided into three groups according to clinical features: No clinically observable diabetic retinopathy (NoDR), nonproliferative diabetic retinopathy (NPDR), and proliferative diabetic retinopathy (PDR). OCTAs of the superficial and deep vascular layers centered at the fovea were superimposed to form a full vascular layer on which the FAZ was manually traced. Acircularity index and axis ratio were calculated for each FAZ. Significant differences in acircularity index were observed between all groups except for controls vs. NoDR. Similar results were found for axis ratio, although there was no significant difference observed between NPDR and PDR. We demonstrate that acircularity index and axis ratio can be used to help noninvasively stage DR using OCTA, and show promise as methods to monitor disease progression and detect response to treatment.

Large individual differences in cone densities occur even in healthy, young adults with low refractive error. We investigated whether cone density follows a simple model that some individuals have more cones, or whether individuals differ in both number and distribution of cones. We quantified cones in the eyes of 36 healthy young adults with low refractive error using a custom adaptive optics scanning laser ophthalmoscope. The average cone density in the temporal meridian was, for the mean ± SD, 43216 ± 6039, 27466 ± 3496, 14996 ± 1563, and 12207 ± 1278 cones /mm2 for 270, 630, 1480, and 2070 microns from the foveal center. Cone densities at 630 microns retinal eccentricity were uncorrelated to those at 2070 microns, ruling out models with a constant or proportional relation of cone density to eccentricity. Subjects with high central macula cone densities had low peripheral cone densities. The cone density ratio (2070 : 630 microns) was negatively correlated with cone density at 630 microns, consistent with variations in the proportion of peripheral cones migrating towards the center. We modelled the total cones within a central radius of 7 deg, using the temporal data and our published cone densities for temporal, nasal, superior, and inferior meridians. We computed an average of 221,000 cones. The coefficient of variation was 0.0767 for total cones, but higher for samples near the fovea. Individual differences occur both in total cones and other developmental factors related to cone distribution.

Purpose To characterise longitudinal changes in the retinal microvasculature of type 2 diabetes mellitus (T2DM) as exemplified in a patient with proliferative diabetic retinopathy (PDR) using an adaptive optics scanning light ophthalmoscope (AOSLO). Methods A 35-year-old T2DM patient with PDR treated with scatter pan-retinal photocoagulation at the inferior retina 1 day prior to initial AOSLO imaging along with a 24-year-old healthy control were imaged in this study. AOSLO vascular structural and perfusion maps were acquired at four visits over a 20-week period. Capillary diameter and microaneurysm area changes were measured on the AOSLO structural maps. Imaging repeatability was established using longitudinal imaging of microvasculature in the healthy control. Results Capillary occlusion and recanalisation, capillary dilatation, resolution of local retinal haemorrhage, capillary hairpin formation, capillary bend formation, microaneurysm formation, progression and regression were documented over time in a region 2° superior to the fovea in the PDR patient. An identical microvascular network with same capillary diameter was observed in the control subject over time. Conclusions High-resolution serial AOSLO imaging enables in vivo observation of vasculopathic changes seen in diabetes mellitus. The implications of this methodology are significant, providing the opportunity for studying the dynamics of the pathological process, as well as the possibility of identifying highly sensitive and non-invasive biomarkers of end organ damage and response to treatment.

Retinal vascular diseases are a leading cause of blindness and visual disability. The advent of adaptive optics retinal imaging has enabled us to image the retinal vascular at cellular resolutions, but imaging of the vasculature can be difficult due to the complex nature of the images, including features of many other retinal structures, such as the nerve fiber layer, glial and other cells. In this paper we show that varying the size and centration of the confocal aperture of an adaptive optics scanning laser ophthalmoscope (AOSLO) can increase sensitivity to multiply scattered light, especially light forward scattered from the vasculature and erythrocytes. The resulting technique was tested by imaging regions with different retinal tissue reflectivities as well as within the optic nerve head.

