The 100 most-cited articles in Investigative Ophthalmology & Visual Science, 2019

Purpose: We provide a standardized set of terminology, definitions, and thresholds of myopia and its main ocular complications. Methods: Critical review of current terminology and choice of myopia thresholds was done to ensure that the proposed standards are appropriate for clinical research purposes, relevant to the underlying biology of myopia, acceptable to researchers in the field, and useful for developing health policy. Results: We recommend that the many descriptive terms of myopia be consolidated into the following descriptive categories: myopia, secondary myopia, axial myopia, and refractive myopia. To provide a framework for research into myopia prevention, the condition of "pre-myopia" is defined. As a quantitative trait, we recommend that myopia be divided into myopia (i.e., all myopia), low myopia, and high myopia. The current consensus threshold value for myopia is a spherical equivalent refractive error ≤ -0.50 diopters (D), but this carries significant risks of classification bias. The current consensus threshold value for high myopia is a spherical equivalent…

The results of many studies in a variety of species have significantly advanced our understanding of the role of visual experience and the mechanisms of postnatal eye growth, and the development of myopia.This paper surveys and reviews the major contributions that experimental studies using animal models have made to our thinking about emmetropization and development of myopia.These studies established important concepts informing our knowledge of the visual regulation of eye growth and refractive development and have transformed treatment strategies for myopia.Several major findings have come from studies of experimental animal models.These include the eye's ability to detect the sign of retinal defocus and undergo compensatory growth, the local retinal control of eye growth, regulatory changes in choroidal thickness, and the identification of components in the biochemistry of eye growth leading to the characterization of signal cascades regulating eye growth and refractive state.Several of these findings provided the proofs of concepts that form the scientific basis of new and…

Myopia has been predicted to affect approximately 50% of the world's population based on trending myopia prevalence figures. Critical to minimizing the associated adverse visual consequences of complicating ocular pathologies are interventions to prevent or delay the onset of myopia, slow its progression, and to address the problem of mechanical instability of highly myopic eyes. Although treatment approaches are growing in number, evidence of treatment efficacy is variable. This article reviews research behind such interventions under four categories: optical, pharmacological, environmental (behavioral), and surgical. In summarizing the evidence of efficacy, results from randomized controlled trials have been given most weight, although such data are very limited for some treatments. The overall conclusion of this review is that there are multiple avenues for intervention worthy of exploration in all categories, although in the case of optical, pharmacological, and behavioral interventions for preventing or slowing progression of myopia, treatment efficacy at an individual level…

The knowledge on the genetic background of refractive error and myopia has expanded dramatically in the past few years. This white paper aims to provide a concise summary of current genetic findings and defines the direction where development is needed. We performed an extensive literature search and conducted informal discussions with key stakeholders. Specific topics reviewed included common refractive error, any and high myopia, and myopia related to syndromes. To date, almost 200 genetic loci have been identified for refractive error and myopia, and risk variants mostly carry low risk but are highly prevalent in the general population. Several genes for secondary syndromic myopia overlap with those for common myopia. Polygenic risk scores show overrepresentation of high myopia in the higher deciles of risk. Annotated genes have a wide variety of functions, and all retinal layers appear to be sites of expression. The current genetic findings offer a world of new molecules involved in myopiagenesis. As the missing heritability is still large, further genetic advances are needed.…

Best practice clinical guidelines for myopia control involve an understanding of the epidemiology of myopia, risk factors, visual environment interventions, and optical and pharmacologic treatments, as well as skills to translate the risks and benefits of a given myopia control treatment into lay language for both the patient and their parent or caregiver. This report details evidence-based best practice management of the pre-, stable, and the progressing myope, including risk factor identification, examination, selection of treatment strategies, and guidelines for ongoing management. Practitioner considerations such as informed consent, prescribing off-label treatment, and guides for patient and parent communication are detailed. The future research directions of myopia interventions and treatments are discussed, along with the provision of clinical references, resources, and recommendations for continuing professional education in this growing area of clinical practice.

Purpose: In this study, we aim to investigate whether mesenchymal stem cell (MSC)-derived exosomes (MSC-Exos) could regulate hyperglycemia-induced retinal inflammation by transferring microRNA-126 (miR-126). Methods: MSC-Exos were isolated from the media of human umbilical cord-derived mesenchymal stem cells (hUCMSCs), and this isolation was followed by the transfer of miR-126. MSC-Exos or MSC-Exos overexpressing miR-126 were intravitreally injected into diabetic rats in vivo and were cocultured with high glucose-affected human retinal endothelial cells (HRECs) in vitro. Plasma samples were obtained from the vitreous of rats and from HREC cells after treatment for ELISA assay. Retinal sections were examined using immunohistochemistry. RT-PCR and Western blotting were conducted to assess the levels of high-mobility group box 1 (HMGB1), NLRP3 inflammasome, and NF-κB/P65 in retinas and HRECs. Results: Our results showed that hyperglycemia greatly increased inflammation in diabetic rats or HRECs exposed to high glucose, increasing the levels of caspase-1, interleukin-1β (IL-1β) and…

Purpose: The purpose of this study was to study changes in choroidal thickness (ChT) and choroidal blood perfusion (ChBP), and the correlation between them, in guinea pig myopia. Methods: The reliability of optical coherence tomography angiography (OCTA) for measuring ChT and ChBP was verified in guinea pigs, after cervical dislocation (n = 7) or temporal ciliary artery transection (n = 6). Changes in refraction, axial length, ChT, and ChBP were measured during spontaneous myopia (n = 9), monocular form-deprivation myopia (FDM, n = 13), or lens-induced myopia (LIM, n = 14), and after 4 days of recovery from FDM and LIM. Results: The abolition (by cervical dislocation) or reduction (by temporal ciliary artery transection) of ChBP, and of the associated changes in ChT, were verified by OCTA, thus validating the method of measurement. In the spontaneous myopia group, ChT and ChBP were reduced by 25.2% and 31.9%, respectively. In FDM eyes, mean ± SD ChT and ChBP decreased significantly compared with the untreated fellow eyes (ChT fellow: 76.13 ± 9.34 μm versus 64.76 ± 11.15 μm for FDM;…

With the growing prevalence of myopia, already at epidemic levels in some countries, there is an urgent need for new management approaches. However, with the increasing number of research publications on the topic of myopia control, there is also a clear necessity for agreement and guidance on key issues, including on how myopia should be defined and how interventions, validated by well-conducted clinical trials, should be appropriately and ethically applied. The International Myopia Institute (IMI) reports the critical review and synthesis of the research evidence to date, from animal models, genetics, clinical studies, and randomized controlled trials, by more than 85 multidisciplinary experts in the field, as the basis for the recommendations contained therein. As background to the need for myopia control, the risk factors for myopia onset and progression are reviewed. The seven generated reports are summarized: (1) Defining and Classifying Myopia, (2) Experimental Models of Emmetropization and Myopia, (3) Myopia Genetics, (4) Interventions for Myopia Onset and Progression, (5)…

The evidence-basis based on existing myopia control trials along with the supporting academic literature were reviewed; this informed recommendations on the outcomes suggested from clinical trials aimed at slowing myopia progression to show the effectiveness of treatments and the impact on patients. These outcomes were classified as primary (refractive error and/or axial length), secondary (patient reported outcomes and treatment compliance), and exploratory (peripheral refraction, accommodative changes, ocular alignment, pupil size, outdoor activity/lighting levels, anterior and posterior segment imaging, and tissue biomechanics). The currently available instrumentation, which the literature has shown to best achieve the primary and secondary outcomes, was reviewed and critiqued. Issues relating to study design and patient selection were also identified. These findings and consensus from the International Myopia Institute members led to final recommendations to inform future instrumentation development and to guide clinical trial protocols.

Myopia is fast becoming a global public health burden with its increasing prevalence, particularly in developed countries. Globally, the prevalence of myopia and high myopia (HM) is 28.3% and 4.0%, respectively, and these numbers are estimated to increase to 49.8% for myopia and 9.8% for HM by 2050 (myopia defined as -0.50 diopter [D] or less, and HM defined as -5.00 D or less). The burden of myopia is tremendous, as adults with HM are more likely to develop pathologic myopia (PM) changes that can lead to blindness. Accordingly, preventive measures are necessary for each step of myopia progression toward vision loss. Approaches to prevent myopia-related blindness should therefore attempt to prevent or delay the onset of myopia among children by increased outdoor time; retard progression from low/mild myopia to HM, through optical (e.g., defocus incorporated soft contact lens, orthokeratology, and progressive-additional lenses) and pharmacological (e.g., low dose of atropine) interventions; and/or retard progression from HM to PM through medical/surgical treatments (e.g., anti-VEGF…

Purpose: We studied biometry changes before and after myopia onset in a cohort of Singaporean children. Methods: All data were taken from the Singapore Cohort Study of the Risk Factors for Myopia (SCORM). Participants underwent refraction and biometry measurements with a follow-up of 3 to 6 years. The longitudinal ocular biometry (spherical equivalent refraction, axial length, and lens power) changes were compared between children who suffered myopia during the study (N = 303), emmetropic children (N = 490), and children myopic at baseline (N = 509). Results: At myopia onset, the myopic shift increased to 0.50 diopters (D)/y or more in new myopes compared to the minor changes in emmetropes of the same age. New myopes had higher axial growth rates than emmetropes, even years before myopia onset (0.37 and 0.14 mm/y, respectively; ANOVA with Bonferroni post hoc test, P < 0.001). After onset, the change in both parameters slowed down gradually, but significantly (P < 0.05). In new myopes, lens power loss (-0.71 D/y) was significantly higher up to 1 year before myopia onset compared to…

Purpose: We investigate macular perfusion and the systemic and ocular associations in a population-based setting. Methods: In this cross-sectional study, 2018 adults residing in Hong Kong underwent detailed ophthalmic examinations after consenting to participate. Macular perfusion was measured with optical coherence tomography angiography (OCTA) using the split-spectrum amplitude decorrelation angiography algorithm. The parafoveal flow index and vessel area density were quantified using automated custom-built software. Results: Of the 2018 participants, the OCTA measurements were available for 1940, and 1631 (84.1%) had good quality scans. The right eyes of these 1631 participants (43.1% men) were included for final analysis. Mean age was 49.8 years (range, 18-92 years). Mean global macular vessel density was 47.3% and 55.1% for the superficial and deep retinal layers, respectively. In multivariate analysis, lower superficial vessel density remained significantly associated with lower signal strength index (SSI; P < 0.001, standardized β = 0.607) and male sex (P < 0.001, β = 0.162),…

Purpose: Single-cell RNA-sequencing (scRNA-seq) was used to interrogate the relatively rare stem (SC) and early transit amplifying (TA) cell populations in limbal/corneal epithelia from wild-type and autophagy-compromised mice. Methods: We conducted scRNA-seq on ocular anterior segmental tissue from wild-type and beclin 1-deficient (beclin1+/-) mice, using a 10X Gemomics pipeline. Cell populations were distinguished by t-distributed stochastic neighbor embedding. Seurat analysis was conducted to compare gene expression profiles between these two groups of mice. Differential protein expression patterns were validated by immunofluorescence staining and immunoblotting. Results: Unbiased clustering detected 10 distinct populations: three clusters of mesenchymal and seven clusters of epithelial cells, based on their unique molecular signatures. A discrete group of mesenchymal cells expressed genes associated with corneal stromal SCs. We identified three limbal/corneal epithelial cell subpopulations designated as stem/early TA, mature TA, and differentiated corneal epithelial cells.…

Purpose: To elucidate the mechanism whereby miR-590-3p regulates pyroptosis in diabetic retinopathy (DR). Methods: Human retinal microvascular endothelial cells (HRMECs) incubated with high glucose (HG) were used to establish cell models, and the expression levels of miR-590-3p, caspase-1, IL-1β, NLRP1, NOX4, TXNIP, NLRP3, and ROS were determined. Additionally, miR-590-3p was altered using a mimic or an inhibitor, and siRNAs targeting NLRP1 and NOX4 were applied to explore the regulatory mechanism of miR-590-3p in DR. The relationships between miR-590-3p and NLRP1/NOX4 also were investigated using a luciferase reporter assay. Furthermore, vitreous tissue samples were collected to confirm pyroptosis in clinical DR. Results: Downregulated miR-590-3p and upregulated NLRP1/NOX4 levels were observed in a cell culture model of DR. Inhibiting miR-590-3p upregulated NLRP1, the NOX4/ROS/TXNIP/NLRP3 pathway, and caspase-1. NLRP1 and NOX4 were confirmed as direct target genes of miR-590-3p. The overexpression of miR-590-3p or knockdown of NLRP1 and NOX4 increased cell activity and suppressed…

