Purpose:To investigate the regional and layer-specific vascular reactivity of the healthy human retina and choriocapillaris to changes in systemic carbon dioxide or oxygen.Methods:High-resolution 3 × 3-mm2 optical coherence tomography angiography (OCTA) images were acquired from the central macula, temporal macula, and peripapillary retina while participants were exposed to three gas breathing conditions-room air, 5%CO2, and 100% O2. OCTA from all three regions were extracted and the apparent skeletonized vessel density (VSD) was assessed. The mean flow deficit sizes (MFDSs) of the choriocapillaris were also assessed. Repeated-measures analysis of variance was used to compare the ratio of intrasubject VSD change induced by the gas conditions from baseline in the superficial retinal layer (SRL) and deep retinal layer (DRL) for each retinal region independently, as well as the MFDS of the choriocapillaris. We also compared the vessel reactivity between the retinal capillaries and the choriocapillaris.Results:The cumulative intrasubject response to the gas conditions differed significantly among regions of the SRL (F(2, 7) = 28.22, P < 0.001), with the temporal macula showing the largest response (15%) compared to the macula (8%) and radial peripapillary capillaries (7%). A similar trend was found in the DRL. The choriocapillaris reactivity was similar between the macula (5.8%) and temporal macula (5.6%). There was also a significant heterogeneity in the layer-specific gas responses, with the DRL showing the largest response (28.2%) and the choriocapillaris showing the smallest response (2.8%).Conclusions:Capillary reactivity to changes in inhaled O2 and CO2 is spatially heterogeneous across the retina but not choriocapillaris.

Purpose: To compare optical coherence tomography angiography (OCTA)–derived flux with conventional OCTA measures of retinal vascular density in assessment of physiological changes in retinal blood flow. Methods: Healthy subjects were recruited, and 3 × 3-mm2 fovea-centered scans were acquired using commercially available swept-source OCTA (SS-OCTA) while participants were breathing room air, 100% O2, or 5% CO2. Retinal perfusion was quantified using vessel area density (VAD) and vessel skeleton density (VSD), as well as novel measures of retinal perfusion, vessel area flux (VAF) and vessel skeleton flux (VSF). Flux is proportional to the number of red blood cells moving through a vessel segment per unit time. The percentage change in each measure was compared between the O2 and CO2 gas conditions for images of all vessels (arterioles, venules, and capillaries) and capillary-only images. Statistical significance was determined using paired t-tests and a linear mixed-effects model. Results: Eighty-four OCTA scans from 29 subjects were included (age, 45.9 ± 19.5 years; 14 male, 48.3%). In capillary-only images, the change under the CO2 condition was 168% greater in VAF than in VAD (P = 0.002) and 124% greater in VSF than in VSD (P = 0.004). Similarly, under the O2 condition, the change was 94% greater in VAF than in VAD (P = 0.004) and 57% greater in VSF than in VSD (P = 0.01). Flux measures showed significantly greater change in capillary-only images compared with all-vessels images. Conclusions: OCTA-derived flux measures quantify physiological changes in retinal blood flow at the capillary level with a greater effect size than conventional vessel density measures. Translational Relevance: OCTA-derived flux is a useful measure of subclinical changes in retinal capillary perfusion.

PURPOSE:To assess continuity of perimetric defects corresponding to arcuate defects seen on optical coherence tomography (OCT) en face reflectance images of the retinal nerve fibre layer (RNFL) in patients with glaucoma. METHODS:Seven patients with glaucoma who had arcuate structural defects on OCT RNFL en face images were recruited. Static suprathreshold stimuli were presented along different meridians to localise perimetric defects in the corresponding hemifield. Then two contrasts, one 6 dB greater than the other, were used with kinetic perimetry to assess the slope of the defect. Findings with kinetic and 24-2 perimetry were compared. RESULTS:Static perimetry found that regions of perimetric abnormality spatially corresponded with the regions of en face RNFL hyporeflectivity. Kinetic perimetry found that the slopes of the edges of the defects ranged from 3-12 dB degree-1 , and that the functional abnormalities were continuous with the physiologic blind spot even when the 24-2 protocol only showed paracentral defects. CONCLUSIONS:Perimetric abnormalities and arcuate RNFL en face defects were spatially correspondent. Perimetric testing guided by OCT en face reflectance images can reveal greater functional detail of glaucomatous abnormality than 24-2 testing.