We evaluated the relationship between the size of the peripapillary crescent and the axial length (AL) of the eye as well as the fine structure of the peripapillary crescent in selected eyes. Infrared fundus imaging and spectral domain optical coherence tomography (SDOCT) (Spectralis HRA + OCT, Heidelberg Engineering, Germany) centered at the fovea were performed on 72 healthy adults. On the infrared fundus images, we measured (a) the distance between the foveola and the temporal edge of the optic disc (FOD) and (b) the distance between the foveola and the temporal edge of the peripapillary crescent (FOC) (if present). A peripapillary crescent presented at the nasal margin of the disc in 64% of the subjects. The FOD and FOC were 4.22 mm ± 0.46 and 3.97 mm ± 0.25, respectively. Only the FOD was significantly correlated with axial length. As AL increased by 10%, the FOD increased by 13%, the outer neural retina only expanded by 4% (as indicated by the FOC). This result emphasizes that retinal stretching may not mirror scleral growth, and the existence in some eyes of a difference between the photoreceptor margin and retinal pigment epithelium (RPE) margin suggests that within the retina there could be slippage during eye growth.

Enlargement of the foveal avascular zone (FAZ) due to progressive capillary nonperfusion is associated with visual deterioration in patients with diabetic retinopathy. The FAZ area has long been considered an important clinical marker of advancing retinopathy. However, a large body of literature shows that the FAZ area varies considerably in healthy eyes, resulting in substantial overlap between controls and diabetics, thus reducing its discriminatory value. In this study, within-subject FAZ area enlargement was obtained by the comparison of the structural FAZ area to the functional FAZ area using simultaneously-acquired, corresponding en face OCT reflectance and OCT angiography images. Our study suggests that en face OCT reflectance images provide useful anatomic baselines of structural FAZ morphology prior to the onset of disease. Measurements of within-subject FAZ area enlargement appear to be a more sensitive method for identifying the onset of diabetic retinopathy as compared to using OCT angiographic measurements of FAZ alone.

Purpose To describe a new technique for mapping parafoveal intercapillary areas (PICAs) using optical coherence tomography angiography (OCTA), and demonstrate its utility for quantifying parafoveal nonperfusion in diabetic retinopathy (DR). Methods Nineteen controls, 15 diabetics with no retinopathy (noDR), 15 with nonproliferative diabetic retinopathy (NPDR), and 15 with proliferative diabetic retinopathy (PDR) were imaged with 10 macular OCTA scans. PICAs were automatically delineated on the averaged superficial OCTA images. Following creation of an eccentricity-specific reference database from the controls, all PICAs greater than 2 SD above the reference means for PICA area and minor axis length were identified as nonperfused areas. Regions of interest (ROI) at 300 μm and 1000 μm from the foveal avascular zone (FAZ) margin were analyzed. Percent nonperfused area was defined as summed nonperfused areas divided by ROI area. Values were compared using Kruskal-Wallis and post-hoc Mann-Whitney U tests. Results Median values for total percent nonperfused area at the 300-μm ROI were 2.09, 2.44, 18.08, and 27.55 in the control, noDR, NPDR, and PDR groups, respectively. Median values at the 1000-μm ROI were 3.10, 3.31, 13.42, and 23.00. While there were no significant differences between the control and noDR groups, significant differences were observed between all other groups at both ROIs. Conclusions Percent nonperfused area can quantify parafoveal nonperfusion in DR and can be calculated through automatic delineation of PICAs in an eccentricity-specific manner using a standard deviation mapping approach. Translational relevance Percent nonperfused area shows promise as a metric to measure disease severity in diabetic retinopathy.

IMPORTANCE: This study used optical coherence tomographic angiography to assess for impaired blood flow in myopic eyes with or without open-angle glaucoma. OBJECTIVE: To compare the peripapillary perfused capillary density (PCD) between eyes with and without glaucoma. DESIGN, SETTING, AND PARTICIPANTS: In this cross-sectional study at a tertiary glaucoma referral practice, we recruited patients with myopic eyes of spherical equivalent of more than −3.0 diopters with and without open-angle glaucoma, patients with nonmyopic eyes with glaucoma, and patients with no disease from February 2016 to October 2016. We obtained 4.5 × 4.5-mm optical coherence tomographic angiography images of the optic nerve head and calculated PCD as the ratio of pixels associated with capillaries to the number of pixels in the region of interest after large blood vessel removal. Both eyes of each patient were used in the analysis. Continuous variables were assessed by analysis of variance and Tukey tests. A marginal model of generalized estimating equations was performed to adjust for confounding factors and intraclass correlations. MAIN OUTCOMES AND MEASURES: Mean PCD. RESULTS: We matched 87 patients with myopic eyes with glaucoma (of whom 39 [45%] were women), 17 with myopic eyes without glaucoma (of whom 10 [59%] were women), and 93 with non-myopic eyes with glaucoma (of whom 55 [59%] were women) for visual field defects and included 51 control participants (or whom 38 [75%] were women). Mean (SD) ages were 67.5 (12.0) years for patients with myopia and glaucoma, 48.2 (19.0) years for those with myopia without glaucoma, 67.3 (11.0) years for those with glaucoma without myopia, and 64.7 (8.9) years in control participants. Global PCD demonstrated a progressive decrease from the control group (mean [SD], 41.0 [4.2]) to those with myopia without glaucoma (38.4 [5.8]) to those with glaucoma without myopia (31.9 [7.5]) to those with both (28.2 [6.0]; all P