Purpose: Previous studies reported that hyperlipidemia and blood lipid levels were associated with glaucoma, ocular hypertension (OHT), and intraocular pressure (IOP). However, studies aimed at investigating this association have yielded conflicting results. Therefore, to shed light on these inconclusive findings, we performed multiple distinct meta-analyses to clarify the association of hyperlipidemia and blood lipid levels with glaucoma, OHT, and IOP. Methods: A systematic literature search from Embase, Web of Science, and PubMed was performed to identify relevant studies. To assess the association between hyperlipidemia and glaucoma, we used the pooled odds ratio (OR) with 95% confidence interval (CI). When we assessed the association between blood lipid levels and IOP levels, the pooled mean difference in IOP associated with a 10 mg/dL increase in the blood lipid level was estimated. The pooled difference in IOP was also estimated between patients with and without hyperlipidemia. All the papers that assessed the correlation between hyperlipidemia and glaucoma, between blood…

Purpose: Comparison of retinal microvasculature within the macula and the optic nerve head in the eyes of patients with Alzheimer's disease (AD), primary open-angle glaucoma (POAG), and in a healthy control (HC) group, using optical coherence tomography angiography (OCTA). Methods: In this cross-sectional study, 27 patients with AD, 27 with POAG, and 27 healthy controls were enrolled. The Mini-Mental State Examination test was used to assess cognitive function. Ophthalmic examination included OCTA, which was used for the imaging of vascular flow within the layer of radial peripapillary capillaries (RPCs), and also in the superficial vascular plexus (SVP) and deep vascular plexus (DVP) of the retina. Results: In the AD group, the density of vessels in DVP was significantly reduced and the foveal avascular zone was increased when compared to POAG and HC groups (P < 0.001). Patients with POAG had a significantly reduced vessel density in RPCs and SVP as compared to AD and HC groups (P < 0.001). The average thickness of peripapillary retinal nerve fiber layer was correlated with the…

Purpose: To detect visual field (VF) progression by analyzing spatial pattern changes. Methods: We selected 12,217 eyes from 7360 patients with at least five reliable 24-2 VFs and 5 years of follow-up with an interval of at least 6 months. VFs were decomposed into 16 archetype patterns previously derived by artificial intelligence techniques. Linear regressions were applied to the 16 archetype weights of VF series over time. We defined progression as the decrease rate of the normal archetype or any increase rate of the 15 VF defect archetypes to be outside normal limits. The archetype method was compared with mean deviation (MD) slope, Advanced Glaucoma Intervention Study (AGIS) scoring, Collaborative Initial Glaucoma Treatment Study (CIGTS) scoring, and the permutation of pointwise linear regression (PoPLR), and was validated by a subset of VFs assessed by three glaucoma specialists. Results: In the method development cohort of 11,817 eyes, the archetype method agreed more with MD slope (kappa: 0.37) and PoPLR (0.33) than AGIS (0.12) and CIGTS (0.22). The most frequently progressed…

Purpose: Glaucoma is more common in urban populations than in others. Ninety percent of the world's population are exposed to air pollution above World Health Organization (WHO) recommended limits. Few studies have examined the association between air pollution and glaucoma. Methods: Questionnaire data, ophthalmic measures, and ambient residential area air quality data for 111,370 UK Biobank participants were analyzed. Particulate matter with an aerodynamic diameter < 2.5 μm (PM2.5) was selected as the air quality exposure of interest. Eye measures included self-reported glaucoma, intraocular pressure (IOP), and average thickness of macular ganglion cell-inner plexiform layer (GCIPL) across nine Early Treatment Diabetic Retinopathy Study (ETDRS) retinal subfields as obtained from spectral-domain optical coherence tomography. We examined the associations of PM2.5 concentration with self-reported glaucoma, IOP, and GCIPL. Results: Participants resident in areas with higher PM2.5 concentration were more likely to report a diagnosis of glaucoma (odds ratio = 1.06, 95% confidence…

Purpose: To investigate the plasma concentration of nicotinamide in primary open-angle glaucoma (POAG). Methods: Plasma of 34 POAG individuals was compared to that of 30 age- and sex-matched controls using a semiquantitative method based on liquid chromatography coupled to high-resolution mass spectrometry. Subsequently, an independent quantitative method, based on liquid chromatography coupled to mass spectrometry, was used to assess nicotinamide concentration in the plasma from the same initial cohort and from a replicative cohort of 20 POAG individuals and 15 controls. Results: Using the semiquantitative method, the plasma nicotinamide concentration was significantly lower in the initial cohort of POAG individuals compared to controls and further confirmed in the same cohort, using the targeted quantitative method, with mean concentrations of 0.14 μM (median: 0.12 μM; range, 0.06-0.28 μM) in the POAG group (-30%; P = 0.022) and 0.19 μM (median: 0.18 μM; range, 0.08-0.47 μM) in the control group. The quantitative dosage also disclosed a significantly lower plasma nicotinamide…

Purpose: AMD is the leading cause of irreversible blindness in older individuals in the Western world, and there are currently no therapies to halt disease progression. Studies suggest that the commonly prescribed antidiabetic drug, metformin, is associated with decreased risk of several ocular diseases, but no work has investigated the effect of metformin use on development of AMD. Thus, we aim to investigate whether metformin use is associated with decreased risk of developing AMD. Methods: In this retrospective case-control study, we used medical records from patients older than 55 who have visited a University of Florida health clinic. Three controls were matched for every AMD case, defined by International Classification of Diseases, Ninth Revision code, based on the Charlson Comorbidity Index to ensure comparable baseline overall health status. Univariate and conditional multivariable logistic regressions were used to determine the association between a variety of covariates, including metformin use, and AMD diagnosis. Results: Metformin use was associated with decreased odds…

Purpose: To develop and assess a method for predicting the likelihood of converting from early/intermediate to advanced wet age-related macular degeneration (AMD) using optical coherence tomography (OCT) imaging and methods of deep learning. Methods: Seventy-one eyes of 71 patients with confirmed early/intermediate AMD with contralateral wet AMD were imaged with OCT three times over 2 years (baseline, year 1, year 2). These eyes were divided into two groups: eyes that had not converted to wet AMD (n = 40) at year 2 and those that had (n = 31). Two deep convolutional neural networks (CNN) were evaluated using 5-fold cross validation on the OCT data at baseline to attempt to predict which eyes would convert to advanced AMD at year 2: (1) VGG16, a popular CNN for image recognition was fine-tuned, and (2) a novel, simplified CNN architecture was trained from scratch. Preprocessing was added in the form of a segmentation-based normalization to reduce variance in the data and improve performance. Results: Our new architecture, AMDnet, with preprocessing, achieved an area under the…

Purpose: Choroidal neovascularization (CNV) is the principal pathological factor contributing to blindness in neovascular age-related macular degeneration (nAMD). Infiltration of M2 macrophage is thought to contribute to CNV progress, although the way that regulates its differentiation remains unclear. Here, we investigate the role of CHI3L1 in M2 differentiation and angiogenesis in CNV. Methods: Serums from nAMD patients were tested for CHI3L1 expression. Mice were subjected to laser injury to induce CNV, and lesion expansion were tracked using fundus fluorescence angiography (FFA) and immunofluorescence analysis. Several strategies were taken to verify the contribution of M2 macrophage and CHI3L1: macrophage depletion by clodrosome, local CHI3L1 inhibition using intravitreally injection neutralize antibody (mAY), and depletion of CHI3L1 receptor (IL13-Ra2) by small-interfering RNA (siRNA). Tuber analysis was used to further determine angiogenetic effect of CHI3L1. Anti-VEGFA was used as positive control for mAY. Results: Serum levels of CHI3L1 were highly elevated in nAMD…

Purpose: The goal of this study was to perform an extensive temporal characterization of the early pathologic processes in the streptozotocin (STZ)-induced diabetic retinopathy (DR) mouse model, beyond the vascular phenotype, and to investigate the potential of clinically relevant compounds in attenuating these processes. Methods: Visual acuity and contrast sensitivity (CS) were studied in the mouse STZ model until 24 weeks postdiabetes onset. ERG, spectral domain optical coherence tomography (SD-OCT), leukostasis, and immunohistochemistry were applied to investigate neurodegeneration, inflammation, and gliosis during early-, mid- and late-phase diabetes. Aflibercept or triamcinolone acetonide (TAAC) was administered to investigate their efficacy on the aforementioned processes. Results: Visual acuity and CS loss started at 4 and 18 weeks postdiabetes onset, respectively, and progressively declined over time. ERG amplitudes were diminished and OP latencies increased after 6 weeks, whereas SD-OCT revealed retinal thinning from 4 weeks postdiabetes. Immunohistochemical analyses linked…

Purpose: To investigate the composition and diversity of bacterial community on the ocular surface of patients with meibomian gland dysfunction (MGD) via 16S rDNA sequencing. Methods: Forty-seven patients with MGD, who were divided into groups of mild, moderate, and severe MGD, and 42 sex- and age-matched participants without MGD (control group) were enrolled. Samples were collected from the upper and lower conjunctival sac of one randomly chosen eye of each participant. Through sequencing the hypervariable region of 16S rDNA gene obtained from samples, differences in the taxonomy and diversity between groups were compared. Results: Principle coordinate analysis showed significantly distinct clustering of the conjunctival sac bacterial community between the severe MGD group and the other groups. At the phylum level, the relative abundances of Firmicutes (31.70% vs. 19.67%) and Proteobacteria (27.46% vs. 14.66%) were significantly higher (P < 0.05, Mann-Whitney U), and the abundance of Actinobacteria (34.17% vs. 56.98%) was lower in MGD than controls (P < 0.05, Mann-Whitney U). At…

Purpose: To determine age-related alterations in the retinal tissue perfusion (RTP) and volumetric vessel density (VVD) in healthy subjects. Methods: Total 148 healthy subjects (age 18 to 83 years) were enrolled and divided into four groups (G1, <35 years; G2, 35 ∼ 49 years; G3, 50 ∼ 64 years; and G4, ≥65 years). The RTP and VVD were measured at the macula. The RTP was calculated as the blood flow supplying the macular area (ϕ 2.5 mm) divided by the perfused tissue volume of the inner retina from the inner limiting membrane to the outer plexiform layer. The VVD of the macula (ϕ 2.5 mm) was calculated as the vessel density divided by the corresponding tissue volume. Results: The RTP and VVD of the retinal vascular network and deep vascular plexus (DVP) reached a peak in G2. Compared to G2, G4 had significantly lower RTP and VVD of DVP (P < 0.05). After 35 years old, age was negatively related to the RTP (r = -0.26, P = 0.02) and VVD of the DVP (r = -0.47, P < 0.001). However, age was positively related to VVD of the superficial vascular plexus (SVP; r = 0.24, P = 0.04) in subjects…

Purpose: To determine plasma metabolite and metabolic pathway differences between patients with type 2 diabetes with diabetic retinopathy (DR) and without retinopathy (diabetic controls), and between patients with proliferative DR (PDR) and nonproliferative DR (NPDR). Methods: Using high-resolution mass spectrometry with liquid chromatography, untargeted metabolomics was performed on plasma samples from 83 DR patients and 90 diabetic controls. Discriminatory metabolic features were identified through partial least squares discriminant analysis, and linear regression was used to adjust for age, sex, diabetes duration, and hemoglobin A1c. Pathway analysis was performed using Mummichog 2.0. Results: In the adjusted analysis, 126 metabolic features differed significantly between DR patients and diabetic controls. Pathway analysis revealed alterations in the metabolism of amino acids, leukotrienes, niacin, pyrimidine, and purine. Arginine, citrulline, glutamic γ-semialdehyde, and dehydroxycarnitine were key contributors to these pathway differences. A total of 151 features distinguished…