Purpose: This work assesses longitudinal microvascular changes in eyes treated with iodine-125 episcleral plaque brachytherapy (EPB). Methods: High-resolution optical coherence tomography angiograms (OCTAs) of the central 3 × 3-mm macula were obtained from iodine-125 EPB–treated and untreated fellow eyes of 61 patients. Previously validated semiautomated algorithms quantified capillary density (vessel skeleton density [VSD]) and caliber (vessel diameter index). Nonperfusion was also quantified as flow impairment region (FIR). Examinations from treated and fellow eyes obtained before treatment and at 6-month, 1-year, and 2-year intervals were compared using generalized estimating equation linear models. Dosimetry maps evaluated spatial correlation between radiation dose and microvascular metrics. Results: At 6 months, treated eyes had significantly lower VSD (0.145 ± 0.003 vs 0.155 ± 0.002; P = .009) and higher FIR (2.01 ± 0.199 vs 1.46 ± 0.104; P = .01) compared with fellow eyes. There was a significant decrease in VSD and a corresponding increase in FIR even for treated eyes without clinically identifiable retinopathy at 6 months. Vessel diameter index was significantly higher in treated eyes than in fellow eyes at 2 years (2.92 ± 0.025 vs 2.84 ± 0.018; P < .001). We categorized our cohort into low-dose (< 15 Gy) and high-dose (> 45 Gy) radiation to the fovea and noted significant differences in VSD and FIR between groups. Conclusions: OCTA can quantify and monitor EPB-induced retinopathy and can detect vascular abnormalities even without clinically observable retinopathy. OCTA may be useful in investigating treatment interventions aiming to delay EPB-induced radiation retinopathy.

Purpose To investigate layer specific retinal vascular reactivity (RVR) in capillaries of diabetic subjects without DR or with only mild non-proliferative diabetic retinopathy (NPDR). Methods A previously described nonrebreathing apparatus was used to deliver room air, 5% CO2, or 100% O2 to 41 controls and 22 diabetic subjects (with mild or no NPDR) while simultaneously acquiring fovea-centered 3x3mm2 Swept-Source Optical Coherence Tomography Angiography (SS-OCTA) images. Vessel skeleton density (VSD) and vessel diameter index (VDI) were calculated for each gas condition for the superficial retinal layer (SRL) and deep retinal layer (DRL). The superficial layer analysis excluded arterioles and venules. Data analysis was performed using mixed factorial analysis of covariance stratified by diabetic status. All models were adjusted for age, gender, and hypertension, and statistical significance for multiple comparisons from posthoc comparisons were defined at p<0.017. Results Among controls, there was a significant difference in capillary VSD between all gas conditions (p<0.001). This difference was present in both the SRL and DRL. Among diabetics, there was no significant difference in response to CO2 conditions in the SRL (p = 0.072), and a blunted response to both CO2 (p = 0.9) and O2 in the DRL (p = 0.019). A significant gas effect was detected in the capillary VDI in the SRL of controls (p = 0.001), which was driven by higher VDI in the oxygen condition compared to that of carbon dioxide. Conclusions Impairment in RVR in diabetic subjects is characterized by a paradoxical response to CO2 in both the SRL and DRL as well as an attenuated response to O2 in the DRL. These layer and gas specific impairments in diabetics seem to occur early in the disease and to be driven primarily at the capillary level.

We investigated the hypothesis that retinal capillary perfusion is a biomarker of early cognitive decline and cerebrovascular perfusion associated with small vessel disease in a pilot data set of Latinx adults at high risk for vascular cognitive impairment and dementia.