Purpose Eyes fellow to nonischemic central retinal vein occlusion (CRVO) were examined for abnormalities, which might explain their increased risk for future occlusion, using adaptive optics scanning light ophthalmoscope fluorescein angiography. Methods Adaptive optics scanning light ophthalmoscope fluorescein angiography foveal microvascular densities were calculated. Nonperfused capillaries adjacent to the foveal avascular zone were identified. Spectral domain optical coherence tomography, ultrawide field fluorescein angiographies, and microperimetry were also performed. Results Ten fellow eyes of nine nonischemic CRVO and 1 nonischemic hemi-CRVO subjects and four affected eyes of three nonischemic CRVO and one nonischemic hemi-CRVO subjects were imaged. Ninety percent of fellow eyes and 100% of affected eyes demonstrated at least 1 nonperfused capillary compared with 31% of healthy eyes. Fellow eye microvascular density (35 ± 3.6 mm(-1)) was significantly higher than that of affected eyes (25 ± 5.2 mm(-1)) and significantly lower than that of healthy eyes (42 ± 4.2 mm(-1)). Compared with healthy controls, spectral domain optical coherence tomography thicknesses showed no significant difference, whereas microperimetry and 2/9 ultrawide field fluorescein angiography revealed abnormalities in fellow eyes. Conclusion Fellow eye changes detectable on adaptive optics scanning light ophthalmoscope fluorescein angiography reflect subclinical pathology difficult to detect using conventional imaging technologies. These changes may help elucidate the pathogenesis of nonischemic CRVO and help identify eyes at increased risk of future occlusion.

In vivo microvascular imaging was performed on healthy and diseased retinas using an adaptive optics scanning light ophthalmoscope (AOSLO) employed with fluorescein angiography and non-confocal imaging techniques. With qualitative and quantitative image analyses, AOSLO provides a platform for characterizing retinal microvascular changes cross-sectionally and longitudinally due to disease progression or response to treatment.

International audience; Recent advances to the adaptive optics scanning light ophthalmoscope (AOSLO) have enabled finer in vivo assessment of the human retinal microvasculature. AOSLO confocal reflectance imaging has been coupled with oral fluorescein angiography (FA), enabling simultaneous acquisition of structural and perfusion images. AOSLO offset pinhole (OP) imaging combined with motion contrast post-processing techniques, are able to create a similar set of structural and perfusion images without the use of exogenous contrast agent. In this study, we evaluate the similarities and differences of the structural and perfusion images obtained by either method, in healthy control subjects and in patients with retinal vasculopathy including hypertensive retinopathy, diabetic retinopathy, and retinal vein occlusion. Our results show that AOSLO OP motion contrast provides perfusion maps comparable to those obtained with AOSLO FA, while AOSLO OP reflectance images provide additional information such as vessel wall fine structure not as readily visible in AOSLO confocal reflectance images. AOSLO OP offers a non-invasive alternative to AOSLO FA without the need for any exogenous contrast agent.

