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Purpose To compare choroidal vascular features of eyes with and without subretinal drusenoid deposits (SDD), using swept-source optical coherence tomography (SS OCT). Design Multicenter, cross-sectional study. Methods We prospectively recruited patients with intermediate age-related macular degeneration (AMD), without other vitreoretinal pathology. All participants underwent complete ophthalmic examination, color fundus photography (used for AMD staging), and spectral-domain OCT (to evaluate the presence of SDD). SS OCT was used to obtain automatic macular choroidal thickness (CT) maps, according to the Early Treatment Diabetic Retinopathy Study (ETDRS) sectors. For data analysis, we considered mean choroidal thickness as the arithmetic mean value of the 9 ETDRS sectors. SS OCT en face images of choroidal vasculature were also captured and converted to binary images. Choroidal vascular density (CVD) was calculated as a percent area occupied by choroidal vessels in a 6-mm-diameter submacular circular. Choroidal vessel volume was calculated by multiplying the average CVD by macular area and CT. Multilevel mixed linear models (to account for the inclusion of 2 eyes of same subject) were performed for analysis. Results We included 186 eyes (n = 118 subjects), 94 (50.5%) presenting SDD. Multiple regression analysis revealed that, controlling for age, eyes with SDD presented a statistically thinner mean CT (s = −21.9, P = .006) and CT in all the individual ETDRS fields (s ≤ −18.79, P ≤ .026). Mean choroidal vessel volume was also significantly reduced in eyes with SDD (s = −0.003, P = .007). No significant associations were observed with mean CVD. Conclusion In subjects with intermediate AMD, choroidal thickness and vessel volume are reduced in the presence of subretinal drusenoid deposits.

Purpose To investigate the inter-individual variability in duration of anti-vascular endothelial growth factor (VEGF) treatment effect in neovascular age-related macular degeneration (nvAMD). Design Prospective observational multi-centered study. Participants Forty-eight patients with nvAMD treated with anti-VEGF injections were included. Both treatment naive (n=25) as well as patients who had previously received treatment with ranibizumab (n=23) more than one month prior to their enrollment were recruited. Methods Patients received injection with ranibizumab (0.5 mg/0.05 ml) and were followed weekly for 4 weeks with spectral-domain OCT (SD-OCT) assessing the time to maximal reduction of central retinal thickness (CRT) and the presence of intraretinal and subretinal fluid. Other data collected included age, gender, visual acuity, axial length, lens status, and previous injections. The Shapiro-Wilk test was used to examine normal distributions for all variables. Correlations were examined by calculating Spearman's correlation coeficient. Distributions of quantitative variables are described as means (±SD). Qualitative variables are summarized by counts and percentage. Main Outcome Measures Time to maximal reduction of CRT and intra- and subretinal fluid after ranibizumab injection. Results A total of 48 eyes of 48 patients (age 74.8±8.3 years, 62.5% female, 52% treatment naive, 35.4% pseudophakic) were assessed. Two-thirds (64.6%) reached maximal CRT reduction earlier than the standard 4-week interval: 6.3% at 1 week postinjection, 22.9% at 2 weeks postinjection, and 35.4% at 3 weeks postinjection. Only 35.4% of patients had maximal CRT reduction at 4 weeks. Twenty percent of treatment-naive and 34.8% of non-naive patients had a week-4 CRT that was >35 μm thicker than the earlier occuring lowest CRT value (nadir). The time to maximal CRT reduction was not related to axial length, age, lens status, or history of injections. Conclusions Optimal dosing interval for maximal CRT reduction may be less than 4 weeks for a significant proportion of patients. Most patients will be classified as complete responders if intervals less than 4 weeks are used to assess anti-VEGF treatment response. Disease load rather than eye size appears to be the driver of anti-VEGF treatment duration and therefore, dosing interval needs to be optimized in the cohort of short-term responders.

Significance Extracellular tissue debris accumulates with aging and in the most prevalent central-vision-threatening eye disorder, age-related macular degeneration (AMD). In this work, we discovered that lysosome-associated membrane protein-2 (LAMP2), a glycoprotein that plays a critical role in lysosomal biogenesis and maturation of autophagosomes/phagosomes, is preferentially expressed in the outermost, neuroepithelial layer of the retina, the retinal pigment epithelium (RPE), and contributes to the prevention of ultrastructural changes in extracellular basolaminar deposits including lipids and apolipoproteins. LAMP2 thus appears to play an important role in RPE biology, and its apparent decrease with aging and in AMD specimens suggests that its deficiency may accelerate the basolaminar deposit formation and RPE dysfunction seen in these conditions., The early stages of age-related macular degeneration (AMD) are characterized by the accumulation of basal laminar deposits (BLamDs). The mechanism for BLamDs accumulating between the retinal pigment epithelium (RPE) and its basal lamina remains elusive. Here we examined the role in AMD of lysosome-associated membrane protein-2 (LAMP2), a glycoprotein that plays a critical role in lysosomal biogenesis and maturation of autophagosomes/phagosomes. LAMP2 was preferentially expressed by RPE cells, and its expression declined with age. Deletion of the Lamp2 gene in mice resulted in age-dependent autofluorescence abnormalities of the fundus, thickening of Bruch’s membrane, and the formation of BLamDs, resembling histopathological changes occurring in AMD. Moreover, LAMP2-deficient mice developed molecular signatures similar to those found in human AMD—namely, the accumulation of APOE, APOA1, clusterin, and vitronectin—adjacent to BLamDs. In contrast, collagen 4, laminin, and fibronectin, which are extracellular matrix proteins constituting RPE basal lamina and Bruch’s membrane were reduced in Lamp2 knockout (KO) mice. Mechanistically, retarded phagocytic degradation of photoreceptor outer segments compromised lysosomal degradation and increased exocytosis in LAMP2-deficient RPE cells. The accumulation of BLamDs observed in LAMP2-deficient mice was eventually followed by loss of the RPE and photoreceptors. Finally, we observed loss of LAMP2 expression along with ultramicroscopic features of abnormal phagocytosis and exocytosis in eyes from AMD patients but not from control individuals. Taken together, these results indicate an important role for LAMP2 in RPE function in health and disease, suggesting that LAMP2 reduction may contribute to the formation of BLamDs in AMD.