Purpose: To provide a histopathologic, morphometric analysis of the retina in Alzheimer's disease (AD). Methods: Human postmortem retinas from eight patients with AD (mean age: 80 ± 12.7 years) and from 11 age-matched controls (mean age: 78 ± 16.57 years) were analyzed. The retinas were sampled from the superior quadrant on both the temporal and nasal sides with respect to the optic nerve. Thickness of the inner and outer layers involving the retinal nerve fiber layer (RNFL), retinal ganglion cell layer (RGCL), inner plexiform layer (IPL), inner nuclear layer (INL), and outer nuclear layer (ONL) were measured and compared between controls and AD. A total of 16 measurements of retinal thickness were acquired for each layer. Results: RNFL thinning supero-temporally was significant closest to the optic nerve (∼35% thickness reduction; P < 0.001). Supero-nasally, RNFL was thinner throughout all points (∼40% reduction; P < 0.001). Supero-temporally, RGCL thinning was pronounced toward the macula (∼35% thickness reduction; P < 0.001). Supero-nasally, RGCL showed uniform thinning…

Purpose: To enumerate the ocular surface fungal microbiome of healthy human eyes by using next-generation sequencing (NGS). Methods: Tarsal and fornix conjunctiva from the lower and upper lids of both eyes of healthy individuals were swabbed in duplicate separately. A total of 34 samples were collected from both the eyes of 17 individuals, which were used for the generation of ocular surface fungal microbiomes by NGS. Twenty-four swabs were used for the detection of culturable fungi by the conventional cultivable method. Microbiome generation involved DNA extraction, internal transcribed spacer 2 (ITS2) amplification, library preparation, amplicon sequencing, taxonomic assignment of sequences, diversity analyses, and identification of genera. Results: The cultivable method detected fungi in 3 out of 24 (12.5 %) ocular surface swabs, whereas NGS identified fungi in 25 of the 34 (73.5 %) swabs. In the cultivable method Aspergillus was the only genus detected, whereas NGS detected 65 distinct genera with 12 to 24 genera per microbiome. Genera Aspergillus, Setosphaeria, Malassezia, and…

Purpose: To assess axial elongation-associated characteristics in Bruch's membrane opening (BMO) as the inner optic nerve head lamella. Methods: Participants of the population-based Beijing Eye Study without glaucoma underwent optical coherence tomography for measurement of the BMO size and shape. Results: The study included 365 individuals (mean age, 61.0 ± 8.7 years; range, 50-88 years; axial length, 24.45 ± 1.99 mm; range, 21.32-30.88 mm). Larger horizontal (mean:1.62 ± 0.28 mm) and vertical (mean: 1.74 ± 0.27 mm) BMO diameters were linearly associated with longer axial length beyond an axial length of 26.0 mm (horizontal diameter: P < 0.001; standardized regression coefficient β: 0.66; nonstandardized regression coefficient B: 0.22; 95% confidence interval (CI): 0.16, 0.27; vertical diameter: P < 0.001; β: 40; B: 0.12; 95% CI: 0.06, 0.18). In multivariable analysis, wider largest gamma zone was associated with larger intrapapillary Bruch's membrane (BM) overhanging on the side opposite to the largest gamma zone (P = 0.006; β: 0.14; B: 0.35; 95% CI: 0.10, 0.60) and with longer…

Purpose: We determine the changes in intestinal microbiota and/or disruptions in intestinal homeostasis during uveitis. Methods: Experimental autoimmune uveitis (EAU) was induced in B10.RIII mice with coadministration of interphotoreceptor retinoid-binding protein peptide (IRBP) and killed mycobacterial antigen (MTB) as an adjuvant. Using 16S rRNA gene sequencing, we looked at intestinal microbial differences during the course of uveitis, as well as intestinal morphologic changes, changes in intestinal permeability by FITC-dextran leakage, antimicrobial peptide expression in the gastrointstinal tract, and T lymphocyte prevalence before and at peak intraocular inflammation. Results: We demonstrate that increased intestinal permeability and antimicrobial peptide expression in the intestinal tract coincide in timing with increased effector T cells in the mesenteric lymph nodes, during the early stages of uveitis, before peak inflammation. Morphologic changes in the intestine were most prominent during this phase, but also occurred with adjuvant MTB alone, whereas increased intestinal…

Purpose: To systematically analyze thicknesses of retinal layers in an older population and their link to early age-related macular degeneration (AMD). Methods: In the AugUR baseline survey from a population aged ≥70 years, we conducted multimodal retinal imaging, including spectral-domain optical coherence tomography. Autosegmentation of eight distinct retinal layers was followed by manual correction of segmentation errors. AMD status was graded on color fundus images according to the Three Continent AMD Consortium Severity Scale. We tested the association of early AMD on retinal layer thicknesses by using linear mixed models and replicated significant results in independent data also from the AugUR platform. Results: When comparing layer thicknesses between early AMD and no AMD (822 eyes, 449 participants), the retinal pigment epithelium/Bruch's membrane complex demonstrated a statistically significant thickening (e.g., P = 6.41 × 10-92 for severe early versus no AMD) and photoreceptor layers showed a significant thinning. Autosegmented retinal layer thicknesses revealed similar…

Purpose: Glaucoma, a leading cause of blindness worldwide, often remains undetected until irreversible vision loss has occurred. Treatments focus on lowering intraocular pressure (IOP), the only modifiable and readily measurable risk factor. However, IOP can vary and does not always predict disease progression. MicroRNAs (miRNAs) are promising biomarkers. They are abundant and stable in biological fluids, including plasma and aqueous humor (AqH). We aimed to identify differentially expressed miRNAs in AqH and plasma from glaucoma, exfoliation syndrome (XFS), and control subjects. Methods: Plasma and AqH from two ethnic cohorts were harvested from glaucoma or XFS (often associated with glaucoma, n = 33) and control (n = 31) patients undergoing elective surgery. A custom miRNA array measured 372 miRNAs. Molecular target prediction and pathway analysis were performed with Ingenuity Pathway Analysis (IPA) and DIANA bioinformatical tools. Results: Levels of miRNAs in plasma, a readily accessible biomarker source, correlated with miRNA levels in AqH. Twenty circulating miRNAs were at…

Purpose: To characterize intraocular pressure (IOP) dynamics by identifying the sources of transient IOP fluctuations and quantifying the frequency, magnitude, associated cumulative IOP-related mechanical energy, and temporal distribution. Methods: IOP was monitored at 500 Hz for periods of 16 to 451 days in nine normal eyes of six conscious, unrestrained nonhuman primates using a validated, fully implanted wireless telemetry system. IOP transducers were calibrated every two weeks via anterior chamber cannulation manometry. Analysis of time-synchronized, high-definition video was used to identify the sources of transient IOP fluctuations. Results: The distribution of IOP in individual eyes is broad, and changes at multiple timescales, from second-to-second to day-to-day. Transient IOP fluctuations arise from blinks, saccades, and ocular pulse amplitude and were as high as 14 mm Hg (>100%) above momentary baseline. Transient IOP fluctuations occur ∼10,000 times per waking hour, with ∼2000 to 5000 fluctuations per hour greater than 5 mm Hg (∼40%) above baseline. Transient IOP…

Purpose: The purpose of this study was to study the status and association among tear-soluble factors, corneal dendritic cell density, vitamin D, and signs and symptoms in dry eye disease (DED). Methods: A total of 33 control subjects and 47 evaporative dry eye patients were included in the study. DED diagnosis and classification was based on the 2017 Report of the Tear Film & Ocular Surface Society International Dry Eye Workshop (TFOS DEWS II). DED workup, including tear film break-up time (TBUT), Schirmer's test I (STI), corneal and conjunctival staining, ocular surface disease index (OSDI) scoring, and in vivo confocal microscopy (to assess corneal dendritic cell density [cDCD] and subbasal nerve plexus [SBNP] features) was performed in the study subjects. Tear fluid using Schirmer's strip and serum were collected from the subjects. Multiplex ELISA or single analyte ELISA was performed to measure 34 tear-soluble factors levels including vitamin D. Results: Significantly higher OSDI discomfort score, lower TBUT, and lower STI were observed in DED patients. cDCD was significantly…

Purpose: Lipofuscin and melanolipofuscin organelles in retinal pigment epithelium (RPE) cells are signal sources for clinical fundus autofluorescence (AF). To elucidate the subcellular basis of AF imaging, we identified, characterized, and quantified the frequency of RPE morphology and AF phenotypes in donor eyes with age-related macular degeneration (AMD). Methods: In 25 RPE-Bruch's membrane flat mounts from 25 eyes, we analyzed 0.4-μm z-stack epifluorescence images of RPE stained with phalloidin for actin cytoskeleton. Using a custom ImageJ plugin, we classified cells selected in a systematic unbiased fashion in six phenotypes representing increasing degrees of pathology. For each cell, area, AF intensity, and number of Voronoi neighbors were compared with phenotype 1 (uniform AF, polygonal morphology) via generalized estimating equations. We also analyzed each cell's neighborhood. Results: In 29,323 cells, compared with phenotype 1, all other phenotypes, in order of increasing pathology, had significantly larger area, reduced AF, and more variable number of neighbors.…

Human age-related nuclear cataract is commonly characterized by four biochemical features that involve modifications to the structural proteins that constitute the bulk of the lens: coloration, oxidation, insolubility, and covalent cross-linking. Each of these is progressive and increases as the cataract worsens. Significant progress has been made in understanding the origin of the factors that underpin the loss of lens transparency. Of these four hallmarks of cataract, it is protein-protein cross-linking that has been the most intransigent, and it is only recently, with the advent of proteomic methodology, that mechanisms are being elucidated. A diverse range of cross-linking processes involving several amino acids have been uncovered. Although other hypotheses for the etiology of cataract have been advanced, it is likely that spontaneous decomposition of the structural proteins of the lens, which do not turn over, is responsible for the age-related changes to the properties of the lens and, ultimately, for cataract. Cataract may represent the first and best characterized of a…

Purpose: To develop deep learning (DL) models for the automatic detection of optical coherence tomography (OCT) measures of diabetic macular thickening (MT) from color fundus photographs (CFPs). Methods: Retrospective analysis on 17,997 CFPs and their associated OCT measurements from the phase 3 RIDE/RISE diabetic macular edema (DME) studies. DL with transfer-learning cascade was applied on CFPs to predict time-domain OCT (TD-OCT)-equivalent measures of MT, including central subfield thickness (CST) and central foveal thickness (CFT). MT was defined by using two OCT cutoff points: 250 μm and 400 μm. A DL regression model was developed to directly quantify the actual CFT and CST from CFPs. Results: The best DL model was able to predict CST ≥ 250 μm and CFT ≥ 250 μm with an area under the curve (AUC) of 0.97 (95% confidence interval [CI], 0.89-1.00) and 0.91 (95% CI, 0.76-0.99), respectively. To predict CST ≥ 400 μm and CFT ≥ 400 μm, the best DL model had an AUC of 0.94 (95% CI, 0.82-1.00) and 0.96 (95% CI, 0.88-1.00), respectively. The best deep convolutional neural network…

Purpose: Geographic atrophy (GA) is a clinical phenotype of late age-related macular degeneration (AMD) with no current treatment available. In this study, we investigated markers of chronic inflammation in plasma of patients with GA and how these relate to progression rate. Methods: We prospectively included 42 patients with GA, 41 patients with neovascular AMD, and 27 healthy controls. We quantified levels of interleukin (IL)-1β, IL-6, IL-8, tumor necrosis factor (TNF) receptor 2, and C-reactive protein (CRP). We adapted an inflammation summary score to cluster conceptually related markers of chronic inflammation. Enlargement rate of the atrophic lesion was measured from fundus autofluorescence images performed at baseline and after 1 year. Results: Patients with GA showed an increase in proinflammatory markers of IL-6 (P = 0.009), TNF receptor 2 (P = 0.013), and CRP (P = 0.017) compared to healthy controls. We found that IL-8 levels were markedly higher in patients with GA when compared to patients with neovascular AMD (P = 0.013). The inflammation summary score was high in…

