Diagnostic Role of Oral Fluorescein Angiography in Pediatric Ambulatory Clinics.

Oral ingestion of fluorescein can be done in ambulatory pediatric clinics. We show that oral ultra-widefield fluorescein angiography is a non-invasive approach to rapidly diagnose and manage a diverse set of pediatric retinal vascular diseases.

Immunology of Retinitis Pigmentosa and Gene Therapy-Associated Uveitis.

The underlying immune state of inherited retinal degenerations (IRDs) and retinitis pigmentosa (RP) has been an emerging area of interest, wherein the consequences have never been greater given the widespread recognition of gene therapy-associated uveitis (GTU) in gene therapy clinical trials. Whereas some evidence suggests that the adaptive immune system may play a role, the majority of studies indicate that the innate immune system is likely the primary driver of neuroinflammation in RP. During retinal degeneration, discrete mechanisms activate resident microglia and promote infiltrating macrophages that can either be protective or detrimental to photoreceptor cell death. This persistent stimulation of innate immunity, overlaid by the introduction of viral antigens as part of gene therapy, has the potential to trigger a complex microglia/macrophage-driven proinflammatory state. A better understanding of the immune pathophysiology in IRD and GTU will be necessary to improve the success of developing novel treatments for IRDs.

RARE CASE OF ABIOTROPHIA DEFECTIVA BLEB-ASSOCIATED ENDOPHTHALMITIS.

PURPOSE: To report a rare case of Abiotrophia defectiva bleb-associated endophthalmitis. METHODS: In this case report of a patient with bleb-associated endophthalmitis, the authors describe the features of severe and rapid onset, associated retinitis, and a favorable outcome after aggressive early intervention. RESULTS: A 75-year-old woman presented decreased visual acuity of hand motions with an elevated intraocular pressure of 39 mmHg in the left eye. Her ocular history was notable for severe primary open-angle glaucoma with uneventful trabeculectomy and cataract surgery in both eyes 7 years before. Examination revealed conjunctival injection and an opaque avascular cystic bleb along with anterior chamber cellular reaction with a B-scan ultrasound concerning for vitritis. A. defectiva was isolated from the vitreous tap within 48 hours and confirmed later with 16S ribosomal RNA sequencing. After initial vitreous tap and inject and subsequent pars plana vitrectomy, her best-corrected visual acuity had improved to 20/500 at postoperative week one and then improved to 20/40 at postoperative month 2. Her examination was notable for resolved corneal edema, a deep and quiet anterior chamber, and resolved intraretinal hemorrhages. CONCLUSION: This is one of the first bleb-associated endophthalmitis cases with rapid identification of A. defectiva . A. defectiva was isolated from the initial vitreous tap within 48 hours and confirmed with 16S ribosomal RNA sequencing. This case highlights that rapid identification of A. defectiva may be indicative of a greater bacterial load and should prompt aggressive intervention and that the visual prognosis can be favorable.

Targeted long-read sequencing allows for rapid identification of pathogenic disease-causing variants in retinoblastoma.

BACKGROUND: Identification of disease-causing variants of the retinoblastoma gene (RB1), the predominant cause of retinoblastoma, is challenging. Targeted long-read genome sequencing offers a novel approach to resolve the diverse range of pathogenic variants in RB1 and provides haplotype information rapidly. MATERIALS AND METHODS: Genomic DNA was isolated from a venipuncture blood draw of a retinoblastoma patient. Whole genome sequencing (WGS) was carried out using the short-read Ilumina platform. WGS and targeted sequencing of RB1 was accomplished using the long-read Oxford Nanopore Technologies (ONT) platform. Deep-learning frameworks allowed haplotagging, variant calling, and variant annotation of both short- and long-read data. RESULTS: Targeted long-read sequencing of the RB1 gene allowed for enhanced depth of read coverage for discovery of rare variants and haplotype analysis. A duplication leading to a frameshift and early termination in RB1 was identified as the most deleterious variant by all sequencing methods, with long-read technology providing additional information of methylation signal and haplotype information. More importantly, there was greater than 98% concordance of RB1 variants identified between short-read and targeted long-read sequencing modalities. CONCLUSIONS: Targeted long-read technology allows for focused sequencing effort for variant discovery. Application of this for the first time in a retinoblastoma patient allowed haplotagged variant identification and demonstrated excellent concordance with benchmark short-read sequencing. The added benefit of targeted long-read sequencing to resolve disease-causing genomic variation in RB1 rapidly from a blood draw will provide a more definitive diagnosis of heritable RB and guide management decisions for patients and their families.

Frequency of Urgent or Emergent Vitreoretinal Surgical Procedures in the United States During the COVID-19 Pandemic.

Importance: The American Academy of Ophthalmology (AAO) indicated that urgent or emergent vitreoretinal surgical procedures should continue during the coronavirus disease 2019 (COVID-19) pandemic. Although decreases in the frequency of critical procedures have been reported outside the field of ophthalmology, analyses are limited by volume, geography, and time. Objective: To evaluate whether the frequency of ophthalmic surgical procedures deemed urgent or emergent by the AAO changed across the United States during the COVID-19 pandemic. Design, Setting, and Participants: Vitreoretinal practices from 17 institutions throughout the US participated in this multicenter cross-sectional study. The frequency of 11 billed vitreoretinal Current Procedural Terminology (CPT) codes across respective weeks was obtained from each practice between January 1, 2019, and May 31, 2020. Data were clustered into intravitreal injections (code 67028), lasers and cryotherapy (codes 67141, 67145, and 67228), retinal detachment (RD) repairs (codes 67107, 67108, 67110, and 67113), and other vitrectomies (codes 67036, 67039, and 67040). Institutions were categorized by region (Northeast, Midwest, South, and West Coast), practice setting (academic [tax-exempt] or private [non-tax-exempt]), and date of respective statewide stay-at-home orders. Main Outcomes and Measures: Nationwide changes in the frequency of billing for urgent or emergent vitreoretinal surgical procedures during the COVID-19 pandemic. Results: A total of 526 536 CPT codes were ascertained: 483 313 injections, 19 257 lasers or cryotherapy, 14 949 RD repairs, and 9017 other vitrectomies. Relative to 2019, a weekly institutional decrease in injections was observed from March 30 to May 2, 2020, with a maximal 38.6% decrease (from a mean [SD] of 437.8 [436.3] to 273.8 [269.0] injections) from April 6 to 12, 2020 (95% CI, -259 to -69 injections; P = .002). A weekly decrease was also identified that spanned a longer interval, at least until study conclusion (March 16 to May 31, 2020), for lasers and cryotherapy, with a maximal 79.6% decrease (from a mean [SD] of 6.6 [7.7] to 1.5 [2.0] procedures) from April 6 to 12, 2020 (95% CI, -6.8 to -3.3 procedures; P < .001), for RD repairs, with a maximal 59.4% decrease (from a mean [SD] of 3.5 [4.0] to 1.6 [2.2] repairs) from April 13 to 19, 2020 (95% CI, -2.7 to -1.4 repairs; P < .001), and for other vitrectomies, with a maximal 84.3% decrease (from a mean [SD] of 3.0 [3.1] to 0.4 [0.8] other vitrectomies) from April 6 to 12, 2020 (95% CI, -3.3 to -1.8 other vitrectomies; P < .001). No differences were identified by region, setting, or state-level stay-at-home order adjustment. Conclusions and Relevance: Although the AAO endorsed the continued performance of urgent or emergent vitreoretinal surgical procedures, the frequency of such procedures throughout the country experienced a substantial decrease that may persist after the COVID-19 pandemic's initial exponential growth phase. This decrease appears independent of region, setting, and state-level stay-at-home orders. It is unknown to what extent vitreoretinal intervention would have decreased without AAO recommendations, and how the decrease is associated with outcomes. Although safety is paramount during the COVID-19 pandemic, practices should consider prioritizing availability for managing high-acuity conditions until underlying reasons for the reduction are fully appreciated.

Relationship of Epiretinal Membrane Formation and Macular Edema Development in a Large Cohort of Uveitic Eyes.

Purpose: To identify the temporal relationship and clinical characteristics of epiretinal membrane (ERM) and macular edema (ME) formation in uveitic eyes.Methods: A total of 269 subjects (444 uveitic eyes) met study inclusion criteria. Comprehensive ophthalmic examination, spectral domain-optical coherence tomography (SD-OCT), and clinical testing were carried out.Results: Of the 444 uveitic eyes, 229 eyes (51.6%) developed an ERM, whereas 87 eyes (19.1%) developed ME. The odds ratios (ORs) of systemic disease causing uveitis and resulting in ERM and ME were significantly higher in posterior uveitis (OR 6.56, 95% CI 2.98-14.46; p < .0001) and panuveitis (OR 10.09, 95% CI 4.05-25.15; p < .0001). Temporal analysis revealed that an ERM was noted concurrently or prior to ME development in 93.8% of eyes.Conclusions: ERM and ME are primarily observed in posterior uveitis and panuveitis associated with systemic diseases. The temporal relationship highlights the importance of characterization of ERM as it relates to the development of uveitic ME.

Quantifying Subclinical and Longitudinal Microvascular Changes Following Episcleral Plaque Brachytherapy Using Spectral Domain-Optical Coherence Tomography Angiography.

PURPOSE: To assess longitudinal microvascular changes in eyes treated with I-125 episcleral plaque brachytherapy (EPB). METHODS: High resolution OCT angiograms of the central 3×3mm macula were obtained from I-125 episcleral plaque brachytherapy treated and untreated fellow eyes of 61 patients. Capillary density (vessel skeleton density, VSD) and caliber (vessel diameter index, VDI) were quantified using previously validated semi-automated algorithms. Nonperfusion was also quantified as flow impairment regions (FIR). Exams from treated and fellow eyes obtained pre-treatment and at 6-month, 1-year, and 2-year intervals were compared using generalized estimating equation linear models. Dosimetry maps were used to evaluate spatial correlation between radiation dose and microvascular metrics. RESULTS: At 6 months, treated eyes had significantly lower VSD (0.145 ± 0.003 vs 0.155 ± 0.002; p = 0.009) and higher FIR (2.01 ± 0.199 vs 1.46 ± 0.104; p = 0.010) compared to fellow eyes. There was a significant decrease in VSD and a corresponding increase in FIR even for treated eyes without clinically identifiable retinopathy at 6 months. VDI was significantly higher in treated eyes than in fellow eyes at 2 years (2.92 ± 0.025 vs 2.84 ± 0.018; p < 0.001). When our cohort was categorized into low dose radiation (<15Gy) and high dose radiation (>45Gy) to the fovea, there were significant differences in VSD and FIR between groups. CONCLUSIONS: OCTA can be used to quantify and monitor EPB induced retinopathy, and can detect vascular abnormalities even in the absence of clinically observable retinopathy. OCTA may therefore be useful in investigating treatment interventions that aim to delay EPB-induced radiation retinopathy.

Unilateral Multifocal Choroidal Melanoma.

We report a case of multifocal choroidal melanoma in the same eye, separated in presentation by 20 years. A 57-year-old Caucasian male initially presented with a choroidal melanoma of the right eye that was treated with transpupillary thermotherapy. Due to recurrence, the patient underwent proton beam therapy with subsequent tumor regression. A second small choroidal lesion was noted in the right eye during his surveillance examinations that was closely monitored and demonstrated stable dimensions and features suggestive of a choroidal nevus. Twenty years after his first presentation, the second lesion exhibited accelerated growth with imaging studies indicative of transformation to a distinct choroidal melanoma. The patient underwent a second globe salvage treatment of proton beam therapy. We describe the clinical course, radiographic, and imaging findings of this rare choroidal melanoma.

Systems pharmacology identifies drug targets for Stargardt disease-associated retinal degeneration.

A systems pharmacological approach that capitalizes on the characterization of intracellular signaling networks can transform our understanding of human diseases and lead to therapy development. Here, we applied this strategy to identify pharmacological targets for the treatment of Stargardt disease, a severe juvenile form of macular degeneration. Diverse GPCRs have previously been implicated in neuronal cell survival, and crosstalk between GPCR signaling pathways represents an unexplored avenue for pharmacological intervention. We focused on this receptor family for potential therapeutic interventions in macular disease. Complete transcriptomes of mouse and human samples were analyzed to assess the expression of GPCRs in the retina. Focusing on adrenergic (AR) and serotonin (5-HT) receptors, we found that adrenoceptor α 2C (Adra2c) and serotonin receptor 2a (Htr2a) were the most highly expressed. Using a mouse model of Stargardt disease, we found that pharmacological interventions that targeted both GPCR signaling pathways and adenylate cyclases (ACs) improved photoreceptor cell survival, preserved photoreceptor function, and attenuated the accumulation of pathological fluorescent deposits in the retina. These findings demonstrate a strategy for the identification of new drug candidates and FDA-approved drugs for the treatment of monogenic and complex diseases.

Topology of class A G protein-coupled receptors: insights gained from crystal structures of rhodopsins, adrenergic and adenosine receptors.

Biological membranes are densely packed with membrane proteins that occupy approximately half of their volume. In almost all cases, membrane proteins in the native state lack the higher-order symmetry required for their direct study by diffraction methods. Despite many technical difficulties, numerous crystal structures of detergent solubilized membrane proteins have been determined that illustrate their internal organization. Among such proteins, class A G protein-coupled receptors have become amenable to crystallization and high resolution X-ray diffraction analyses. The derived structures of native and engineered receptors not only provide insights into their molecular arrangements but also furnish a framework for designing and testing potential models of transformation from inactive to active receptor signaling states and for initiating rational drug design.

Nitrogenase complexes: multiple docking sites for a nucleotide switch protein.

Adenosine triphosphate (ATP) hydrolysis in the nitrogenase complex controls the cycle of association and dissociation between the electron donor adenosine triphosphatase (ATPase) (Fe-protein) and its target catalytic protein (MoFe-protein), driving the reduction of dinitrogen into ammonia. Crystal structures in different nucleotide states have been determined that identify conformational changes in the nitrogenase complex during ATP turnover. These structures reveal distinct and mutually exclusive interaction sites on the MoFe-protein surface that are selectively populated, depending on the Fe-protein nucleotide state. A consequence of these different docking geometries is that the distance between redox cofactors, a critical determinant of the intermolecular electron transfer rate, is coupled to the nucleotide state. More generally, stabilization of distinct docking geometries by different nucleotide states, as seen for nitrogenase, could enable nucleotide hydrolysis to drive the relative motion of protein partners in molecular motors and other systems.