PurposeTo investigate the progression characteristics of ellipsoid zone (EZ) loss in eyes with macular telangiectasia type 2 (MacTel) as reflected by area and linear measurements, and their relevance for visual acuity.MethodsParticipants were selected from the MacTel Study cohort. Linear and area measurements of EZ loss were performed in Spectral-Domain Optical Coherence Tomograph (SD-OCT) volume scans. Progression characteristics and correlations between linear and area measurements were analysed using linear mixed effects models.ResultsA total of 134 eyes of 70 patients were included (85 eyes with follow-up, mean 4.7years, range: 1.4-8years). Ellipsoid zone (EZ) loss significantly progressed at a mean annual increment of 0.057mm2 (p=0.005). The progression rate was non-linear and interacted significantly with initial EZ lesion size indicating an exponential growth before reaching a plateau. There was a strong heterogeneity in area sizes between fellow eyes. EZ break length had a significant linear effect on EZ break area (b=1.06, p&nbsp

Purpose To analyze imaging artifacts and segmentation errors with wide-field swept-source optical coherence tomography angiography (SS-OCTA) in diabetic retinopathy (DR). Methods We conducted a prospective, observational study at Massachusetts Eye and Ear from December 2018 to March 2019. Proliferative diabetic retinopathy (PDR), nonproliferative diabetic retinopathy (NPDR), diabetic patients with no diabetic retinopathy (DR), and healthy control eyes were included. All patients were imaged with a SS-OCTA and the Montage Angio (15 × 9 mm) was used for analysis. Images were independently evaluated by two graders using the motion artifact score (MAS). All statistical analyses were performed using SPSS 25.0 and R software. Results One hundred thirty-six eyes in 98 participants with the montage image were included in the study. Patients with more severe stages of DR had higher MAS by trend test analysis (P < 0.05). The occurrence of segmentation error was 0% in the healthy group, 10.53% in the no DR group, 10.00% in the NPDR group, and 50% in the PDR group. Multivariate regression analysis showed that the severity of DR and dry eye were the major factors affecting MAS (P < 0.05). There were some modifiable artifacts that could be corrected to improve image quality. Conclusions Wide field SS-OCTA assesses retinal microvascular changes by noninvasive techniques, yet distinguishing real alterations from artifacts is paramount to accurate interpretations. DR severity and dry eye correlated with MAS. Translational Relevance Understanding contributing factors and methods to reduce artifacts is critical to routine use and clinical trial with wide-field SS-OCTA.

The pathogenesis of age-related macular degeneration (AMD), a leading cause of blindness worldwide, remains only partially understood. This has led to the current lack of accessible and reliable biofluid biomarkers for diagnosis and prognosis, and absence of treatments for dry AMD. This study aimed to assess the plasma metabolomic profiles of AMD and its severity stages with the ultimate goal of contributing to addressing these needs. We recruited two cohorts: Boston, United States (n = 196) and Coimbra, Portugal (n = 295). Fasting blood samples were analyzed using ultra-high performance liquid chromatography mass spectrometry. For each cohort, we compared plasma metabolites of AMD patients versus controls (logistic regression), and across disease stages (permutation-based cumulative logistic regression considering both eyes). Meta-analyses were then used to combine results from the two cohorts. Our results revealed that 28 metabolites differed significantly between AMD patients versus controls (false discovery rate (FDR) q-value: 4.1 &times, 10&minus, 2&ndash, 1.8 &times, 5), and 67 across disease stages (FDR q-value: 4.5 &times, 1.7 &times, 4). Pathway analysis showed significant enrichment of glycerophospholipid, purine, taurine and hypotaurine, and nitrogen metabolism (p-value &le, 0.04). In conclusion, our findings support that AMD patients present distinct plasma metabolomic profiles, which vary with disease severity. This work contributes to the understanding of AMD pathophysiology, and can be the basis of future biomarkers and precision medicine for this blinding condition.

ObjectivesTo investigate the relationship between visual acuity (VA), total area of geographic atrophy (GA) and percentage of foveal GA.MethodsMulticenter, retrospective cross-sectional study of patients with GA due to age-related macular degeneration. Demographics, VA, fundus autofluorescence (FAF) and optical coherence tomography (OCT) images were collected. Using FAF images aided by OCT, foveal sparing status, GA pattern, total GA size, and percentage of GA covering the foveal area - area within a 1.5 mm diameter circle centered on the fovea centralis - were assessed. Univariable and multiple linear regression analyses were performed.Results54 eyes (mean age 78.7 ±7.7 (SD), 60.0% female) were studied. Mean VA was 0.8 ± 0.6 logMAR, mean total GA 8.8 ± 6.7 mm2 and mean percentage of foveal GA was 71.5 ± 30.9%. Of all assessed eyes, 48.2% (n = 26) presented with multifocal GA, and 18.5% (n = 10) had foveal sparing. Multiple regression analysis revealed that, controlling for age and GA pattern, the percentage of foveal GA presented a statistically significant association with VA (ß = 0.41, P = 0.004). No significant associations were observed with mean total GA size, while controlling for the same variables (ß = 0.010, P = 0.440).ConclusionPercentage of foveal GA was significantly associated with VA impairment, while the same was not verified for total GA area. These findings suggest that percentage of foveal GA may represent a more useful tool for assessing the impact of GA on VA. Further validation is needed in larger cohorts.

Age-related macular degeneration (AMD) is the leading cause of blindness in people over age 50 worldwide, and the third leading cause of blindness overall [...]