Purpose: To perform deep phenotyping of subjects with PDE6C achromatopsia and examine disease natural history. Methods: Eight subjects with disease-causing variants in PDE6C were assessed in detail, including clinical phenotype, best-corrected visual acuity, fundus autofluorescence, and optical coherence tomography. Six subjects also had confocal and nonconfocal adaptive optics scanning light ophthalmoscopy, axial length, international standard pattern and full-field electroretinography (ERG), short-wavelength flash (S-cone) ERGs, and color vision testing. Results: All subjects presented with early-onset nystagmus, decreased best-corrected visual acuity, light sensitivity, and severe color vision loss, and five of them had high myopia. We identified three novel disease-causing variants and provide phenotype data associated with nine variants for the first time. No subjects had foveal hypoplasia or residual ellipsoid zone (EZ) at the foveal center; one had an absent EZ, three had a hyporeflective zone, and four had outer retinal atrophy. The mean width of the central EZ lesion on…

Purpose: Deregulated expression of several microRNAs (miRNAs) in sera or salivary glands of patients with Sjögren syndrome (SS) has been reported. However, none have investigated miRNAs in samples that can represent lacrimal glands. We compared the miRNAs expression in the tears of SS patients and healthy controls. Moreover, we investigated the correlation between miRNAs expression and ocular staining score (OSS). Methods: Individual tear samples were collected from 18 SS patients and 8 age-matched controls. Clinical ophthalmologic assessments included Schirmer I test, tear film breakup time (tBUT), and OSS. The expression of 43 different miRNAs in tears was measured using real-time polymerase chain reaction, and compared between the SS patients and controls. And we also compared between the three groups of control, primary SS, and secondary SS patients. The correlation between the miRNA expression and OSS was evaluated. Results: The expression levels of miR-16-5p, miR-34a-5p, miR-142-3p, and miR-223-3p were significantly upregulated in patients with SS when compared with those in…

Purpose: To evaluate peripapillary and macular vessel density in eyes with dysthyroid optic neuropathy (DON) and its correlation with visual function. Methods: Patients diagnosed as thyroid-associated ophthalmopathy (TAO) with or without DON and healthy participants were recruited. All subjects underwent a complete ophthalmic examination and optical coherence tomography angiography centered on the fovea and the optic nerve head. Microvascular measurements were summarized as vessel density in the whole image and in each subfield. Visual function, including best-corrected visual acuity (BCVA), visual field (VF), and visual evoked potential (VEP), were assessed for all TAO patients. Areas under the receiver operating characteristic curves (AUROCs) were applied to evaluate the diagnostic accuracy of vessel density for DON. Results: A total of 23 healthy eyes, 41 TAO eyes without DON, and 30 with DON were studied. The radial peripapillary capillary whole image vessel density (rpc-wiVD) and optic nerve head whole image vessel density (onh-wiVD) were significantly decreased in DON eyes…

Purpose: To investigate the molecular pathways that drive thyroid stimulating hormone receptor (TSHR)-induced cellular proliferation in orbital fibroblasts (OFs) from thyroid eye disease (TED) patients. Methods: Orbital fibroblasts from TED and non-TED patients were treated with TSH and changes in gene expression and proliferation were measured. To determine the role of TSHR, TSHR-specific siRNA was used to deplete TSHR levels. Proliferation was measured by bromodeoxyuridine (BrdU) incorporation. PI3K/Akt activation was analyzed by Western blot. The PI3K inhibitor LY294002 was used to investigate PI3K/Akt signaling in OF proliferation. Expression of TSHR, inflammatory cytokines, proliferation related genes and miR-146a and miR-155 were measured by qPCR. Results: Orbital fibroblasts from TED patients proliferate significantly more than non-TED OFs in response to TSH. TSH-induced proliferation was dependent upon TSHR expression and required the PI3K/Akt signaling cascade. TSHR activation stimulated miR-146a and miR-155 expression. TED OFs produced significantly more miR-146a and…

Purpose: To evaluate the effect of age on global and sectoral vascular parameters of the peripapillary area and macula in healthy eyes by using optical coherence tomography angiography (OCT-A). Methods: This retrospective cross-sectional study included 239 eyes of 172 healthy subjects. Subjects were scanned using the high-definition disc angio scan (4.5 × 4.5 mm), retina angio scan (6 × 6 mm), and optic nerve head/ganglion cell complex (GCC) modes of OCT-A. Global and sectoral circumpapillary vessel density (VD), parafoveal VD, retinal nerve fiber layer (RNFL), and GCC thickness parameters were modeled in terms of age by using linear mixed-effect models incorporating covariates. Normalized slopes were calculated by dividing the absolute slopes by the mean value of the OCT-A parameters. Results: All global and sectoral circumpapillary VDs decreased significantly with increasing age, except for those in the temporal superior sector. The steepest slopes in the circumpapillary region were observed in nasal superior (-0.098%/y) and nasal inferior (-0.096%/y) sectors. The global,…

Purpose: To assess the agreement between structural (optical coherence tomography [OCT]) and functional (visual field [VF]) glaucomatous damage with an automated method and deviation/probability maps, and to compare this method to a metric method. Methods: Wide-field spectral-domain OCT scans, including the disc and macula, and 24-2 and 10-2 VFs were obtained from 45 healthy control (H) eyes/individuals, and 53 eyes/patients with 24-2 mean deviation (MD) better than -6 dB diagnosed as "definite glaucoma" (DG) by experts. Abnormal structure-abnormal function (aS-aF) agreement was assessed with an automated topographic (T) method based upon VF pattern deviation and OCT probability maps. Results were compared to a metric (M) method optimized for accuracy, (abnormal 24-2 glaucoma hemifield test [GHT] or pattern standard deviation [PSD], or 10-2 PSD AND abnormal OCT [quadrant]). Results: For the T-method, 47 (88.7%) of the 53 DG eyes showed aS-aF agreement, compared to 2 (4.5%) of the 45 H eyes. The aS-aF agreement for these two H eyes was easily identified as mistaken, and did not…

Purpose: To determine the riboflavin concentration in the posterior corneal stroma, Descemet's membrane, and endothelium prior to UV irradiation in corneal cross-linking (CXL) in humans. Methods: Five human deepithelialized cadaver corneas were mounted into artificial anterior chambers. After the establishment of stable physiological hydration, two-photon imaging with a certified multiphoton tomograph was used to determine fluorescence intensity and second harmonic generation signals from collagen throughout each cornea by optical sectioning, with a step size of 2.5 μm. Afterward, 0.1% riboflavin solution was applied to the anterior corneal surface, similar to the standard CXL protocol. To determine the absolute riboflavin concentration immediately before UV irradiation, corneas were measured by two-photon imaging just at the end of the riboflavin imbibition and after riboflavin saturation. Results: The topical application of 0.1% riboflavin results in a riboflavin concentration that decreases to 0.035% in the posterior stroma. Inside Descemet's membrane and endothelium, the…

Purpose: We quantify volumetric changes of subretinal hyperreflective material (SHRM) and determine the conversion toward subretinal fibrosis, the angiofibrotic switch, under anti-VEGF therapy using polarization-sensitive optical coherence tomography (PS-OCT). Methods: A total of 50 eyes of 50 patients with treatment-naïve neovascular age-related macular degeneration (AMD) were included in this prospective observational study: 26 diagnosed with type 1 choroidal neovascularization (CNV), seven with type 2 CNV, 11 with mixed type CNV, three with a retinal angiomatous proliferation (RAP) lesion and three with a polypoidal choroidal vasculopathy (PCV). Patients were imaged at baseline and at the end of the loading phase (after treatment with three intravitreal anti-VEGF injections) using a PS-OCT system with a scanning angle of 30° × 30° and a scan pattern of 1024 × 250 A-scans. The device is capable of detecting fibrosis based on birefringence and the RPE based on depolarization. The volume of SHRM was quantified by manual delineation in each PS-OCT B-scan and interpolation between…

Purpose: The purpose of this study is to evaluate differences in optical coherence tomography angiography (OCT-A) findings between patients with papilledema and pseudopapilledema. Methods: In this prospective, comparative study, 41 eyes of 21 subjects with papilledema, 27 eyes of 15 subjects with pseudopapilledema, and 44 eyes of 44 healthy normal subjects were included and were imaged using OCT-A. In addition to peripapillary total vasculature maps obtained with commercial vessel density mapping, major vessel removal using customized image analysis software was also used to measure whole image capillary density and peripapillary capillary density (PCD). Peripapiilary retinal nerve fiber layer (RNFL) and macular ganglion cell complex (GCC) were recorded. Results: Average RNFL thicknesses were greater in papilledema eyes than in pseudopapilledema and control subjects. GCC thickness was not different among three groups. Peripapillary vasculature values were significantly lower in papilledema (58.5 ± 6.1%) and pseudopapilledema (58.9 ± 4.7%) eyes compared with healthy eyes (63.2 ±…

Purpose: Explore in vivo whether there is direct communication between the cerebrospinal fluid (CSF) and extravascular compartment of human visual pathway structures. Methods: A prospective and observational study included 10 subjects who underwent intrathecal gadolinium-enhanced magnetic resonance imaging (MRI) for suspected CSF circulation disorder, but with a negative result and with no known ophthalmic diseases. After precontrast T1-weighted MRI, 0.5 mL of gadobutrol (Gadovist, 1.0 mmol/mL) was injected intrathecally. Gadobutrol distributes in the extravascular space, and served as a CSF tracer. Consecutive MRI scans were obtained throughout 24 to 48 hours. To assess gadobutrol contrast enrichment, regions of interest (ROIs) were placed at multiple locations along the visual pathway, from the primary visual cortex to the eye's vitreous body. CSF tracer dependent T1 signal was measured in each ROI. A linear mixed-model was used for statistical analyses. Results: CSF tracer enrichment was found within the optic nerve, optic chiasm, optic tract, and primary visual cortex (P <…

Purpose: Our study aimed to evaluate the side effects of selective serotonin reuptake inhibitors (SSRIs) on the ocular surface. Methods: Twenty patients with depression and dry eye disease (DED) were randomly picked to receive SSRI treatment, whereas another 20 patients received placebo treatment. The serotonin, inflammatory cytokine, and proapoptotic protein levels were determined by using protein chip, qRT-PCR, and ELISA analyses. A rat depression model was established, and SSRIs were applied for 3 or 6 weeks. Tear production and corneal epithelial barrier function were evaluated. The serotonin and inflammatory cytokine levels were analyzed by qRT-PCR, immunohistochemical staining, and ELISA. Human corneal epithelial cells were subjected to serotonin, a HTR antagonist, and/or an NF-κB signaling inhibitor. The inflammatory cytokine and proapoptotic protein levels were determined by qRT-PCR, Western blot analysis, and ELISA. The cell apoptosis rate was assessed by using flow cytometry. Results: The SSRI group had higher tear serotonin levels and more serious inflammation and cell…

Purpose: To investigate the contribution and mechanism of miRNAs and autophagy in diabetic peripheral neuropathy. Methods: In this study, we used streptozotocin (STZ)-induced type I diabetes C57 mice as animal models, and we detected the expression of miR-34c and autophagic intensity in trigeminal ganglion (TG) tissue. The bioinformatics software was used to predict and analyze the potential targets of miR-34c. Primary trigeminal ganglion neurons were cultured in vitro to investigate the effect of miR-34c on axon growth and survival of TG cells. A corneal epithelial damage-healing model was established on the diabetic mice, then miR-34c antagomir was injected subconjunctivally. The condition of corneal epithelial healing was observed through sodium fluorescein staining, and the peripheral nerve degeneration of the cornea was evaluated by β-tublin corneal nerve staining. Results: The expression of miR-34c was significantly increased in TG tissue of type I diabetic mice by real-time PCR. Western blot showed that autophagy-related proteins Atg4B and LC3-II were significantly…

Purpose: Mutations in six genes have been associated with achromatopsia (ACHM): CNGA3, CNGB3, PDE6H, PDE6C, GNAT2, and ATF6. ATF6 is the most recent gene to be identified, though thorough phenotyping of this genetic subtype is lacking. Here, we sought to test the hypothesis that ATF6-associated ACHM is a structurally distinct form of congenital ACHM. Methods: Seven genetically confirmed subjects from five nonconsanguineous families were recruited. Foveal hypoplasia and the integrity of the ellipsoid zone (EZ) band (a.k.a., IS/OS) were graded from optical coherence tomography (OCT) images. Images of the photoreceptor mosaic were acquired using confocal and nonconfocal split-detection adaptive optics scanning light ophthalmoscopy (AOSLO). Parafoveal cone and rod density values were calculated and compared to published normative data as well as data from two subjects harboring CNGA3 or CNGB3 mutations who were recruited for comparative purposes. Additionally, nonconfocal dark-field AOSLO images of the retinal pigment epithelium were obtained, with quantitative analysis performed in one…