PURPOSE Microaneurysms (MAs) are considered a hallmark of retinal vascular disease, yet what little is known about them is mostly based upon histology, not clinical observation. Here, we use the recently developed adaptive optics scanning light ophthalmoscope (AOSLO) fluorescein angiography (FA) to image human MAs in vivo and to expand on previously described MA morphologic classification schemes. METHODS Patients with vascular retinopathies (diabetic, hypertensive, and branch and central retinal vein occlusion) were imaged with reflectance AOSLO and AOSLO FA. Ninety-three MAs, from 14 eyes, were imaged and classified according to appearance into six morphologic groups: focal bulge, saccular, fusiform, mixed, pedunculated, and irregular. The MA perimeter, area, and feret maximum and minimum were correlated to morphology and retinal pathology. Select MAs were imaged longitudinally in two eyes. RESULTS Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging revealed microscopic features of MAs not appreciated on conventional images. Saccular MAs were most prevalent (47%). No association was found between the type of retinal pathology and MA morphology (P = 0.44). Pedunculated and irregular MAs were among the largest MAs with average areas of 4188 and 4116 μm(2), respectively. Focal hypofluorescent regions were noted in 30% of MAs and were more likely to be associated with larger MAs (3086 vs. 1448 μm(2), P = 0.0001). CONCLUSIONS Retinal MAs can be classified in vivo into six different morphologic types, according to the geometry of their two-dimensional (2D) en face view. Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging of MAs offers the possibility of studying microvascular change on a histologic scale, which may help our understanding of disease progression and treatment response.

International audience; The adaptive optics scanning light ophthalmoscope (AOSLO) allows visualization of microscopic structures of the human retina in vivo. In this work, we demonstrate its application in combination with oral and intravenous (IV) fluorescein angiography (FA) to the in vivo visualization of the human retinal microvasculature. Ten healthy subjects ages 20 to 38 years were imaged using oral (7 and/or 20 mg/kg) and/or IV (500 mg) fluorescein. In agreement with current literature, there were no adverse effects among the patients receiving oral fluorescein while one patient receiving IV fluorescein experienced some nausea and heaving. We determined that all retinal capillary beds can be imaged using clinically accepted fluorescein dosages and safe light levels according to the ANSI Z136.1-2000 maximum permissible exposure. As expected, the 20 mg/kg oral dose showed higher image intensity for a longer period of time than did the 7 mg/kg oral and the 500 mg IV doses. The increased resolution of AOSLO FA, compared to conventional FA, offers great opportunity for studying physiological and pathological vascular processes.

Purpose To compare perfused peripapillary capillary density in primary open-angle glaucoma (POAG), normal-tension glaucoma (NTG), and normal patients using optical coherence tomography angiography (OCT-A). Methods A retrospective review of POAG, NTG, and normal patients imaged with OCT-A was performed. En face OCT angiograms identifying peripapillary vessels were obtained using a spectral-domain OCT system (Avanti RTVue-XR). A custom image analysis approach identified perfused peripapillary capillaries, quantified perfused capillary density (PCD), and generated color-coded PCD maps for 3.5- and 4.5-mm-diameter scans. We compared PCD values, PCD maps, standard automated perimetry (Humphrey visual field [HVF]) parameters, and OCT retinal nerve fiber layer (RNFL) thickness analyses across all groups. Results Forty POAG, 26 NTG, and 26 normal patients were included. Annular PCD in POAG (34.24 ± 6.76%) and NTG (37.75 ± 3.52%) patients was significantly decreased compared to normal patients (42.99 ± 1.81%) in 4.5-mm scans (P < 0.01 and P < 0.01, respectively). Similar trends and statistical significances were seen in 3.5-mm scans. Linear regression analysis resulted in moderate correlations between annular PCD values and other glaucomatous parameters. Pearson coefficients comparing annular PCD from 4.5-mm scans in POAG and NTG groups to HVF mean deviation, HVF pattern standard deviation, and average RNFL thickness all showed statistical significance (P < 0.05). Color maps showed that POAG and NTG patients had a reduction of perfused capillaries that progressed in size when comparing early, moderate, and severe glaucoma groups. Conclusions Optical coherence tomography angiography can uniquely identify changes in peripapillary PCD in glaucoma patients. Optical coherence tomography angiography may offer insights into the pathophysiology of glaucomatous damage and risk factors for disease progression.