Objectives To assess resident cataract surgery outcomes at an academic teaching institution using 2 Physician Quality Reporting System (PQRS) cataract measures, which are intended to serve as a proxy for quality of surgical care. Design A retrospective review comparing cataract surgery outcomes of resident and attending surgeries using 2 PQRS measures: (1) 20/40 or better best-corrected visual acuity following cataract surgery and (2) complications within 30 days following cataract surgery requiring additional surgical procedures. Setting An academic ophthalmology center. Participants A total of 2487 surgeries performed at the Massachusetts Eye and Ear Infirmary from January 1, 2011 to December 31, 2012 were included in this study. Results Of all 2487 cataract surgeries, 98.95% achieved a vision of at least 20/40 at or before 90 days, and only 0.64% required a return to the operating room for postoperative complications. Of resident surgeries, 98.9% (1370 of 1385) achieved 20/40 vision at or before 90 days follow-up. Of attending surgeries, 99.0% (1091 of 1102) achieved 20/40 vision at or before 90 days (p = 1.00). There were no statistically significant differences between resident and attending cases regarding postoperative complications needing a return to the operating room (i.e., 0.65%, or 9 of 1385 resident cases vs 0.64%, or 7 of 1102 attending cases; p = 1.00). Conclusions Using PQRS Medicare cataract surgery criteria, this study establishes new benchmarks for cataract surgery outcomes at a teaching institution and supplemental measure for assessing resident surgical performance. Excellent cataract outcomes were achieved at an academic teaching institution, with results exceeding Medicare thresholds of 50%. There appears to be no significant difference in supervised trainee and attending cataract surgeon outcomes using 2 PQRS measures currently used by Medicare to determine physician reimbursement and quality of care.

Purpose: To study the clinical features of endogenous endophthalmitis (EE) in sample patient populations from the USA and South Korea over an 8-year period.Methods: We reviewed data from 128 eyes of 60 American and 48 Korean patients diagnosed with EE and compared their clinical characteristics.Results: Fungemia and liver abscess were the most common extraocular infection sources among American (26.7%) and Korean patients (33.3%), respectively. Klebsiella pneumoniae and Candida species were the most common pathogens of EE in the Korean and the American patients, respectively. Endophthalmitis caused by fungi had a better visual prognosis than that caused by bacteria (p = 0.001). Vitrectomy was beneficial for eyes with EE due to virulent bacteria presenting with worse than counting finger vision.Conclusions: The predisposing conditions and responsible organisms for EE vary in different regions of the world. The visual prognosis was strongly influenced by the underlying pathogen.

Anterior ischemic optic neuropathy (AION) is the most common cause of non-glaucomatous optic nerve head (ONH) injury among older adults. AION results from a sudden ischemic insult to the proximal portion of the optic nerve, typically leading to visual impairment. Here, we present an experimental model of photodynamically induced ONH injury that can be used to study neuroprotective modalities.Intraperitoneal injection of mesoporphyrin IX was followed by photodynamic treatment of the ONH in one eye of Brown-Norway rats; the fellow eye received the reverse sequence as a sham control. Fluorescein angiography (FA), spectral domain optical coherence tomography (SD-OCT), and visual evoked potential (VEP) recordings were performed at different time points following laser treatment. Immunohistochemistry was used to monitor apoptotic cell death (TUNEL) and macrophage infiltration (CD68). Cytokine levels were evaluated using enzyme-linked immunosorbent assay (ELISA).FA showed early hyperfluorescence and late leakage of the ONH, while SD-OCT revealed optic nerve edema. No leakage or other abnormalities were detected in control eyes. VEPs were significantly reduced in amplitude and showed prolonged responses compared to sham eyes. The number of apoptotic retinal ganglion cells was elevated one day after laser treatment (13.77 ± 4.49, p0.01) and peaked on day 7 (57.22 ± 11.34, p0.01). ONH macrophage infiltration also peaked on day 7 (101.8 ± 9.8, p0.05). ELISAs performed showed upregulation of macrophage chemoattractant protein-1 and macrophage inflammatory protein-2 on days 3 and 1, respectively.Photodynamic treatment of the ONH after administration of mesoporphyrin IX leads to macroscopic, histologic, and physiologic evidence of ONH injury. Given the long half-life of mesoporphyrin IX and the ease of intraperitoneal injections, this new model of photodynamically induced ONH injury may be a useful tool for studying optic nerve injury and possible neuroprotective treatments.

Multiple synthetic polymer nanoparticles (NPs) have been widely used as drug delivery systems. However, their toxicity to the retinal pigment epithelium and retinal endothelium remains unclear. In this study, we analyze the cytotoxic effects of three different kinds of NPs, made of poly lactic-co-glycolic acid (PLGA), polycaprolactone (PCL), and PEGylated PLGA (PEG-PLGA), in a retinal pigment epithelium cell line (ARPE-19) and in primary human retinal vascular endothelial cells (RVEC). PEG-PLGA NPs presented the lowest cytotoxicity on ARPE-19 cells and RVEC as assessed by MTT viability assay. While PLGA and PCL exhibited variable amounts of toxicity, no significant toxicity was observed when incubating cells with high PEG-PLGA concentrations (100 µg/ml), for up to 6 days. On both transmission electron microscopy and confocal microscopy, Rhodamine 6G-loaded PEG-PLGA NPs were observed intracellularly in multiple subcellular organelles. PEG-PLGA NPs are a potentially viable option for the treatment of eye diseases.