Purpose: Relationships between tear film lipid (TFL) layer composition, structure, and function could provide insight into the etiology of dry eye. The molar ratio of cholesteryl ester (CE)/wax ester (WE) was measured in meibum from normal donors (Mn) and compared with meibum from donors with meibomian gland dysfunction (MMGD). Methods: CE/WE was measured using nuclear magnetic resonance spectroscopy. Results: CE/WE was distributed into two populations with 81% distributed near 0.55 and 19% near 0.3. CE/WE were higher in donors 13 to 19 years old compared with donors 1 to 12 years old and 20 to 88 years old. CE/WE for MMGD was 30% lower, 0.34 ± 0.04, compared with Mn, 0.49 ± 0.04. There were no sex differences in CE/WE. There were no significant racial differences between the CE/WE ratios for Asians and Caucasians. The CE/WE ratio was higher for blacks and lower for Hispanics compared to Caucasians. Due to the small number sampled, confirmation of the later racial results is needed. The packing of CE and WE in the TFL layer was proposed. Conclusions: Although MMGD contains much less…

Purpose: To quantify the thickness of the outer retinal sublayers and choroid in pathological myopia and examine associations between these factors and best-corrected visual acuity (BCVA). Methods: The cohort was composed of 21 eyes with emmetropia and 70 eyes with high myopia (49 simple high myopia; 21 pathological myopia). Optical coherence tomography images were segmented to determine macular thicknesses of the choroid and the following outer retinal sublayers: outer plexiform layer (OPL), Henle fiber layer and outer nuclear layer (HFL + ONL), myoid and ellipsoid zone (MEZ), outer segment of photoreceptors (OS), and interdigitation zone and RPE/Bruch complex (IZ + RPE). Correlations between BCVA and thickness of the outer retinal sublayers and choroid were determined. Results: In pathological myopia, the choroid, HFL + ONL, MEZ, and IZ + RPE were thinner than in emmetropia and simple high myopia (P < 0.05). Simple and multiple regression models showed that MEZ thickness was correlated with BCVA (both P < 0.001). The relationship between MEZ thickness and BCVA varied with…

Purpose: Aging is a risk factor for dry eye disease. The aim of this study was to investigate if aging is associated with a range of signs of dry eye disease, including tear hyperosmolarity, reduced nerve density, and increased dendritic cell number, in mice. Method: Healthy C57BL/6 female mice, aged 2 months (young, n = 10) and 22 months (aged, n = 11) were used. Clinical assessments included corneal sensitivity (Cochet-Bonnet esthesiometry), tear secretion (Phenol red thread test), tear film osmolarity (TearLab osmometer), and corneal thickness (optical coherence tomography). The sum length of the corneal superficial terminals and sub-basal nerves, density of vertical nerve projections, and density and tree area of resident epithelial dendritic cells, were assessed using immunofluorescence and confocal microscopy. Results: Aged mice had significantly higher tear secretion, lower corneal sensitivity, and a thicker corneal stroma but thinner epithelium. There was no significant intergroup difference for tear osmolarity. Aged mice showed a significantly lower sum length of nerves in…

Purpose: To investigate retinal structure in subjects with CNGA3-associated achromatopsia and evaluate disease symmetry and intrafamilial variability. Methods: Thirty-eight molecularly confirmed subjects underwent ocular examination, optical coherence tomography (OCT), and nonconfocal split-detection adaptive optics scanning light ophthalmoscopy (AOSLO). OCT scans were used for evaluating foveal hypoplasia, grading foveal ellipsoid zone (EZ) disruption, and measuring outer nuclear layer (ONL) thickness. AOSLO images were used to quantify peak foveal cone density, intercell distance (ICD), and the coefficient of variation (CV) of ICD. Results: Mean (±SD) age was 25.9 (±13.1) years. Mean (± SD) best corrected visual acuity (BCVA) was 0.87 (±0.14) logarithm of the minimum angle of resolution. Examination with OCT showed variable disruption or loss of the EZ. Seven subjects were evaluated for disease symmetry, with peak foveal cone density, ICD, CV, ONL thickness, and BCVA not differing significantly between eyes. A cross-sectional evaluation of AOSLO imaging showed a mean (±SD) peak…

Purpose: The purpose of this study was to investigate the cytokine profiles in plasma and aqueous humor of patients with choroidal neovascularization (CNV) due to exudative AMD and polypoidal choroidal vasculopathy (PCV). Methods: In this cross-sectional study, 16 patients clinically diagnosed with AMD, 18 patients with PCV, and 50 age- and sex-matched cataract patients without AMD/PCV (controls) were enrolled. Study subjects were treatment naïve, and 200 μL undiluted aqueous humor and 5 mL peripheral venous blood were collected from the study subjects. Clinical samples were analyzed for 41 different cytokines by Luminex bead-based multiplex assay. Cytokines concentrations with detection rates of 50% or more were included for the analysis, and the differences in plasma and aqueous humor cytokines levels between each group were analyzed. Results: The age of the patients with AMD and PCV was 70.62 ± 10.15 (mean ± SD) and 71.48 ± 9.08 years, respectively, and that in the control group was 62.8 ± 10.67 years. Aqueous humor cytokines growth-regulated oncogene (GRO), macrophage-derived…

Purpose: To obtain a choroidal vascularity index (CVI) map of macular area on an Early Treatment Diabetic Retinopathy Study (ETDRS) grid. Methods: The study was a cross-sectional study involving 30 eyes of 30 healthy individuals. In brief, a shadow-compensated automated algorithm was used to segment and binarize the individual optical coherence tomography (OCT) B-scans. This was followed by three-dimensional reconstruction of these processed B-scans to obtain the overall thickness and vascularity maps. ETDRS grid was overlaid on both the extrapolated thickness and vascularity maps to obtain the corresponding sector-wise CVI. The main outcome measure was to evaluate the topographical variation of CVI in the macular area. Results: The mean age of the study participants was 44.33 ± 16.29 years (range, 18-70 years). CVI showed no significant difference in different rings, subfields, or quadrants of the ETDRS map. CVI had a negative correlation with age (r = -0.384, P = 0.03). There were no statistically significant differences between CVI of both eyes in either rings or the full ETDRS…

Purpose: The objective of this study was to map the distribution and density of the three major components of the classical scotopic "night vision" pathway (rods, rod bipolar, and AII amacrine cells) in postmortem human retinas. Methods: Four postmortem donor eyes (male and female, aged 44-56 years) were used to cut vertical sections through the temporal horizontal meridian. The sections were processed for immunohistochemistry and imaged using high-resolution multichannel confocal microscopy. Rods, rod bipolar, and AII amacrine cells were counted along the temporal horizontal meridian. Two additional retinas were used for intracellular injections. Results: Rod peak density is close to 150,000 cells/mm2 at 4 to 5 mm (15° to 20°) eccentricity, declining to below 70,000 cells/mm2 in peripheral retina. Rod bipolar density is lower but follows a similar distribution with peak density near 10,000 cells/mm2 between 2 and 4 mm (7° to 15°) eccentricity declining to below 4000 cells/mm2 in peripheral retina. The peak density of AII amacrine cells (near 4000 cells/mm2) is located close to the…

Purpose: To investigate ocular and systemic diurnal rhythms in emmetropic and myopic adults and examine relationships with light exposure. Methods: Adult subjects (n = 42, 22-41 years) underwent measurements every 4 hours for 24 hours, including blood pressure, heart rate, body temperature, intraocular pressure (IOP), ocular biometry, and optical coherence tomography imaging. Mean ocular perfusion pressure (MOPP) was calculated. Saliva was collected for melatonin and cortisol analysis. Acrophase and amplitude for each parameter were compared between refractive error groups. Subjects wore a light, sleep, and activity monitor for 1 week before measurements. Results: All parameters exhibited significant diurnal rhythm (ANOVA, P < 0.05 for all). Choroidal thickness peaked at 2.42 hours, with a diurnal variation of 25.8 ± 13.44 μm. Axial length peaked at 12.96 hours, with a variation of 35.71 ± 6.6 μm. Melatonin peaked at 3.19 hours during the dark period, while cortisol peaked after light onset at 8.86 hours. IOP peaked at 11.24 hours, with a variation of 4.92 ± 1.57 mm Hg, in antiphase…

Purpose: To assess how DNA damage-inducible transcript 4 (DDIT4) and autophagic flux are altered in dry eye disease and reveal the underlying mechanisms. Methods: C57BL/6 mice were exposed to desiccating stress (subcutaneous scopolamine [0.5 mg/0.2 mL] 3 times a day, humidity < 30%) for 7 days. Primary human corneal epithelial cells and cells from a human corneal epithelial cell line were cultured under hyperosmolarity. Western blot assays and immunofluorescence staining were used to measure changes in protein expression. mRNA expression was analyzed by RT-PCR and quantitative real-time PCR. Autophagosomes were observed through electron microscopy. Cellular reactive oxygen species and mitochondrial function were detected with 2',7'-dichlorodihydrofluorescein diacetate and mitochondrial membrane potential assays. Cell Counting Kit-8 and lactate dehydrogenase assays were used to measure cell death. Apoptosis was analyzed by Annexin V-PI flow cytometry. Results: Increased expression of microtubule-associated protein 1 light chain 3 (LC3-II), sequestosome 1 (SQSTM1), and DDIT4 were…

Purpose: Recent evidence suggests that retinal photoreceptor cells have an important role in the pathogenesis of retinal microvascular lesions in diabetes. We investigated the role of rod cell phototransduction on the pathogenesis of early diabetic retinopathy (DR) using Gnat1-/- mice (which causes permanent inhibition of phototransduction in rod cells without degeneration). Methods: Retinal thickness, oxidative stress, expression of inflammatory proteins, electroretinograms (ERG) and optokinetic responses, and capillary permeability and degeneration were evaluated at up to 8 months of diabetes. Results: The diabetes-induced degeneration of retinal capillaries was significantly inhibited in the Gnat1-/- diabetics. The effect of the Gnat1 deletion on the diabetes-induced increase in permeability showed a nonuniform accumulation of albumin in the neural retina; the defect was inhibited in diabetic Gnat1-/- mice in the inner plexiform layer (IPL), but neither in the outer plexiform (OPL) nor inner nuclear (INL) layers. In Gnat1-deficient animals, the diabetes-induced increase in…

Purpose: To investigate the presence and role of fibroblast senescence in the dynamic process of corneal wound healing involving stromal cell apoptosis, proliferation, and differentiation. Methods: An in vivo corneal wound healing model was performed using epithelial debridement in C57BL/6 mice. The corneas were stained using TUNEL, Ki67, and α-smooth muscle actin (α-SMA) as markers of apoptosis, proliferation, and myofibroblastic differentiation, respectively. Cellular senescence was confirmed by senescence-associated β-galactosidase (SA-β-gal) staining and P16Ink4a expression. Mitogenic response and gene expression were compared among normal fibroblasts, H2O2-induced senescent fibroblasts, and TGF-β-induced myofibroblasts in vitro. The senescence was further detected in mouse models of corneal scarring, alkali burn, and penetrating keratoplasty (PKP). Results: The apoptosis and proliferation of corneal stromal cells were found to peak at 4 and 24 hours after epithelial debridement. Positive staining of SA-β-gal was observed clearly in the anterior stromal cells at 3 to 5 days. The…

Purpose: To examine trends in the prevalence of myopia and myopic maculopathy in a general Japanese population. Methods: Residents of a Japanese community aged 40 years and older participated in surveys conducted in 2005, 2012, and 2017. Each participant underwent comprehensive eye examinations that included measurements of refractive error, axial lengths, and color fundus photography. Myopic maculopathy was defined according to the criteria of the Meta-analysis of Pathologic Myopia Study Group classification system. Trends in the prevalence of myopia and myopic maculopathy were tested by using a logistic regression analysis fitted by generalized estimating equations to account for individuals submitting to repeated examination. Results: The age-adjusted frequencies of myopia increased significantly from 2005 to 2017 (myopia, 37.7%-45.8%; high myopia 5.8%-9.5%; all P for trend <0.001). The age-adjusted frequency of an axial length level of 26.5 mm or more increased significantly from 2005 to 2017 (3.6%-6.0%; P for trend <0.001). The age-adjusted prevalence of myopic maculopathy also…

Purpose: Besides glia-driven neuroinflammation, growing evidence from analysis of human blood samples, isolated autoantibodies, and postmortem tissues also support systemic immune responses during neurodegeneration in glaucoma patients. To explore the T-cell-mediated component of systemic immunity, this study analyzed T lymphocytes in patients' blood. Methods: Blood samples were collected from 32 patients with glaucoma and 21 nonglaucomatous controls, and mononuclear cells were isolated by Histopaque density gradient centrifugation. T-cell subset distribution was analyzed by multicolor flow cytometry after helper (Th) and cytotoxic fractions, and Th subpopulations, were stained with antibodies to CD4, CD8, or distinctive markers, such as IFN-γ (for Th1), IL-4 (for Th2), IL-17A (for Th17), and CD25/FoxP3 (for T regulatory cells [Tregs]). In addition, proliferative activity and cytokine secretion of T cells were analyzed after in vitro stimulation. Results: Analysis of T-cell subset distribution detected a glaucoma-related shift. Despite similar frequencies of CD4+ or CD8+ T cells, or…

Purpose: Light-emitting diodes that emit high-intensity blue light are associated with blue-light hazard. Here, we report that blue light disturbs circadian rhythms by interfering with the clock gene in the suprachiasmatic nucleus (SCN) and that suppression of blue light at night ameliorates metabolic abnormalities by controlling circadian rhythms. Methods: C57BL/6J mice were exposed to 10-lux light for 30 minutes at Zeitgeber time 14 for light pulse with blue light or blue-light cut light to induce phase shift of circadian rhythms. Phase shift, clock gene expression in SCN, and metabolic parameters were analyzed. In the clinical study, healthy participants wore blue-light shield eyewear for 2 to 3 hours before bed. Anthropometric data analyses, laboratory tests, and sleep quality questionnaires were performed before and after the study. Results: In mice, phase shift induced with a blue-light cut light pulse was significantly shorter than that induced with a white light pulse. The phase of Per2 expression in the SCN was also delayed after a white light pulse. Moreover, blood glucose…

Purpose: Corneal endothelial dysfunction leads to corneal edema, pain, and vision loss. Adequate animal models are needed to study the safety and efficacy of novel cell therapies as an alternative to corneal transplantation. Methods: Primary human corneal endothelial cells (HCECs) were isolated from cadaveric donor corneas, expanded in vitro, transduced to express green fluorescent protein (GFP), loaded with superparamagnetic nanoparticles, and injected into the anterior chamber of adult rabbits immediately after endothelial cell or Descemet's membrane stripping. The same volume of balanced salt solution plus (BSS+) was injected in control eyes. We compared different models for inducing corneal edema in rabbits, and examined the ability of transplanted HCECs to reduce corneal edema over time by measuring central corneal thickness and tracking corneal clarity. GFP-positive donor cells were tracked in vivo using optical coherence tomography (OCT) fluorescence angiography module, and the transplanted cells were confirmed by human nuclei immunostaining. Results: Magnetic HCECs…

Purpose: To assess retinal microvascular reactivity during dark adaptation and the transition to ambient light and after flicker stimulation using optical coherence tomography angiography (OCTA). Methods: Fifteen eyes of 15 healthy participants were dark adapted for 45 minutes followed by OCTA imaging in the dark-adapted state. After 5 minutes of normal lighting, subjects underwent OCTA imaging. Participants were then subjected to a flashing light-emitting diode (LED) light and repeat OCTA. Parafoveal vessel density and adjusted flow index (AFI) were calculated for superficial (SCP), middle (MCP), and deep capillary plexuses (DCP), and then compared between conditions after adjusting for age, refractive error, and scan quality. SCP vessel length density (VLD) was also evaluated. Between-condition capillary images were aligned and subtracted to identify differences. We then analyzed images from 10 healthy subjects during the transition from dark adaptation to ambient light. Results: SCP vessel density was significantly higher while SCP VLD was significantly lower during ambient light…

Purpose: To investigate whole transcriptional differences between proliferative diabetic retinopathy (PDR) neovascular membranes (NVMs) and retinas, and the regulatory genes participating in retinal neovascularization in PDR. Methods: We used high-throughput sequencing technology to capture the whole-genome gene expression levels of all participants, including 23 patients with PDR or branch retinal vein occlusion (BRVO), 3 normal retinal samples, and 2 retinal samples from type II diabetic (T2D) eyes by donation, followed by analyses of expression patterns using bioinformatics methods, then validation of the data by in situ hybridization and Western blotting. Results: We showed that transcriptional profiles of the NVMs were distinct from those of the retinas. Angiogenesis growth factors VEGFC, ANGPT1, ANGPT2, and EFNB2, and their receptors FLT4, TIE1, TIE2, and EPHB4, respectively, were overexpressed. Expression of VEGFA was highly upregulated in T2D retina, but low in the NVMs, while angiogenesis transcription factors, including ETS1 and ERG, were coordinately upregulated in NVMs.…

Purpose: To assess the ocular surface in volunteers who consider themselves as healthy, in order to evaluate how para-inflammatory mechanisms fail with age, and thus investigate the phenomenon of "InflammAging." Methods: In this observational prospective cohort study, volunteers were categorized into three groups according to age: young (19-40 years), middle-aged (41-60 years), and older adults (61-93 years). Clinical assessments included tear breakup time (T-BUT) and Schirmer test type I. Dry eye symptoms were evaluated by the Ocular Surface Disease Index (OSDI) questionnaire. Conjunctival mRNA and protein expression of intercellular adhesion molecule-1 (ICAM-1), MUC5AC, and IL-8 were measured by real-time PCR and immunofluorescence. Results: A total of 82 volunteers (38 males and 44 females) were enrolled. T-BUT decreased significantly with increasing age (young: 11.13 ± 0.18 seconds; middle-aged: 10.83 ± 0.56 seconds; older: 9.00 ± 1.00 seconds, P < 0.05). Schirmer test values decreased significantly with age (young: 20.6 ± 1.0 mm; middle-aged: 19.2 ± 1.2 mm; older: 16.0 ± 1.1…

Purpose: We investigate the impact of parental myopia on spherical equivalent (SE) progression and axial length (AL) elongation. Methods: Children and their parents were invited for annual examinations from 2006 (baseline). Cycloplegic autorefraction and AL were measured at each visit. Parental refractive status was determined using refraction data from their baseline visit. Children were classified into five groups: no myopic parents (non-non), only one moderately myopic parent (non-moderate), only one highly myopic parent (non-high), two moderately myopic parents (moderate-moderate), and one moderately myopic or more severe and one highly myopic parent (moderate-high/high-high). The relationship between progression of SE and AL with parental refractive status was estimated by linear mixed-effects models. Data from 2006 to 2017 were analyzed in the current study. Results: A total of 1831 children were enrolled (mean age, 11 ± 2.7 years; mean standard error, -0.49 ± 2.16 diopters [D] at baseline. Myopia progressed faster for children with parental myopia (non-non group as reference,…

Purpose: To study the characteristics and associated factors of retinal microvascular abnormalities in children with type 1 diabetes mellitus (T1DM) without visual impairment and diabetic retinopathy (DR). Methods: Case-control hospital-based study including children with or without DM. Optical coherence tomography angiography (OCTA; CIRRUS HD-OCT model 5000) was used to scan 6 × 6 mm square area of posterior retina and optic disc. The indexes analyzed mainly included vascular length density (VD), perfusion density (PD), and foveal avascular zone area, perimeter, and morphology. Independent risk factors were analyzed by multifactor linear regression. Results: A total of 47 children with T1DM and 44 healthy subjects were enrolled. Statistical analysis showed that VD within 1 to 3 mm (inner ring) of the macula in the case group was smaller than that in the control group (18.561 ± 1.151/mm: 19.161 ± 0.464/mm; P< 0.001), and mother's excessive weight gain during pregnancy was an independent factor (P = 0.004); VD within 3 to 6 mm (outer ring) of the macula in the case group was smaller…

Purpose: We previously reported increased expression of cell proliferation and Jak-Stat pathway-related genes in chronic experimental glaucoma model optic nerve heads (ONH) with early, mild injury. Here, we confirm these observations by localizing, identifying, and quantifying ONH cellular proliferation and Jak-Stat pathway activation in this model. Methods: Chronic intraocular pressure (IOP) elevation was achieved via outflow pathway sclerosis. After 5 weeks, ONH longitudinal sections were immunolabeled with proliferation and cell-type markers to determine nuclear densities in the anterior (unmyelinated) and transition (partially myelinated) ONH. Nuclear pStat3 labeling was used to detect Jak-Stat pathway activation. Nuclear density differences between control ONH (uninjected) and ONH with either early or advanced injury (determined by optic nerve injury grading) were identified by ANOVA. Results: Advanced injury ONH had twice the nuclear density (P < 0.0001) of controls and significantly greater astrocyte density in anterior (P = 0.0001) and transition (P = 0.006) ONH regions. An…

Purpose: To investigate the therapeutic effects of targeting signal transducer and activator of transcription-3 (STAT3) activation on the ocular surface damage of dry eye in mice. Methods: Adult Balb/C and C57BL/6 mice with benzalkonium chloride (BAC) treatment, lacrimal gland excision, and meibomian gland dysfunction were used as dry eye models. The levels of phosphorylated STAT3 (p-STAT3) were detected with immunofluorescence staining and Western blotting. STAT3 inhibition was performed by topical application of STAT3 inhibitor S3I-201. Corneal epithelial barrier function, tear production, and conjunctival goblet cell density were quantified with fluorescein sodium staining, phenol red cotton test, and histochemical staining. The expressions of matrix metalloproteinase (MMP)-3/9, TUNEL, and inflammation cytokines were assessed with immunofluorescence staining, qPCR, and ELISA assays. The therapeutic effect of S3I-201 was further compared with the Janus kinase inhibitor tofacitinib and ruxolitinib. Results: Elevated levels of nuclear p-STAT3 were detected in the corneal and…

Purpose: To evaluate retinal vascular status during pregnancy by using optical coherence tomography angiography (OCTA). Methods: Women in their third trimester of pregnancy and nonpregnant age-matched women were recruited for this prospective, case-control study. Subjects were imaged with OCTA. Main outcome measures were foveal avascular zone parameters, perfusion density (PD) percentage in the superficial retinal capillary plexus (SCP), PD percentage in the deep retinal capillary plexus (DCP), SCP vessel length density (VLD), DCP-VLD, and choriocapillaris (CC) flow voids (i.e., flow deficits in the CC). Results: Nineteen eyes of 10 pregnant subjects and 44 eyes of 27 nonpregnant control women were included. Mean ages were 36 ± 7 and 35 ± 8 years (SD), respectively (P value = 0.78). Mean gestational age of pregnant women was 33 weeks (range = 29-39, SD = 3). There was a significant reduction in the SCP-PD in the entire scan and in the nasal Early Treatment Diabetic Retinopathy Study subfield (47.9 vs. 49.7, P = 0.04 and 49.3 vs. 51.6, P = 0.03, respectively) in the pregnant cohort…

Purpose: To evaluate changes in peripapillary microvascular parameters in the fellow eyes of patients with unilateral retinal vein occlusion (RVO) using optical coherence tomography angiography (OCTA) and to determine the relationships between peripapillary microvasculature and retinal nerve fiber layer (RNFL) and ganglion cell-inner plexiform layer (GC-IPL) thickness. Methods: Eighty-three patients with unilateral RVO (50 patients with branch RVO and 33 with central RVO) and 83 normal controls were enrolled. OCTA (Cirrus HD-OCT 5000 with AngioPlex) 6 × 6-mm scans centered on the optic disc were acquired. Peripapillary vessel density (VD) and perfusion density (PD) were automatically calculated. Results: The average RNFL and GC-IPL thicknesses in the fellow eyes of RVO patients were significantly thinner than in normal controls (93.5 vs. 96.6 μm, P = 0.013 and 81.3 vs. 84.1 μm, P = 0.003, respectively). In the fellow eyes of patients with unilateral RVO, the peripapillary VD of the inner ring, outer ring, and full area (17.47, 18.50, and 17.89, respectively) were significantly lower…

Purpose: Mesenchymal stem cells (MSCs) have been extensively studied for their capacity to enhance wound healing and represent a promising research field for generating cell therapies for corneal scars. In the present study, we investigated MSCs from different tissues and their potential to differentiate toward corneal keratocytes. Methods: Adipose-derived stem cells, bone marrow MSCs, umbilical cord stem cells, and corneal stromal stem cells (CSSCs) were characterized by their expression of surface markers CD105, CD90, and CD73, and their multilineage differentiation capacity into adipocytes, osteoblasts, and chondrocytes. MSCs were also evaluated for their potential to differentiate toward keratocytes, and for upregulation of the anti-inflammatory protein TNFα-stimulated gene-6 (TNFAIP6) after simulation by IFN-γ and TNF-α. Results: Keratocyte lineage induction was achieved in all MSCs as indicated by the upregulated expression of keratocyte markers, including keratocan, lumican, and carbohydrate sulfotransferase. TNFAIP6 response to inflammatory stimulation was observed only in…

Purpose: In this study, the gut fungal microbiome of uveitis (UVT) patients was generated and compared with healthy controls (HC) to identify dysbiosis in UVT patients and ascertain the role of gut fungal microbiome in disease pathology. Methods: In the present study, gut fungal microbiomes were analyzed in the fecal samples of HC (n = 24) and UVT patients (n = 14) using high-throughput Illumina sequencing of ITS2 region of the fungal ribosomal RNA. QIIME and R software were used for data analysis. Results: The gut fungal richness and diversity were significantly decreased in UVT patients compared to HC. Our analyses showed enrichment of several pathogenic fungi including Malassezia restricta, Candida albicans, Candida glabrata, and Aspergillus gracilis in UVT patients. Heatmap and discriminatory OTUs further confirmed the disparities between UVT and HC microbiomes. Conclusions: This is the first study demonstrating dysbiosis in the gut fungal communities of UVT patients indicating the importance of fungal microbiome in the disease pathology. These initial findings might warrant…

Purpose: Reoxygenation after hypoxia can increase reactive oxygen species and upregulate autophagy. We determined, for the first time, the impact of elevated IOP on hypoxia induction, superoxide accumulation, and autophagy in a bead model of glaucoma. Method: Ocular hypertension was achieved with magnetic bead injection into the anterior chamber. Before mice were killed, they were injected with pimonidazole for hypoxia detection and dihydroethidium (DHE) for superoxide detection. Total retinal ganglion cells (RGCs) and optic nerve (ON) axons were quantified, total glutathione (GSH) was measured, and retinal and ON protein and mRNA were analyzed for hypoxia (Hif-1α and Hif-2α), autophagy (LC3 and p62), and SOD2. Results: With IOP elevation (P < 0.0001), the retina showed significantly (P < 0.001) decreased GSH compared with control, and a significant decrease (P < 0.01) in RGC density compared with control. Pimonidazole-positive Müller glia, microglia, astrocytes, and RGCs were present in the retinas after 4 weeks of ocular hypertension but absent in both the control and after only 2…

Purpose: Investigate the impact matrix metalloproteinase 14 (MMP14) delivered via exosomes produced by corneal fibroblasts on vascular endothelial growth factor receptor 1 (VEGFR1) cleavage on endothelial cells, and other key processes of angiogenesis. Methods: Proteolysis of VEGFR1 and R2 by the catalytic domain of MMP14 was investigated via immunocytochemistry with anti-VEGFR1, anti-VEGFR2, and anti-MMP14 antibodies. Exosomes were isolated via precipitation and serial ultracentrifugation from wild-type (WT) and MMP14 exon4-deficient corneal fibroblasts. Transmission electron microscopy and nanotracking analysis were used to characterize the isolated exosomes. The presence of MMP14 in exosomes from WT fibroblasts was confirmed by Western blotting. VEGFR1 cleavage upon treatment with WT-derived exosomes, Δexon4-derived exosomes, or the pan-MMP inhibitor GM60001 was examined via in vitro proteolysis analysis using recombinant mouse (rm) VEGFR1/R2. Endothelial cell migration and proliferation were investigated using a Boyden chamber assay and BrdU incorporation, respectively. Results:…

Purpose: To confirm the association of all reported common polymorphisms with POAG. Methods: We searched in PubMed and Web of Science (up to January 10, 2018) for genetic studies of POAG. All case control studies investigating the association between single-nucleotide polymorphisms (SNPs) and POAG risk were included. Odds ratios (ORs) with 95% confidence intervals (CIs) were calculated by fixed- or random-effect model. Results: This meta-analysis included 108 case control studies involving 35,389 POAG patients and 51,742 controls. The pooled results showed a significant association between 20 SNPs in 12 genes (148Asp/Glu in APE1 gene; rs449647 in APOE gene; rs1052990 and rs4236601 in CAV1/CAV2 gene; rs1799750 in MMP gene; c.603T3A (Met98Lys) in OPTN gene; rs7081455 in PLXDC2 gene; rs1279683 in SLC23A2 gene; 372 T/C in TIMP1 gene; rs1927911, rs2149356, rs4986791, rs7037117, and rs10759930 in TLR4 gene; rs4656461 in TMCO1 gene; 399Arg/Gln in XRCC1 gene; and rs540782, rs547984, and rs693421 in ZP4 gene) with POAG. Conclusions: Based on the current meta-analysis, we indicate 20 SNPs in…

Purpose: We assess if α retinal ganglion cells (αRGCs) and intrinsically photosensitive retinal ganglion cells (ipRGCs) survive in mouse models of glaucoma. Methods: Two microliters of N-methyl-D-aspartate (NMDA; 1 mM) or PBS were injected intraocularly 7 days before sacrifice. Immunohistochemical analyses of the retina were performed using antibodies against RNA-binding protein with multiple splicing (RBPMS), osteopontin, and melanopsin. Immunohistochemical analyses also were performed in adult mice with glutamate/aspartate transporter (GLAST) deletion (GLAST knockout [KO] mice), a mouse model of normal tension glaucoma. Results: NMDA-induced loss of RBPMS-positive total RGCs was 58.4% ± 0.4% compared to PBS-treated controls, whereas the loss of osteopontin-positive αRGCs was 5.0% ± 0.6% and that of melanopsin-positive ipRGCs was 7.6% ± 1.6%. In GLAST KO mice, the loss of total RGCs was 48.4% ± 0.9% compared to wild-type mice, whereas the loss of αRGCs and ipRGCs was 3.9% ± 0.4% and 9.3% ± 0.5%, respectively. The distribution of survived total RGCs, αRGCs, and ipRGCs was similar…

Purpose: To quantify retinal vascular density in amblyopic children and to investigate the relationship between superficial and deep retinal vessel density and retinal thickness. Method: In this prospective cross-sectional study, 85 amblyopic children (5-12 years) and 66 age-matched control subjects participated at a pediatric ophthalmology clinic. Macular vessel densities of the superficial and deep capillary plexuses (SCP, DCP, respectively) and retinal thickness were measured by clinical optical coherence tomographic angiography (OCTA). Vessel density and retinal thickness were compared between amblyopic groups and the control group, and correlations among these two variables were analyzed. Results: Of the 85 amblyopic children, 52 children had anisometropic amblyopia, 16 had strabismic amblyopia, and 17 had bilateral amblyopia. The foveal and parafoveal macular vessel density in the SCP was lower in amblyopic than control children (P ≤ 0.008). Compared to strabismic and bilateral amblyopias, anisometropic amblyopia SCP differed the most from controls (P ≤ 0.005). Macular vessel…

Several retinal conditions have been recently revealed by optical coherence tomography (OCT) to occur more frequently in glaucoma than in healthy eyes: paravascular defects, peripapillary retinoschisis, and pseudo-cysts of the inner nuclear layer (INL). Here the clinical OCT findings described in these reports are reviewed and a framework that could explain why they are related and occur more frequently in glaucoma is proposed. Evidence suggests that these conditions all share in common a strong tendency to develop in association with severe and/or rapidly progressing disease and a likelihood of involving biomechanical forces and differential tissue deformation. Müller glia are mechanosensitive and known to react to shear and axial strain, and to participate in homeostasis of water and ion flux through the retina, and to provide spring-like capability to buffer of mechanical forces. Thus, Müller cell integrity is also likely to be involved in the development and/or response to such events. OCT has also revealed that Müller cell optical properties (scatter and attenuation) appear to…

Purpose: Metastatic uveal melanoma (UM) has a very poor prognosis and no effective therapy. Despite remarkable advances in treatment of cutaneous melanoma, UM remains recalcitrant to chemotherapy, small-molecule kinase inhibitors, and immune-based therapy. Methods: We assessed two sets of oxidative phosphorylation (OxPhos) genes within 9858 tumors across 31 cancer types. An OxPhos inhibitor was used to characterize differential metabolic programming of highly metastatic monosomy 3 (M3) UM. Seahorse analysis and global metabolomics profiling were done to identify metabolic vulnerabilities. Analyses of UM TCGA data set were performed to determine expressions of key OxPhos effectors in M3 and non-M3 UM. We used targeted knockdown of succinate dehydrogenase A (SDHA) to determine the role of SDHA in M3 UM in conferring resistance to OxPhos inhibition. Results: We identified UM to have among the highest median OxPhos levels and showed that M3 UM exhibits a distinct metabolic profile. M3 UM shows markedly low succinate levels and has highly increased levels of SDHA, the enzyme that couples…

Purpose: To determine whether equiluminant artificial dynamic ON or OFF stimuli on a computer screen can induce bidirectional changes in choroidal thickness (ChTh) in both humans and chickens, and whether such changes are associated with bidirectional changes in retinal dopamine release in chickens. Methods: Experiment 1: Before and after ON or OFF stimulation for 1 hour, ChTh was measured with optical coherence tomography (OCT). Experiment 2: chicks (n = 14) were raised under ON or OFF stimulation for 3 hours. ChTh was determined by OCT. Experiment 3: chicks were raised for 7 days either under room light (500 lux, n = 11), dynamic ON stimulus (700 lux, n = 15), or dynamic OFF stimulus (700 lux, n = 7). In addition, negative lenses were attached to their right eyes. After experiments 2 and 3, retinal and vitreal dopamine (DA), and its metabolites, were measured by HPLC-electrochemical detection. Results: Experiment 1: Dynamic ON stimuli caused thicker choroids (+5.3 ± 2.0 μm), whereas OFF stimuli caused choroidal thinning (-4.7 ± 0.5 μm) (right eye data only, P < 0.001). Experiment…

Purpose: To assess the variations induced by exercise in retinal vascular density (VD), foveal avascular zone (FAZ) area, and fractal dimension (FD) at the superficial (SCP) and deep (DCP) capillary plexa in healthy subjects by means of optical coherence tomography angiography (OCT-A). Methods: Consecutive healthy subjects were prospectively included into two groups, ranging in age from 18 to 29 years for group 1 and from 30 to 40 years for group 2. Data from 3 × 3-mm OCT-A acquisition centered on the macula at SCP and DCP (VD, FAZ area, and FD), heart rate, and systolic-diastolic blood pressure were collected before and after a 20-minute standardized physical exercise on a stationary bicycle. Results: Both eyes of 32 healthy volunteers were prospectively included (15 in group 1 and 17 in group 2). Mean age was 27 ± 7 years. In the overall analysis and for each group, a decrease of VD at the level of SCP and an increase of FD at the level of DCP were significant after exercise. A significant correlation was found between these modifications of retinal vascularization and the…

Purpose: To define the nature and extent of cone photoreceptor abnormalities in diabetic individuals who have mild or no retinopathy by assessing the activation phase of cone phototransduction and the flicker ERG in these individuals. Methods: Light-adapted single-flash and flicker ERGs were recorded from 20 diabetic individuals who have no clinically apparent retinopathy (NDR), 20 diabetic individuals who have mild nonproliferative diabetic retinopathy (NPDR), and 20 nondiabetic, age-equivalent controls. A-waves elicited by flashes of different retinal illuminance were fit with a delayed Gaussian model to derive Rmp3 (maximum amplitude of the massed photoreceptor response) and S (phototransduction sensitivity). Fundamental amplitude and phase of ERGs elicited by full-field sinusoidal flicker were obtained across a frequency range of 6 to 100 Hz. Results: ANVOA indicated that both diabetic groups had significant S losses compared with the controls, whereas mean Rmp3 did not differ significantly among the groups. ANOVA also indicated significantly reduced flicker ERG amplitude for…

Purpose: To explore the characteristics of tilted optic disc and peripapillary atrophy (PPA), and their associations with choroidal thickness (ChT) in young myopic patients. Methods: A total of 821 patients were enrolled in this cross-sectional study. Optic disc tilt ratio, PPA area, macular ChT (mChT), and peripapillary ChT (pChT) were measured. Subjects were divided into four groups purely on the basis of the axial length (AL). Relationships between ChT and the morphologic characteristics of optic disc were analyzed using logistic regression. Results: The prevalence of tilted optic disc and PPA increased as myopia severity increased. Every 0.1-mm2 increase in PPA area was associated with a 14.93-μm decrease in mChT and a 9.54-μm decrease in pChT; and every 0.1 increase in tilt ratio was correlated with a 5.38-μm increase in mChT and a 6.21 decrease in pChT. After stratifying by myopia severity, these trends were still observed in the high myopia group. A larger PPA area (odds ratio [OR] = 2.33; P < 0.01), a longer AL (OR = 1.34; P < 0.01), an increased pChT (OR = 1.11; P < 0.01),…

Purpose: The purpose of this study was to evaluate the effect of removal of Descemet's basement membrane and endothelium compared with removal of the endothelium alone on posterior corneal fibrosis. Methods: Twelve New Zealand White rabbits were included in the study. Six eyes had removal of the Descemet's membrane-endothelial complex over the central 8 mm of the cornea. Six eyes had endothelial removal with an olive-tipped cannula over the central 8 mm of the cornea. All corneas developed stromal edema. Corneas in both groups were cryofixed in optimum cutting temperature (OCT) formula at 1 month after surgery. Immunohistochemistry (IHC) was performed for α-smooth muscle actin (SMA), keratocan, CD45, nidogen-1, vimentin, and Ki-67, and a TUNEL assay was performed to detect apoptosis. Results: Six of six corneas that had Descemet's membrane-endothelial removal developed posterior stromal fibrosis populated with SMA+ myofibroblasts, whereas zero of six corneas that had endothelial removal alone developed fibrosis or SMA+ myofibroblasts (P < 0.01). Myofibroblasts in the fibrotic zone…

Purpose: To measure the deformation of the human optic nerve head (ONH) and peripapillary tissue (PPT) in response to acute intraocular pressure (IOP) elevation. Methods: The ONH and PPT of 14 human donor globes were imaged with high-frequency ultrasonography during inflation testing from 5 to 30 mm Hg. A correlation-based speckle tracking algorithm was used to compute tissue displacements, and the through-thickness, in-plane, and shear strains were calculated by using least-squares strain estimation methods. The ONH and PPT were segmented along the anterior-posterior direction and the nasal-temporal direction. Regional displacements and strains were analyzed and compared. Results: The ONH displaced more posteriorly than the PPT in response to an acute IOP increase. Scleral canal expansion was minimal but correlated with ONH posterior displacement at all IOP levels. Through-thickness compression was concentrated in the anterior of both the ONH and the PPT. Shear was concentrated in the vicinity of the canal with higher shear in the peripheral ONH than the central ONH and higher…

Purpose: DNA methylation is a key epigenetic modification involved in various biological processes and diseases. Corneal epithelial wound healing (CEWH) is essential for restoring corneal integrity and transparency after injury. However, the role of DNA methylation in CEWH remains elusive. Here, we investigate the function and underlying mechanism of DNA methylation in regulating CEWH. Methods: Dot blots and global methylation assays determined DNA methylation levels during CEWH. Quantitative RT-PCR and Western blot analysis examined the expression of DNA methyltransferases (DNMTs), cyclin-dependent kinase inhibitor 2B (CDKN2B), and miR-200a during CEWH, respectively. MTS assays and flow cytometry were used to analyze human corneal epithelial cell (HCEC) proliferation and cell cycle, respectively. The in vitro scratch wound assay assessed HCEC migration and an in vivo murine corneal epithelial debridement model evaluated wound healing. Using bisulfite sequencing PCR, we determined the DNA methylation status of the candidate genes. Transfection of miR-200a mimic or inhibitor assessed…

Purpose: A chemical approach to examine the role of Wnt signaling in maintaining the stemness and/or proliferation of limbal stem/progenitor cells (LSCs). Methods: LSCs were isolated from human donor eyes and cultured as single cells for 12 to 14 days with the following small molecules: IIIC3, an antagonist of the Wnt signaling inhibitor Dickkopf (DKK), and IC15, a Wnt signaling inhibitor. Proliferation of LSCs in the presence of IIIC3 and IC15 was determined by the number of cells and colonies established. Maintenance of stemness was determined by p63α, cytokeratin (K)12, and K14 expression. Results: Activation of Wnt, through IIIC3-mediated DKK inhibition, resulted in similar colony forming efficiency (CFE) as in the untreated LSCs, but significantly increased the number of cultivated cells 7.21% with 5 μM. Inhibition of Wnt with IC15 significantly reduced the CFE (P ≤ 0.01) and the number of cultivated cells by 16% to 29%. Percentage of cells expressing high levels of p63α (p63αbright) and quantity of small cells (≤12 μm), which contain the LSCs, increased 4.71% and 11.26% (both…

Purpose: Long noncoding RNAs (lncRNAs) are important in disease progression and cellular functions. This study aimed to conduct global lncRNA profiling and characterize the role of lncRNA 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase delta 3-sence RNA 1 (PLCD3-OT1) in the progression of age-related cataract (ARC). Methods: We performed lncRNA expression profiling of lens capsule from ARC groups and age-matched groups using high-throughput RNA-sequencing. Real-time PCR was conducted to detect the expression pattern of lncRNA and mRNA in the clinical samples and cell model. Assays of cell-counting kit-8, 5'-ethynyl-2'-deoxyuridine, TUNEL, and propidium iodide staining were used to detect cell viability, proliferation, apoptosis, and cell cycle. We also performed fluorescence in situ hybridization assay to detect the location of lncRNA, and verified the endogenous competitive RNA mechanism between miRNAs, lncRNAs, and target genes via double-luciferase reporter analyses. Results: The expression of lncRNA PLCD3-OT1 and PLCD3 were significantly decreased in ARC. PLCD3-OT1…

Purpose: The purpose of this study was to characterize the ability of applied electrical fields (EFs) to direct retinal ganglion cell (RGC) axon growth as well as to assess whether Rho GTPases play a role in translating electrical cues to directional cues. Methods: Full-thickness, early postnatal mouse retina was cultured in electrotaxis chambers and exposed to EFs of varying strengths (50-200 mV/mm). The direction of RGC axon growth was quantified from time-lapsed videos. The rate of axon growth and responsiveness to changes in EF polarity were also assessed. The effect of toxin B, a broad-spectrum inhibitor of Rho GTPase signaling, and Z62954982, a selective inhibitor of Rac1, on EF-directed growth was determined. Results: In the absence of an EF, RGC axons demonstrated indiscriminate directional growth from the explant edge. Retinal cultures exposed to an EF of 100 and 200 mV/mm showed markedly asymmetric growth, with 74.2% and 81.2% of axons oriented toward the cathode, respectively (P < 0.001). RGC axons responded to acute changes in EF polarity by redirecting their growth…

Purpose: Visual (retinoid) cycle anomalies induce aberrant build-up of all-trans retinal (atRAL) in the retinal pigment epithelium (RPE), which is a cause of RPE atrophy in Stargardt disease type 1 and age-related macular degeneration. NLR family pyrin domain containing 3 (NLRP3) inflammasome activation is implicated in the etiology of age-related macular degeneration. Here, we elucidated the relationship between NLRP3 inflammasome activation and atRAL-induced death of RPE cells. Methods: Cellular toxicities were assessed by MTS or MTT assays. Expression levels of mRNAs and proteins were determined by quantitative reverse transcription-polymerase chain reaction, Western blotting, or enzyme-linked immunosorbent assay. Fluorescence microscopy was used to examine intracellular signals. Ultrastructural features of organelles were examined by transmission electron microscope. Results: Abnormal accumulation of atRAL was associated with a significant increase in the proportion of human ARPE-19 cells exhibiting features of apoptosis and Caspase-3/gasdermin E (GSDME)-mediated pyroptosis.…

Purpose: To investigate structural changes in the retinal outer layers and choroid using adaptive optics (AO) and optical coherence tomography (OCT) in eyes with myopia, and to correlate the changes with decreased macular light sensitivity (MLS). Methods: This prospective study included 27 subjects with emmetropia and low myopia (EM/LM), 25 with moderate myopia (MM), and 25 with high myopia (HM). Microperimetry was used to quantify MLS in each subject, while AO and OCT images of fundus were analyzed to quantify cone density and regularity and thickness of outer retinal sublayers and choroid. Differences of MLS, cone distribution, and chorioretinal thicknesses were compared among the three groups, and the associations among photoreceptor morphological alterations, MLS, and other parameters were analyzed. Results: In HM, the MLS, cone density and regularity, and thicknesses of the myoid and ellipsoid zone (MEZ), Henle fiber layer and outer nuclear layer, interdigitation zone and RPE/Bruch complex, and choroid were lower than in EM/LM. Decreased MLS was correlated with lower cone…

Purpose: Several pieces of epidemiologic evidence have indicated PM2.5 (particulate matter with a diameter of 2.5 μm or less) as a causing factor of allergic conjunctivitis, but without experimental elucidation of mechanism. In the present study, PM2.5 in solution was directly applied to the mouse ocular surface to elucidate whether PM2.5 might cause allergic conjunctivitis, and its underlying mechanisms were analyzed. Methods: ICR mice were challenged for 18 consecutive days with eye drops containing PM2.5 at 3.2, 6.4, and 12.8 mg/mL in 0.9% NaCl saline, along with the controls prepared in parallel without PM2.5 and another control group treated with both PM2.5 at 12.8 mg/mL and artificial tears. On day 19, the whole eyes and meibomian glands were harvested for histopathological analyses and assessment of clinical scoring, tear volume, tear breakup time, and tear ferning. Furthermore, goblet cells by periodic acid Schiff stain and infiltrated eosinophils by Giemsa stain were quantified and compared among study groups. Results: Clinical scoring showed more eyelid edema, tearing, and…

Purpose: Cigarette smoking is a known risk factor for vascular dysfunction. This study evaluated choroidal structural changes in smokers using the choroidal vascularity index (CVI) derived from image binarization on spectral domain optical coherence tomography scans with enhanced depth imaging (EDI-OCT). Methods: This cross-sectional study included 39 smokers and 44 non-smokers. Choroidal images on EDI-OCT were binarized into luminal area (LA) and stromal area (LA). CVI was calculated as the ratio of LA to total choroid area (TCA). CVI, foveal retinal thickness (FRT), and subfoveal choroidal thickness (SFCT) between smokers and non-smokers were compared using likelihood ratio test with linear mixed model. Trend and subgroup analysis were performed to investigate the dose-dependent relationship between CVI/FRT/SFCT and pack-years. Results: CVI in smokers (65 ± 2%) was lower compared to non-smokers (67 ± 2%, P = 0.0001). The difference remained significant after adjusting for age (P = 0.001). There was no significant association between cigarette smoking and FRT/SFCT. CVI decreased by…

Purpose: To investigate macular vascular alterations by using optical coherence tomography angiography (OCTA) in patients with a history of long-term anti-vascular endothelial growth factor (VEGF) therapy for neovascular age-related macular degeneration (nAMD). Methods: Japanese patients with nAMD with a history of long-term anti-VEGF monotherapy at study entry were studied retrospectively. OCTA images were obtained, and the vessel densities (mm-1) of the superficial capillary plexus, deep capillary plexus (DCP), choriocapillaris (CC), and the plexus foveal avascular zone area (mm2) were calculated. Results: One hundred twenty-four eyes (124 patients) were included. The mean ± standard deviation follow-up period between the first and last OCTA imaging sessions was 14.5 ± 3.1 months; the duration of the anti-VEGF monotherapy before the first OCTA imaging session was 68.0 ± 23.6 months, with a mean of 3.6 ± 3.0 injections during the follow-up period. The vessel densities of the DCP and CC significantly decreased (P = 0.001 and P = 0.009, respectively) from 10.62 ± 2.72 mm-1 and 11.84…