Purpose Small artery and arteriolar walls thicken due to elevated blood pressure. Vascular wall thickness show a correlation with hypertensive subject history and risk for stroke and cardiovascular events. Methods The inner and outer diameter of retinal arterioles from less than 10 to over 150 μm were measured using a multiply scattered light adaptive optics scanning laser ophthalmoscope (AOSLO). These measurements were made on three populations, one with habitual blood pressures less than 100/70 mm Hg, one with normal blood pressures without medication, and one with managed essential hypertension. Results The wall to lumen ratio was largest for the smallest arterioles for all three populations. Data from the hypotensive group had a linear relationship between outer and inner diameters (r2 = 0.99) suggesting a similar wall structure in individuals prior to elevated blood pressures. Hypertensive subjects fell below the 95% confidence limits for the hypotensive relationship and had larger wall to lumen ratios and the normotensive group results fell between the other two groups. Conclusion High-resolution retinal imaging of subjects with essential hypertension showed a significant decrease in vessel inner diameter for a given outer diameter, and increases in wall to lumen ratio and wall cross-sectional areas over the entire range of vessel diameters and suggests that correcting for vessel size may improve the ability to identify significant vascular changes. Translational relevance High-resolution imaging allows precise measurement of vasculature and by comparing results across risk populations may allow improved identification of individuals undergoing hypertensive arterial wall remodeling.

Purpose To compare the use of optical coherence tomography angiography (OCTA) and adaptive optics scanning light ophthalmoscope fluorescein angiography (AOSLO FA) for characterizing the foveal microvasculature in healthy and vasculopathic eyes. Methods Four healthy controls and 11 vasculopathic patients (4 diabetic retinopathy, 4 retinal vein occlusion, and 3 sickle cell retinopathy) were imaged with OCTA and AOSLO FA. Foveal perfusion maps were semiautomatically skeletonized for quantitative analysis, which included foveal avascular zone (FAZ) metrics (area, perimeter, acircularity index) and vessel density in three concentric annular regions of interest. On each set of OCTA and AOSLO FA images, matching vessel segments were used for lumen diameter measurement. Qualitative image comparisons were performed by visual identification of microaneurysms, vessel loops, leakage, and vessel segments. Results Adaptive optics scanning light ophthalmoscope FA and OCTA showed no statistically significant differences in FAZ perimeter, acircularity index, and vessel densities. Foveal avascular zone area, however, showed a small but statistically significant difference of 1.8% (P = 0.004). Lumen diameter was significantly larger on OCTA (mean difference 5.7 μm, P < 0.001). Microaneurysms, fine structure of vessel loops, leakage, and some vessel segments were visible on AOSLO FA but not OCTA, while blood vessels obscured by leakage were visible only on OCTA. Conclusions Optical coherence tomography angiography is comparable to AOSLO FA at imaging the foveal microvasculature except for differences in FAZ area, lumen diameter, and some qualitative features. These results, together with its ease of use, short acquisition time, and avoidance of potentially phototoxic blue light, support OCTA as a tool for monitoring ocular pathology and detecting early disease.

Purpose To characterize the effects of intraframe distortion due to involuntary eye motion on measures of cone mosaic geometry derived from adaptive optics scanning light ophthalmoscope (AOSLO) images. Methods We acquired AOSLO image sequences from 20 subjects at 1.0, 2.0, and 5.0° temporal from fixation. An expert grader manually selected 10 minimally distorted reference frames from each 150-frame sequence for subsequent registration. Cone mosaic geometry was measured in all registered images (n = 600) using multiple metrics, and the repeatability of these metrics was used to assess the impact of the distortions from each reference frame. In nine additional subjects, we compared AOSLO-derived measurements to those from adaptive optics (AO)-fundus images, which do not contain system-imposed intraframe distortions. Results We observed substantial variation across subjects in the repeatability of density (1.2%-8.7%), inter-cell distance (0.8%-4.6%), percentage of six-sided Voronoi cells (0.8%-10.6%), and Voronoi cell area regularity (VCAR) (1.2%-13.2%). The average of all metrics extracted from AOSLO images (with the exception of VCAR) was not significantly different than those derived from AO-fundus images, though there was variability between individual images. Conclusions Our data demonstrate that the intraframe distortion found in AOSLO images can affect the accuracy and repeatability of cone mosaic metrics. It may be possible to use multiple images from the same retinal area to approximate a "distortionless" image, though more work is needed to evaluate the feasibility of this approach. Translational relevance Even in subjects with good fixation, images from AOSLOs contain intraframe distortions due to eye motion during scanning. The existence of these artifacts emphasizes the need for caution when interpreting results derived from scanning instruments.

We investigated the effect of localized visual stimulation on human retinal blood velocity using an adaptive optics scanning laser ophthalmoscope (AOSLO). To measure the blood velocity response, the AOSLO scanning raster was moved over the target arteries and red blood cell velocity was measured. Localized visual stimuli were delivered by projecting flicker patterns inside or outside the target artery's downstream region. The blood velocity increased in the presence of a flicker stimulus in the downstream region but not when outside the downstream region. The blood velocity increased more with larger area of stimulation. This increase was significant even when the stimulus was smaller than 600 μm × 600 μm. These findings suggest that when the retina regulates its blood flow to metabolic demands, it regulates blood velocity in the vascular system selectively, according to activity of neurons within its field of influence.

We evaluated the relationship between cone photoreceptor packing density and outer nuclear layer (ONL) thickness within the central 15 degrees.Individual differences for healthy subjects in cone packing density and ONL thickness were examined in 8 younger and 8 older subjects, mean age 27.2 versus 56.2 years. Cone packing density was obtained using an adaptive optics scanning laser ophthalmoscope (AOSLO). The ONL thickness measurements included the ONL and the Henle fiber layer (ONL + HFL), and were obtained using spectral domain optical coherence tomography (SDOCT) and custom segmentation software.There were sizeable individual differences in cone packing density and ONL + HFL thickness. Older subjects had on average lower cone packing densities, but thicker ONL + HFL measurements. Cone packing density and ONL + HFL thickness decreased with increasing retinal eccentricity. The ratio of the cone packing density-to-ONL2 was larger for the younger subjects group, and decreased with retinal eccentricity.The individual differences in cone packing density and ONL + HFL thickness are consistent with aging changes, indicating that normative aging data are necessary for fine comparisons in the early stages of disease or response to treatment. Our finding of ONL + HFL thickness increasing with aging is inconsistent with the hypothesis that ONL measurements with SDOCT depend only on the number of functioning cones, since in our older group cones were fewer, but thickness was greater.

Adaptive optics scanning laser ophthalmoscopes (AOSLO) allow real-time imaging of cellular and sub-cellular structure in the living human eye. In recent years, AOSLO systems have been employed for a growing range of scientific and clinical applications, including detailed measurements of the photoreceptors,1-4 vasculature,5-8 and other retinal structures.9-10 However, adaptive optics (AO) imaging systems typically have a field of view (FOV) of only a few degrees due to optical limitations11 and this can lead to problems of targeting the high resolution imaging to specified regions of interest on the retina. A typical approach is to image a recognizable landmark first and then move the imaging region towards the target region by asking the subject to fixate different targets in spatial sequence, often using a movable fixation spot or a fixation array. If a montage of the affected retinal region is required, the patient is asked to sequentially fixate a series of neighboring fixation targets or a movable target. Burns et.al 12 introduced a different approach by designing a steerable AOSLO combined with a wide-field line scanning ophthalmoscope (LSO). The steering angle in the most recent version is extended to almost 30 degrees of the posterior pole without requiring re-fixation by the subject.13 The wide-field imaging video can guide the high resolution imaging session in real-time. Both approaches are convenient for both the experimenter and the subject, but can be difficult to relate to other clinical imaging modalities. For the Burns’ approach,12,13 this arises because the wide field image quality was made subordinate to AOSLO image quality and thus identifying and navigating to specific clinical features can be difficult. Therefore, for both of these currently used approaches for AOSLO imaging, there can be difficulties in relating the very high resolution images to clinical images during the measurement session and thus be assured that the entire region of interest has been imaged. For this reason we developed a clinical planning module intended to relate regions of interest from a common clinical imaging system directly to the AOSLO steering system. This module would allow the experimenter to guide the AOSLO directly to regions of interest using a fundus image available in clinics, and then to rapidly acquire a montage of the region around that location without requiring re-fixation by the subject. The overall goal of this development was to simplify the imaging session for the patient and also to improve the efficiency and throughput of an AOSLO imaging session without sacrificing the high quality of AOSLO imaging that has been widely demonstrated. In the current paper we present the techniques involved and provide initial data generated using this clinical planning module.

The human fovea undergoes numerous changes during normal aging. Optical and psychophysical techniques allow us to measure these changes in vivo, using non-invasive techniques.