Purpose To review advances made in the treatment of age-related macular degeneration (AMD) and share perspectives on the future of AMD treatment. Methods Review of published clinical and experimental studies. Results Inhibitors of vascular endothelial growth factor (VEGF) truly revolutionized the treatment of AMD. However, available results from longer-term studies suggest that a degenerative process is unveiled, and continues to occur, even when neovascularization is controlled. Furthermore, anti-VEGF therapy may play a role in the development of atrophic changes. We have proposed using neuroprotection to prevent atrophy, and multiple models of retinal degeneration have shown that it is necessary to block both apoptotic and necrotic cell death pathways. Despite the success of anti-VEGF therapy and the promise of neuroprotection, neither addresses the underlying cause of AMD. It has been postulated that in early AMD, the retention and abnormal accumulation of lipids in Bruch's membrane and below the retinal pigmented epithelium (RPE) lead to drusen. Thus, it is conceivable to target the retained lipoproteins and seek to remove them. In a case study and pilot multicenter clinical trial, we observed significant regression of drusen and an improvement in visual acuity in patients taking high-dose statin therapy. These results, though preliminary, warrant further investigation. Conclusion Future treatment of AMD should be based on biology, which will require continued elucidation of the pathogenic mechanisms of AMD development. Neuroprotection represents a potential therapeutic approach, and other promising targets include immune pathways (e.g., inflammation, complement, and inflammasomes) and lipid/lipoprotein accumulation. Finally, due to the heterogeneity of AMD, future progress in therapy will benefit from improved phenotyping and classification.

To investigate the choroidal thickness (CT) and choroidal vascular densities (CVD) of patients with macular telangiectasia type 2 (MacTel2) and their association with other multimodal imaging features, using swept-source optical coherence tomography (SS-OCT).Prospective, cross-sectional study. Consecutive patients with MacTel2 along with controls without any macular disease were included. Fundus photography, confocal blue reflectance, near-infrared reflectance, autofluorescence, fluorescein angiography, spectral domain OCT and SS-OCT were performed. Images were independently analysed by two graders, and CVD was calculated from binarisedThirty-nine eyes of 20 patients with MacTel2 and 29 eyes of 15 control patients were included. Average CT and perifoveal temporal CT did not differ significantly between eyes with MacTel2 and control eyes (p≥0.350), when accounting for confounding factors. Overall and temporal CVD also did not significantly differ between the two groups (p≥0.490).CT and CVD did not significantly differ between MacTel2 and control eyes in this study using SS-OCT. Even though MacTel2 may include abnormalities involving the choroid, these are likely minor in comparison to the predominant retinal changes.

PurposeSodium iodate (NaIO3) has been extensively used as a retinotoxin to induce RPE cell damage and degeneration of photoreceptorsin vitroandin vivo. RIP-Kinase dependent programmed necrosis is an important redundant cell death pathway involved in photoreceptor cell death. We wanted to determine whether these pathways are actively involved in RPE and photoreceptor cell death after NaIO3 insult.MethodsARPE-19 cells were exposed to different concentrations of NaIO3 in the presence or absence of various concentrations of a RIPK inhibitor (Nec-1) or a pan-caspase inhibitor (Z-VAD), individually or combined. Cell death was determined at different time points by MTT (Sigma-Aldrich), LDH (Promega) and TUNEL (Millipore) assay. C57BL/6 and RIP3−/-mice were treated with a peritoneal injection of NaIO3 and eyes were enucleated at day 3 or 7. TUNEL staining was used to evaluate photoreceptor cell death. Photoreceptor cell loss was evaluated by measuring the thickness of outer nuclear layer (ONL). Microglia in the ONL were quantified in a retinal whole mount with Iba-1 antibody. RPE degeneration was also assessed in a RPE whole mount, with ZO-1 antibody.ResultsNaIO3 resulted in significant cell death of ARPE-19 cells. Treatment with Nec-1 resulted in better protection than treatment with Z-VAD (PIn vivoadministration of NaIO3 resulted in significant RPE and photoreceptor destruction with substantial inflammatory cell infiltration. RIP3 knockout animals displayed considerably less RPE and photoreceptor cell loss, as well as drastically less inflammation.ConclusionsProgrammed necrosis is an important cell death pathway mediating NaIO3 RPE and photoreceptor cell toxicity. Blocking the necroptosis pathway may serve as a novel therapeutic strategy for various RPE degenerative diseases.

N-methyl-D-aspartate (NMDA)-induced excitotoxicity is an acute form of experimental retinal injury as a result of overactivation of glutamate receptors. NLRP3 (nucleotide-binding domain, leucine-rich-repeat containing family, pyrin domain containing-3) inflammasome, one of the most studied sensors of innate immunity, has been reported to play a critical role in retinal neurodegeneration with controversial implications regarding neuroprotection and cell death. Thus far, it has not been elucidated whether NMDA-mediated excitotoxicity can trigger NLRP3 inflammasome in vivo. Moreover, it is unknown if NLRP3 is beneficial or detrimental to NMDA-mediated retinal cell death. Here, we employed a murine model of NMDA-induced retinal excitotoxicity by administering 100 nmoles of NMDA intravitreally, which resulted in massive TUNEL+ (TdT-dUTP terminal nick-end labelling) cell death in all retinal layers and especially in retinal ganglion cells (RGCs) 24 h post injection. NMDA insult in the retina potentiates macrophage/microglia cell infiltration, primes the NLRP3 inflammasome in a transcription-dependent manner and induces the expression of interleukin-1β (IL-1β). However, despite NLRP3 inflammasome upregulation, systemic deletion of Nlrp3 or Casp1 (caspase-1) did not significantly alter the NMDA-induced, excitotoxicity-mediated TUNEL+ retinal cell death at 24 h (acute phase). Similarly, the deletion of the two aforementioned genes did not alter the survival of the Brn3a+ (brain-specific homeobox/POU domain protein 3A) RGCs in a significant way at 3- or 7-days post injection (long-term phase). Our results indicate that NMDA-mediated retinal excitotoxicity induces immune cell recruitment and NLRP3 inflammasome activity even though inflammasome-mediated neuroinflammation is not a leading contributing factor to cell death in this type of retinal injury.

SUMMARYRetinal ganglion cell (RGC) death is the hallmark of glaucoma. Axonal injury is thought to precede RGC loss in glaucoma, and thus studies using an optic nerve (ON) crush model have been widely used to investigate mechanisms of cell death that are common to both conditions. Prior work has focused on the involvement of caspases in RGC death, but little is known about the contribution of other forms of cell death such as necrosis. In this study we show that receptor interacting protein (RIP) kinase-mediated necrosis normally plays a role in RGC death and acts in concert with caspase-dependent apoptosis. The expression of RIP3, a key activator of RIP1 kinase, as well as caspase activity, increased following ON injury. Caspase inhibition alone failed to provide substantial protection to injured RGCs and unexpectedly exacerbated necrosis. In contrast, pharmacologic or genetic inhibition of RIP kinases in combination with caspase blockade delayed both apoptotic and necrotic RGC death, although RGCs still continued to die. Furthermore, inhibition of RIP1 kinase promoted a moderate level of axon regeneration that was only minimal affected by caspase inhibition. In conclusion, multiple approaches are required for effective RGC death prevention and axonal regeneration. Further studies are needed to elucidate more effective long term strategies that can lead to sustained neuroprotection and regeneration.

Purpose: To evaluate the deeper choroidal vasculature in eyes with various ocular disorders using spectral domain (SD) optical coherence tomography angiography (OCTA) and swept source (SS) OCTA. Methods: Patients underwent OCTA imaging with either SD-OCTA (Zeiss Cirrus Angioplex or Optovue AngioVue) or SS-OCTA (Topcon Triton). Retinal pigment epithelium (RPE) integrity, structural visualization of deep choroidal vessels on en face imaging, and OCTA of deep choroidal blood flow signal were analyzed. Choroidal blood flow was deemed present if deeper choroidal vessels appeared bright after appropriate segmentation. Results: Structural visualization of choroidal vessels was feasible in all eyes by en face imaging. In both SD-OCTA and SS-OCTA, choroidal blood flow signal was present in all eyes with overlying RPE atrophy (100% of eyes with RPE atrophy, 28.6% of all imaged eyes, P < .001). Conclusions: While choroidal vessels can be visualized anatomically in all eyes by en face imaging, choroidal blood flow detection in deep choroidal vessel is largely restricted to areas with overlying RPE atrophy. Intact RPE acts as a barrier for reliable detection of choroidal flow using current OCTA technology, inhibiting evaluation of flow in deeper choroidal vessels in most eyes.

Retinal detachment (RD) is a sight-threatening complication common in many highly prevalent retinal disorders. RD rapidly leads to photoreceptor cell death beginning within 12 h following detachment. In patients with sustained RD, progressive visual decline due to photoreceptor cell death is common, leading to significant and permanent loss of vision. Microglia are the resident immune cells of the central nervous system, including the retina, and function in the homeostatic maintenance of the neuro-retinal microenvironment. It is known that microglia become activated and change their morphology in retinal diseases. However, the function of activated microglia in RD is incompletely understood, in part because of the lack of microglia-specific markers. Here, using the newly identified microglia marker P2ry12 and microglial depletion strategies, we demonstrate that retinal microglia are rapidly activated in response to RD and migrate into the injured area within 24 h post-RD, where they closely associate with infiltrating macrophages, a population distinct from microglia. Once in the injured photoreceptor layer, activated microglia can be observed to contain autofluorescence within their cell bodies, suggesting they function to phagocytose injured or dying photoreceptors. Depletion of retinal microglia results in increased disease severity and inhibition of macrophage infiltration, suggesting that microglia are involved in regulating neuroinflammation in the retina. Our work identifies that microglia mediate photoreceptor survival in RD and suggests that this effect may be due to microglial regulation of immune cells and photoreceptor phagocytosis.

Purpose Vision loss from age-related macular degeneration (AMD) has a profound effect on vision-related quality of life (VRQoL). The pupose of this study is to identify clinical factors associated with VRQoL using the Rasch- calibrated NEI VFQ-25 scales in bilateral advanced AMD patients. Methods We retrospectively reviewed 47 patients (mean age 83.2 years) with bilateral advanced AMD. Clinical assessment included age, gender, type of AMD, high contrast visual acuity (VA), history of medical conditions, contrast sensitivity (CS), central visual field loss, report of Charles Bonnet Syndrome, current treatment for AMD and Rasch-calibrated NEI VFQ-25 visual function and socioemotional function scales. The NEI VFQ visual function scale includes items of general vision, peripheral vision, distance vision and near vision-related activity while the socioemotional function scale includes items of vision related-social functioning, role difficulties, dependency, and mental health. Multiple regression analysis (structural regression model) was performed using fixed item parameters obtained from the one-parameter item response theory model. Results Multivariate analysis showed that high contrast VA and CS were two factors influencing VRQoL visual function scale (β = -0.25, 95% CI-0.37 to -0.12, p

OBJECTIVES To evaluate the impact of a comprehensive cataract surgery curriculum on the incidence of intraoperative complications. DESIGN We retrospectively compared the total number of cataract surgeries that the residents performed in all of the teaching sites, and the incidences of intraoperative complications (anterior capsule tear, posterior capsule rent, vitreous loss, anterior vitrectomy, zonular dialysis, iris trauma, hemorrhage, dropped lens fragment, corneal wound burn, incorrect intraocular lens) for the surgeries performed at Massachusetts Eye & Ear by residents in the pre-intervention group (residents graduating in 2004 and 2005), before the implementation of a surgical curriculum, and the residents in the post-intervention group (residents graduating in 2014 and 2015). SETTING Ophthalmology residency program at a major academic institution. PARTICIPANTS Residents graduating in 2004, 2005, 2014, and 2015. RESULTS We reviewed 4373 charts. 2086 of those surgeries were performed at Massachusetts Eye & Ear. The incidence of posterior capsule rent/vitreous loss/anterior vitrectomy was lower in the post-intervention group (1.4% versus 7.7%, p CONCLUSIONS Implementation of a comprehensive cataract surgery curriculum focusing on pre-operative, intra-operative and post-operative interventions, with an emphasis on patient outcomes resulted in a decrease in the rate of intraoperative complications.

AimsThe aim of our study was to image choroidal lesions with swept-source optical coherence tomography (SS-OCT) and to identify the morphological characteristics associated with optimal visualisation.MethodsThis was a prospective, cross-sectional study. Patients with choroidal melanocytic lesions ResultsWe included 85 choroidal lesions of 82 patients. There were 24 choroidal lesions (29%) for which image quality was classified as optimal, 31 lesions (37%) as suboptimal and 30 lesions (36%) as poor. The factors associated with optimal image quality were distance closer to the fovea (OR 0.76, pConclusionSS-OCT is useful in imaging most choroidal melanocytic lesions. Image quality is best when the choroidal lesion is closer to the fovea, has a smaller diameter and a lighter choroidal pigmentation.

Metabolomics is the qualitative and quantitative assessment of the metabolites (small molecules 1.5 kDa) in body fluids. The metabolites are the downstream of the genetic transcription and translation processes and also downstream of the interactions with environmental exposures; thus, they are thought to closely relate to the phenotype, especially for multifactorial diseases. In the last decade, metabolomics has been increasingly used to identify biomarkers in disease, and it is currently recognized as a very powerful tool with great potential for clinical translation. The metabolome and the associated pathways also help improve our understanding of the pathophysiology and mechanisms of disease. While there has been increasing interest and research in metabolomics of the eye, the application of metabolomics to retinal diseases has been limited, even though these are leading causes of blindness. In this manuscript, we perform a comprehensive summary of the tools and knowledge required to perform a metabolomics study, and we highlight essential statistical methods for rigorous study design and data analysis. We review available protocols, summarize the best approaches, and address the current unmet need for information on collection and processing of tissues and biofluids that can be used for metabolomics of retinal diseases. Additionally, we critically analyze recent work in this field, both in animal models and in human clinical disease, including diabetic retinopathy and age-related macular degeneration. Finally, we identify opportunities for future research applying metabolomics to improve our current assessment and understanding of mechanisms of vitreoretinal diseases, and to hence improve patient assessment and care.

Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.

Age related macular degeneration is the leading cause of blindness in the developed world. Although its precise cause remains elusive, dysfunction of the retinal pigment epithelium (RPE) and dysregulation of complement have been implicated in its pathogenesis. The goal of this study was to evaluate the role of an AMP-dependent kinase (AMPK) activator, 5-aminoimidazole-4-carboxamide riboside (AICAR), on tumor necrosis factor alpha (TNF-α) induction of complement factor B (CFB) in RPE cells. We found that AICAR inhibited TNF-α-induced CFB expression in ARPE-19 and human primary RPE cells in a dose-dependent fashion. Treatment of cells with dipyridamole, which blocks AICAR cellular uptake abolished these effects. In contrast, the adenosine kinase inhibitor, 5-iodotubericidin, which inhibits the conversion of AICAR to the direct activator of AMPK, ZMP, did not reverse the effects on TNF-α-induced CFB expression, suggesting AMPK-independent effects. Indeed, knockout of AMPK in RPE cells using Clustered Regularly Interspaced Palindromic Repeats (CRISPR)/Cas9 did not abolish the inhibitory effects of AICAR on RPE CFB expression. Collectively, our results suggest that AICAR can suppress TNF-α-induced CFB expression in RPE cells in an AMPK-independent mechanism, and could be used as a therapeutic target in certain complement over-activation scenarios.

Age-related macular degeneration (AMD) is the leading cause of irreversible blindness in the elderly of industrialized nations, and there is increasing evidence to support a role for chronic inflammation in its pathogenesis. Mitochondrial DNA (mtDNA) has been recently reported to be pro-inflammatory in various diseases such as Alzheimer’s and heart failure. Here, we report that intracellular mtDNA induces ARPE-19 cells to secrete IL-6 and IL-8, inflammatory cytokines that have consistently been associated with AMD onset and progression. The induction was dependent on the size of mtDNA, but not on specific sequence. Oxidative stress plays a major role in the development of AMD, and our findings indicate that mtDNA induces IL-6 and IL-8 more potently when oxidized. Cytokine induction was mediated by STING (Stimulator of Interferon Genes) and NF-κB as evidenced by abrogation of the cytokine response with use of specific inhibitors (siRNA and Bay 11–7082, respectively). Finally, mtDNA primed the NLRP3 inflammasome and weakly activated it as shown by caspase-1 activation and mature IL-1β secretion. This study contributes to our understanding of the potential pro-inflammatory role of mtDNA in the pathogenesis of AMD.

Age-related macular degeneration (AMD) is the most common cause of irreversible visual impairment in older populations in industrialized nations. AMD is a late-onset deterioration of photoreceptors and retinal pigment epithelium in the central retina caused by various environmental and genetic factors. Great strides in our understanding of AMD pathogenesis have been made in the past several decades, which have translated into revolutionary therapeutic agents in recent years. In this review, we describe the clinical and pathologic features of AMD and present an overview of current diagnosis and treatment strategies.

Patrick Oellers,1,* Inês Laíns,1,2,* Steven Mach,1 Shady Garas,1 Ivana K Kim,1 Demetrios G Vavvas,1 Joan W Miller,1 Deeba Husain,1 John B Miller1 1Retina Service, Department of Ophthalmology, Massachusetts Eye and Ear, Harvard Medical School, Boston, MA, USA; 2Faculty of Medicine, University of Coimbra, Coimbra, Portugal *These authors contributed equally to this work Purpose: Eyes with age-related macular degeneration (AMD) often harbor pathological changes in the retinal periphery and perimacular region. These extramacular changes have not been well classified, but may be phenotypically and functionally relevant. The purpose of this study was to demonstrate a novel grid to systematically study peripheral retinal abnormalities in AMD using geometric distortion-corrected ultra-widefield (UWF) imaging.Methods: This is a cross-sectional observational case series. Consecutive patients with AMD without any other coexisting vitreoretinal disease and control patients over age 50 without AMD or any other vitreoretinal disease were imaged using Optos 200 Tx. Captured 200° UWF images were corrected for peripheral geometric distortion using Optos transformation software. A newly developed grid to study perimacular and peripheral abnormalities in AMD was then projected onto the images.Results: Peripheral and perimacular changes such as drusen, retinal pigment epithelium changes and atrophy were found in patients with AMD. The presented grid in conjunction with geometric distortion-corrected UWF images allowed for systematic study of these peripheral changes in AMD.Conclusion: We present a novel grid to study peripheral and posterior pole changes in AMD. The grid is unique in that it adds a perimacular zone, which may be important in characterizing certain phenotypes in AMD. Our UWF images were corrected for geometric peripheral distortion to accurately reflect the anatomical dimensions of the retina. This grid offers a reliable and reproducible foundation for the exploration of peripheral retinal pathology associated with AMD. Keywords: ultra-widefield, autofluorescence, macular degeneration, grid, periphery, drusen, retinal pigment epithelium changes

While the development of anti-vascular endothelial growth factor (anti-VEGF) as a therapy for neovascular age-related macular degeneration (AMD) was a great success, the pathologic processes underlying dry AMD that eventually leads to photoreceptor dysfunction, death, and vision loss remain elusive to date, with a lack of effective therapies and increasing prevalence of the disease. There is an overwhelming need to improve the classification system of AMD, to increase our understanding of cell death mechanisms involved in both neovascular and non-neovascular AMD, and to develop better biomarkers and clinical endpoints to eventually be able to identify better therapeutic targets—especially early in the disease process. There is no doubt that it is a matter of time before progress will be made and better therapies will be developed for non-neovascular AMD.

Age-related macular degeneration (AMD) is the most common cause of blindness for individuals age 50 and above in the developed world. Abnormal growth of choroidal blood vessels, or choroidal neovascularization (CNV), is a hallmark of the neovascular (wet) form of advanced AMD and leads to significant vision loss. A growing body of evidence supports a strong link between neovascular disease and inflammation. Metabolites of long-chain polyunsaturated fatty acids derived from the cytochrome P450 (CYP) monooxygenase pathway serve as vital second messengers that regulate a number of hormones and growth factors involved in inflammation and vascular function. Using transgenic mice with altered CYP lipid biosynthetic pathways in a mouse model of laser-induced CNV, we characterized the role of these lipid metabolites in regulating neovascular disease. We discovered that the CYP-derived lipid metabolites epoxydocosapentaenoic acids (EDPs) and epoxyeicosatetraenoic acids (EEQs) are vital in dampening CNV severity. Specifically, overexpression of the monooxygenase CYP2C8 or genetic ablation or inhibition of the soluble epoxide hydrolase (sEH) enzyme led to increased levels of EDP and EEQ with attenuated CNV development. In contrast, when we promoted the degradation of these CYP-derived metabolites by transgenic overexpression of sEH, the protective effect against CNV was lost. We found that these molecules work in part through their ability to regulate the expression of key leukocyte adhesion molecules, on both leukocytes and endothelial cells, thereby mediating leukocyte recruitment. These results suggest that CYP lipid signaling molecules and their regulators are potential therapeutic targets in neovascular diseases.

Purpose To compare choroidal vascular density (CVD) and volume (CVV) in diabetic eyes and controls, using en face swept-source optical coherence tomography (SS-OCT). Design Prospective cross-sectional study. Methods Setting: Multicenter. Patient Population: Total of 143 diabetic eyes—27 with no diabetic retinopathy (DR), 47 with nonproliferative DR (NPDR), 51 with NPDR and diabetic macular edema (DME), and 18 with proliferative DR (PDR)—and 64 age-matched nondiabetic control eyes. Observation Procedures: Complete ophthalmologic examination and SS-OCT imaging. En face SS-OCT images of the choroidal vasculature were binarized. Main Outcome Measures: CVD, calculated as the percent area occupied by choroidal vessels in the central macular region (6-mm-diameter circle centered on the fovea), and throughout the posterior pole (12 × 9 mm). The central macular CVV was calculated by multiplying the average CVD by macular area and choroidal thickness (obtained with SS-OCT automated software). Multilevel mixed linear models were performed for analyses. Results Compared to controls (0.31 ± 0.07), central macular CVD was significantly decreased by 9% in eyes with NPDR + DME (0.28 ± 0.06; s = −0.03, P = .02) and by 15% in PDR (0.26 ± 0.05; s = −0.04, P = .01). The central macular CVV was significantly decreased by 19% in eyes with PDR (0.020 ± 0.005 mm 3 , s = −0.01, P = .01) compared to controls (0.025 ± 0.01 mm 3 ). Conclusions Choroidal vascular density and volume are significantly reduced in more advanced stages of diabetic retinopathy. New imaging modalities should allow further exploration of the contributions of choroidal vessel disease to diabetic eye disease pathogenesis, prognosis, and treatment response.

Phagocytosis of daily shed photoreceptor outer segments is an important function of the retinal pigment epithelium (RPE) and it is essential for retinal homeostasis. RPE dysfunction, especially impairment of its phagocytic ability, plays an essential role in the pathogenesis of age-related macular degeneration (AMD). Statins, or HMG CoA (3-hydroxy-3-methylglutaryl-coenzyme A) reductase inhibitors, are drugs with multiple properties that have been extensively used to treat hyperlipidemia. However, their effect on RPE cells has not been fully elucidated. Here we report that high dose atorvastatin increased the phagocytic function of ARPE-19 cells, as well as rescue the cells from the phagocytic dysfunction induced by cholesterol crystals and oxidized low-density lipoproteins (ox-LDL), potentially by increasing the cellular membrane fluidity. Similar effects were observed when evaluating two other hydrophobic statins, lovastatin and simvastatin. Furthermore, atorvastatin was able to block the induction of interleukins IL-6 and IL-8 triggered by pathologic stimuli relevant to AMD, such as cholesterol crystals and ox-LDL. Our study shows that statins, a well-tolerated class of drugs with rare serious adverse effects, help preserve the phagocytic function of the RPE while also exhibiting anti-inflammatory properties. Both characteristics make statins a potential effective medication for the prevention and treatment of AMD.

The National Eye Institute (NEI) launched the Audacious Goals Initiative (AGI) in 2013 with the aim “to restore vision through the regeneration of neurons and neural connections in the eye and visual system.” An AGI Town Hall held at the Association for Research in Vision and Ophthalmology Annual Meeting in 2016 brought together basic, translational, and clinical scientists to address the clinical implications of the AGI, with a particular emphasis on diseases amenable to regenerative medicine and strategies to deal with barriers to progess. An example of such a barrier is that replacement of lost neurons may be insufficient because damage to other neurons and non-neuronal cells is common in retinal and optic nerve disease. Reparative processes such as gliosis and fibrosis also can make it difficult to replenish and regenerate neurons. Other issues include choice of animal models, selecting appropriate endpoints, ethics of informed consent, and regulatory issues, Another area critical to next steps in the AGI is the choice of target diseases and the stage at which early development studies should be focused. For example, an advantage of doing clinical trials in patients with early disease is that supporting cellular and structural constituents are still likely to be present. On the other hand, regenerative studies in patients with late disease make it easier to detect the effects of replacement therapy against the background of severe visual loss, while it may be harder to detect incremental improvement in visual function in those with early disease and considerable remaining visual function. Achieving the goals of the AGI also requires preclinical advances, new imaging techniques, and optimizing translational issues. The work of the AGI is expected to take at least ten years, but should eventually result in therapies to restore some degree of vision to the blind.

Verteporfin (VP) was first used in Photodynamic therapy, where a non-thermal laser light (689 nm) in the presence of oxygen activates the drug to produce highly reactive oxygen radicals, resulting in local cell and tissue damage. However, it has also been shown that Verteporfin can have non-photoactivated effects such as interference with the YAP-TEAD complex of the HIPPO pathway, resulting in growth inhibition of several neoplasias. More recently, it was proposed that, another non-light mediated effect of VP is the formation of cross-linked oligomers and high molecular weight protein complexes (HMWC) that are hypothesized to interfere with autophagy and cell growth. Here, in a series of experiments, using human uveal melanoma cells (MEL 270), human embryonic kidney cells (HEK) and breast cancer cells (MCF7) we showed that Verteporfin-induced HMWC require the presence of light. Furthermore, we showed that the mechanism of this cross-linking, which involves both singlet oxygen and radical generation, can occur very efficiently even after lysis of the cells, if the lysate is not protected from ambient light. This work offers a better understanding regarding VP’s mechanisms of action and suggests caution when one studies the non-light mediated actions of this drug.

Verteporfin (VP), a benzoporphyrin derivative, is clinically used in photodynamic therapy for neovascular macular degeneration. Recent studies indicate that VP may inhibit growth of hepatoma cells without photoactivation through inhibition of YAP-TEAD complex. In this study, we examined the effects of VP without light activation on human retinoblastoma cell lines. Verteporfin but not vehicle control inhibited the growth, proliferation and viability of human retinoblastoma cell lines (Y79 and WERI) in a dose-dependent manner and was associated with downregulation of YAP-TEAD associated downstream proto-oncogenes such as c-myc, Axl, and surviving. In addition VP affected signals involved in cell migration and angiogenesis such as CTGF, cyr61, and VEGF-A but was not associated with significant effect on the mTOR/autophagy pathway. Of interest the pluripotency marker Oct4 were downregulated by Verteporfin treatment. Our results indicate that the clinically used photosensitizer VP is a potent inhibitor of cell growth in retinoblastoma cells, disrupting YAP-TEAD signaling and pluripotential marker OCT4. This study highlights for the first time the role of the YAP-TEAD pathway in Retinoblastoma and suggests that VP may be a useful adjuvant therapeutic tool in treating Rb patients.

Ocular neovascularization, including age-related macular degeneration (AMD), is a primary cause of blindness in individuals of industrialized countries. With a projected increase in the prevalence of these blinding neovascular diseases, there is an urgent need for new pharmacological interventions for their treatment or prevention. Increasing evidence has implicated eicosanoid-like metabolites of long-chain polyunsaturated fatty acids (LCPUFAs) in the regulation of neovascular disease. In particular, metabolites generated by the cytochrome P450 (CYP)-epoxygenase pathway have been shown to be potent modulators of angiogenesis, making this pathway a reasonable previously unidentified target for intervention in neovascular ocular disease. Here we show that dietary supplementation with ω-3 LCPUFAs promotes regression of choroidal neovessels in a well-characterized mouse model of neovascular AMD. Leukocyte recruitment and adhesion molecule expression in choroidal neovascular lesions were down-regulated in mice fed ω-3 LCPUFAs. The serum of these mice showed increased levels of anti-inflammatory eicosanoids derived from eicosapentaenoic acid and docosahexaenoic acid. 17,18-epoxyeicosatetraenoic acid and 19,20-epoxydocosapentaenoic acid, the major CYP-generated metabolites of these primary ω-3 LCPUFAs, were identified as key lipid mediators of disease resolution. We conclude that CYP-derived bioactive lipid metabolites from ω-3 LCPUFAs are potent inhibitors of intraocular neovascular disease and show promising therapeutic potential for resolution of neovascular AMD.

Aims To evaluate safety and visual outcomes after proton beam irradiation (PBI) therapy for subfoveal choroidal neovascularisation (CNV) secondary to causes other than age-related macular degeneration (AMD). Methods This study is a prospective, unmasked and randomised clinical trial using two dosage regimens, conducted in the Massachusetts Eye and Ear Infirmary. The study included 46 patients with CNV secondary to non-AMD and best-corrected visual acuity of 20/320 or better. Patients were randomly assigned to receive 16 or 24 cobalt gray equivalents (CGE) of PBI in two equal fractions. Complete ophthalmological examinations, fundus photography and fluorescein angiography were performed at baseline and 6, 12, 18 and 24 months after treatment. Results At 1 year after treatment, 82% and 72% lost fewer than 1.5 lines of vision in the 16 CGE and in 24 CGE groups, respectively. At 2 years after therapy, 77% in the lower dose group and 64% in the higher dose group lost fewer than 1.5 lines of vision. