Disruption of the clock components CLOCK and BMAL1 leads to hypoinsulinaemia and diabetes.

The molecular clock maintains energy constancy by producing circadian oscillations of rate-limiting enzymes involved in tissue metabolism across the day and night. During periods of feeding, pancreatic islets secrete insulin to maintain glucose homeostasis, and although rhythmic control of insulin release is recognized to be dysregulated in humans with diabetes, it is not known how the circadian clock may affect this process. Here we show that pancreatic islets possess self-sustained circadian gene and protein oscillations of the transcription factors CLOCK and BMAL1. The phase of oscillation of islet genes involved in growth, glucose metabolism and insulin signalling is delayed in circadian mutant mice, and both Clock and Bmal1 (also called Arntl) mutants show impaired glucose tolerance, reduced insulin secretion and defects in size and proliferation of pancreatic islets that worsen with age. Clock disruption leads to transcriptome-wide alterations in the expression of islet genes involved in growth, survival and synaptic vesicle assembly. Notably, conditional ablation of the pancreatic clock causes diabetes mellitus due to defective beta-cell function at the very latest stage of stimulus-secretion coupling. These results demonstrate a role for the beta-cell clock in coordinating insulin secretion with the sleep-wake cycle, and reveal that ablation of the pancreatic clock can trigger the onset of diabetes mellitus.

Proteus mirabilis Keratitis: Risk Factors, Clinical Features, Treatment Outcomes, and Microbiological Characteristics.

PURPOSE: To characterize the risk factors, clinical presentations, management choices, and outcomes of Proteus mirabilis keratitis. METHODS: In this retrospective study, 26 culture-proven cases of P. mirabilis infections were diagnosed and treated between 1998 and 2019 at the University of Pittsburgh Medical Center. Medical records were available for 14 keratitis cases and were reviewed for demographic information, ocular risk factors, and treatment outcomes. RESULTS: Sixteen eyes of 14 patients were included in the study. The average age was 47.8 ± 19.3 years, with a median follow-up time of 6 months. The most common ocular risk factors were poor ocular surface and contact lens use in 57.1% and 42.9% of cases, respectively. Eleven of the 14 patients (78.6%) had positive corneal cultures, and 13 of the 14 patients (92.9%) had positive conjunctiva or eyelid cultures. All isolates were susceptible to ciprofloxacin, ofloxacin, moxifloxacin, gatifloxacin, and cefazolin. Surgical intervention was required in 4 patients (28.6%). Average LogMAR visual acuity was 1.3 ± 1.0 at presentation and 0.9 6 ± 1.0 at the most recent follow-up visit. CONCLUSIONS: Proteus mirabilis is an uncommon cause of microbial keratitis. Patients with poor ocular surface and those who use contact lens are at increased risk for developing this cause of keratitis. Empiric treatment with fortified antibiotics or fluoroquinolones seemed to provide effective coverage for P. mirabilis.

Earliest Evidence of Preclinical Diabetic Retinopathy Revealed Using Optical Coherence Tomography Angiography Perfused Capillary Density.

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.

Parafoveal Nonperfusion Analysis in Diabetic Retinopathy Using Optical Coherence Tomography Angiography.

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.

Within-subject assessment of foveal avascular zone enlargement in different stages of diabetic retinopathy using en face OCT reflectance and OCT angiography.

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.

A method for age-matched OCT angiography deviation mapping in the assessment of disease- related changes to the radial peripapillary capillaries.

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<0.01). Average healthy LPV density was 21.5±3.07%. Linear regression models indicated that LPV density decreased with age, however ANOVA and pairwise Tukey-Kramer tests did not reach statistical significance. Deviation mapping enabled us to quantitatively and visually elucidate the significance of RPC density changes in disease. CONCLUSIONS: It is important to consider changes that occur with aging when analyzing RPC and LPV density changes in disease. RPC density, coupled with age-matched deviation mapping techniques, represents a potentially clinically useful method in detecting changes to peripapillary perfusion in disease.

Noninvasive Detection of Mitochondrial Dysfunction in Ocular Hypertension and Primary Open-angle Glaucoma.

PURPOSE: To assess mitochondrial dysfunction in vivo in ocular hypertension (OHT) and primary open-angle glaucoma (POAG) using retinal metabolic analysis. PATIENTS AND METHODS: This was an observational, cross-sectional study performed from November 2015 to October 2016 at the New York Eye and Ear Infirmary of Mount Sinai. Thirty-eight eyes with varying stages of POAG, 16 eyes with OHT, and 32 control eyes were imaged on a custom fundus camera modified to measure full retinal thickness fluorescence at a wavelength optimized to detect flavoprotein fluorescence (FPF). Optical coherence tomography was used to measure the retinal ganglion cell-plus layer (RGC+) thickness. Macular FPF and the ratio of macular FPF to RGC+ thickness were the primary outcome variables and were compared among the three groups using an age-adjusted linear regression model. A mixed-effects model was used to assess correlations between FPF variables and clinical characteristics. RESULTS: Both macular FPF and the macular FPF/RGC+ thickness ratio were significantly increased in OHT compared with control eyes (P<0.05 and <0.01, respectively). In POAG eyes, macular FPF was not significantly increased compared with controls (P=0.24). However, the macular FPF/RGC+ thickness ratio in POAG eyes was significantly increased compared with controls (P<0.001). FPF was significantly correlated to age in POAG eyes. CONCLUSIONS: Despite lacking clinical evidence of glaucomatous deterioration, OHT eyes displayed significantly elevated macular FPF, suggesting that mitochondrial dysfunction may be detected before structural changes visible on current clinical imaging. Our preliminary results suggest that macular FPF analysis may prove to be a useful tool in assessing and evaluating OHT and POAG eyes.

Acircularity index and axis ratio of the foveal avascular zone in diabetic eyes and healthy controls measured by optical coherence tomography angiography.

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.

Visualization of Radial Peripapillary Capillaries Using Optical Coherence Tomography Angiography: The Effect of Image Averaging.

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.

Peripapillary perfused capillary density in primary open-angle glaucoma across disease stage: an optical coherence tomography angiography study.

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 Pearson's 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.

Human retinal microvascular imaging using adaptive optics scanning light ophthalmoscopy.

BACKGROUND: Retinal microvascular imaging is an especially promising application of high resolution imaging since there are increasing options for therapeutic intervention and need for better structural and functional biomarkers to characterize ocular and systemic vascular diseases. MAIN BODY: Adaptive optics scanning light ophthalmoscopy (AOSLO) is an emerging technology for improving in vivo imaging of the human retinal microvasculature, allowing unprecedented visualization of retinal microvascular structure, measurements of blood flow velocity, and microvascular network mapping. This high resolution imaging technique shows significant potential for studying physiological and pathological conditions of the retinal microvasculature noninvasively. CONCLUSION: This review will briefly summarize the abilities of in vivo human retinal microvasculature imaging in healthy controls, as well as patients with diabetic retinopathy, retinal vein occlusion, and sickle cell retinopathy using AOSLO and discuss its potential contribution to scientific research and clinical applications.

Imaging Foveal Microvasculature: Optical Coherence Tomography Angiography Versus Adaptive Optics Scanning Light Ophthalmoscope Fluorescein Angiography.

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.

Optical Coherence Tomography Angiography Analysis of Perfused Peripapillary Capillaries in Primary Open-Angle Glaucoma and Normal-Tension Glaucoma.

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.