The vascular supply to the retina has been shown to dynamically adapt through vasoconstriction and vasodilation to accommodate the metabolic demands of the retina. This process, referred to as retinal vascular reactivity (RVR), is mediated by neurovascular coupling, which is impaired very early in retinal vascular diseases such as diabetic retinopathy. Therefore, a clinically feasible method of assessing vascular function may be of significant interest in both research and clinical settings. Recently, in vivo imaging of the retinal vasculature at the capillary level has been made possible by the FDA approval of optical coherence tomography angiography (OCTA), a noninvasive, minimal risk and dyeless angiography method with capillary level resolution. Concurrently, physiological and pathological changes in RVR have been shown by several investigators. The method shown in this manuscript is designed to investigate RVR using OCTA with no need for alterations to the clinical imaging procedures or device. It demonstrates real time imaging of the retina and retinal vasculature during exposure to hypercapnic or hyperoxic conditions. The exam is easily performed with two personnel in under 30 min with minimal subject discomfort or risk. This method is adaptable to other ophthalmic imaging devices and the applications may vary based on the composition of the gas mixture and patient population. A strength of this method is that it allows for an investigation of retinal vascular function at the capillary level in human subjects in vivo. Limitations of this method are largely those of OCTA and other retinal imaging methods including imaging artifacts and a restricted dynamic range. The results obtained from the method are OCT and OCTA images of the retina. These images are amenable to any analysis that is possible on commercially available OCT or OCTA devices. The general method, however, can be adapted to any form of ophthalmic imaging.

Background I-125 episcleral plaque brachytherapy (EPB) is standard-of-care for globe-conserving treatment of medium-sized choroidal melanomas. Radiation retinopathy is a potential consequence of treatment, characterized by deleterious effects on retinal microvasculature. We investigated the application of Optical Coherence Tomography Angiography (OCTA) for detecting and longitudinally monitoring I-125 episcleral plaque brachytherapy induced radiation retinopathy. Methods High resolution OCTA of the central 3×3mm macula were obtained from I-25 episcleral plaque brachytherapy treated and untreated fellow eyes of 62 patients. Capillary density (vessel skeleton density, VSD) and caliber (vessel diameter index, VDI) were quantified using previously validated semi-automated algorithms. Nonperfusion was also quantified as flow impairment regions (FIR). Exams from treated and fellow eyes obtained pre-treatment and at 6-month, 1-year, and 2-year intervals were compared using generalized estimating equation linear models. Dosimetry maps were used to evaluate spatial correlation between radiation dose and microvascular metrics. Results Mean time from treatment to last follow-up was 10.8 months. Mean±SD and median radiation dose at the fovea were 64.5 ± 76 Gy and 32.0 Gy, respectively. Preoperative logMAR (Snellen) mean visual acuity was 0.26 ± 0.05 (∼20/35) and 0.08 ± 0.02 (∼20/25) in treated and fellow eyes, respectively. At 6 months, treated eyes had significantly lower VSD (0.147 ± 0.003 vs 0.155 ± 0.002; p = 0.023) and higher FIR (1.95 ± 0.176 vs 1.45 ± 0.099; p = 0.018) compared to fellow eyes. There was a significant decrease in VSD and a corresponding increase in FIR even for treated eyes without clinically identifiable retinopathy at 6 months. VDI was significantly higher in treated eyes than in fellow eyes at 2 years (2.93 ± 0.022 vs 2.84 ± 0.016; p = 0.002). Microvascular changes were spatially correlated with a radiation gradient of 85-250 Gy across the fovea. Conclusions OCTA can be used to quantify and monitor EPB induced radiation, and can detect vascular abnormalities even in the absence of clinically observable retinopathy. OCTA may therefore be useful in investigating treatment interventions that aim to delay EPB-induced radiation retinopathy. * EPB : Episcleral Plaque Brachytherapy FA : Fluorescein Angiography FIR : Flow Impairment Region OCT(A) : Optical Coherence Tomography (Angiography) RR : Radiation Retinopathy VDI : Vessel Diameter Index VSD : Vessel Skeleton Density

Retinal findings are being investigated as indices of neurodegenerative disease. A cross-sectional, observational pilot study was conducted to investigate retinal changes in individuals with a mutation causing early-onset autosomal dominant Alzheimer disease (AD). Retinal images were examined in 14 subjects. 13 subjects had at least one first-degree relative with the A431E mutation in PSEN1; 1 subject had a first-degree relative with the V7171 mutation in APP. Not all subjects had all imaging modalities available. Mutation status and Clinical Dementia Rating Sum of Boxes (CDR-SOB) score, representing overall severity of dementia, were determined. Nonmutation-carrying subjects (NCs) and 11 healthy age-matched subjects (HCs) served as controls. Color fundus photographs (CF), ultra-widefield photographs (UWF), and autofluorescence images (AF) were inspected for fundus lesions. Retinal layer thicknesses were measured using optical coherence tomography (OCT). Retinal microvasculature was assessed using previously described vessel skeleton density (VSD) and vessel diameter index (VDI) metrics from OCTangiograms (OCTA). Generalized estimating equations and Pearson correlations were used to assess group differences. 10 subjects were mutation carriers (MCs; 1 homozygote, 9 heterozygotes); 4 were NCs. In MCs, global CDR scores ranged from normal to severe dementia (0-3), with the most severely affected MC being a homozygote. Hypopigmented and/or refractile lesions were identified in all MCs with CF images available (Fig 1). Nothing notable was observed on AF images. Mean temporal peripapillary RNFL thickness was greater in MCs compared with NCs (76 vs 65μm, P=0.031). However, mean retinal thickness negatively correlated with CDR-SOB score in the inferior region of the peripapillary RNFL and in the inner annulus surrounding the macula (P=0.002 and P=0.045, respectively; Fig 2). In 3x3mm2 OCTA images, mean VDI was greater in MCs compared with HCs (2.85 vs 2.83, P=0.012; Fig 3). No difference was observed in VSD between any groups.

Purpose: To assess retinal vascular reactivity in healthy controls and subjects with diabetic retinopathy (DR). Methods: A total of 22 healthy control eyes and 16 eyes with DR were enrolled. Images were acquired using a commercially available swept-source optical coherence tomography angiography (SS-OCTA) system. Three conditions were tested for each patient (hyperoxia, hypercapnia, and room-air) by employing a non-rebreathing apparatus that delivered appropriate gas mixtures (100% O2, 5% CO2, room air). Vessel skeleton density (VSD) and vessel diameter index (VDI) were compared between the conditions using mixed-model ANOVA adjusting for age and hypertension. Significant gas or interaction effects were followed by a Bonferroni adjusted pairwise post hoc analysis. Statistical significance was defined at P < 0.05. Results: The mixed-model ANOVA of the VSD found a significant intraindividual gas effect (F[2, 70] = 20.3, P < 0.001) and intergroup effect (F[1, 35] = 6.9, P = 0.001), and interaction effects (F[2, 70] = 4.6, P = 0.03). The post hoc pairwise comparison found significant differences among all three gas conditions in the healthy controls. In the subjects with DR, there were significant differences in VSD between hyperoxic and room air, and between hyperoxic and hypercapnic conditions, but not between hypercapnic and room-air conditions. Similar results were found for VDI. Conclusions: The retinal capillaries, assessed with SS-OCTA, in subjects with DR preferentially reacted to hyperoxia but not hypercapnia, while the healthy controls reacted to both. The difference in the vascular reactivity may be indicative of the underlying pathophysiology of DR.

Bright S. Ashimatey, Kyle M. Green, Zhongdi Chu, Ruikang K. Wang, and Amir H. Kashani Department of Ophthalmology, USC Roski Eye Institute, Keck School of Medicine of the University of Southern California, Los Angeles, California, United States Department of Bioengineering, University of Washington, Seattle, Washington, United States Department of Ophthalmology, University of Washington, Seattle, Washington, United States USC Ginsberg Institute for Biomedical Therapeutics, Los Angeles, California, United States

SIGNIFICANCE Glaucomatous nasal visual field abnormalities correspond to damage in the temporal raphe—where individual nerve bundles can be visualized. The ability to quantify structural abnormality in the raphe, with a clinically applicable protocol, sets the stage for investigating the raphe as a potential site for assessing early glaucoma. PURPOSE To develop a clinically applicable imaging and analysis technique for identifying retinal nerve fiber bundle abnormalities in the temporal raphe. METHODS Spectralis optical coherence tomography scans customized for the temporal raphe were gathered from 30 younger controls, 30 older controls, and 29 patients with glaucoma. An analysis technique was developed based on the reflectance of the nerve fiber bundles. The technique was first developed in the younger controls, and then applied to the older controls to generate normative data for quantifying nerve fiber bundle reflectance abnormalities in the patients with glaucoma. Matrix perimetric data were gathered in the patients with glaucoma to evaluate the reflectance technique’s findings. Reflectance abnormality in the patients was defined when the fraction of enface area showing reflectance abnormality was greater than the 95th percentile estimated from controls. Spearman’s rho was used to quantify the relation between the total deviation at the perimetric testing locations and the fraction of corresponding enface area showing reflectance abnormality. RESULTS Twenty-five of the 29 patients had reflectance abnormalities. Eight of these had mild to no perimetric mean deviation abnormality. Similar results were found when perimetric total deviations were compared to reflectance abnormalities in the corresponding enface locations. Spearman’s rho comparing the total deviations to reflectance abnormalities found rs(174) = −0.72, P < .001. CONCLUSIONS The technique typically identified reflectance abnormality when perimetric abnormality was present. It also identified reflectance abnormalities even when perimetric abnormality was mild or absent. The findings support the potential of raphe imaging in detecting early glaucomatous damage.

Optical coherence tomography (OCT) enface visualisation of the retinal nerve fibre layer (RNFL) reflectance has been found to have some advantages over retinal thickness measures. However, it is not yet clear how abnormalities on enface images relate to findings of abnormalities from other clinical measures such as the circumpapillary retinal nerve fibre layer thickness (cRNFLT). We developed a technique to analyse the RNFL reflectance on the OCT enface images, and to investigate its relation with the cRNFLT. Spectralis ( www.heidelbergengineering.com ) OCT scans of the central retinal ±24° were analysed in the study eye of 31 controls and 33 patients, ages 61 (±9) and 69 (±8) years respectively. Enface slab-images were extracted at 16–24, 24–36, and 24–52 μm from the inner limiting membrane in the temporal raphe, perifoveal and disc regions respectively. Reflectance probability maps were generated for the patients based on the control data. Glaucomatous abnormality was defined on the slab-images when the slab-area with reflectance abnormality was greater than the 95th percentile, and on the cRNFLT when the thickness measure was less than the fifth percentile, of that found in controls. The fraction of slab-image showing reflectance abnormality was compared to cRNFLT in the patient group, using Spearman's rho. Agreement between the findings of abnormality based on cRNFLT and slab-image reflectance was assessed using Cohen's kappa. Slab-image and cRNFLT findings were in agreement for 26/33 eyes; four subjects showed cRNFLT abnormality but not slab-image abnormality, and three subjects showed slab-image abnormality but not cRNFLT abnormality. Spearman's rho found r s (31) = −0.82. The reflectance findings and cRNFLT findings were consistent in 27/33 for both the superior temporal (ST) and inferior temporal (IT) sectors, and Cohen's kappa found 0.53 and 0.61 respectively. The surface area of enface slab-images showing RNFL reflectance were strongly related to the cRNFLT measures, and the classification of a subject with glaucoma based on enface reflectance findings and cRNFLT findings had a generally good agreement. The larger retinal area assessed by the enface method preserves the spatial location of the RNFL abnormalities, and makes the technique a useful approach for identifying regions of potential RNFL abnormality for targeted perimetry.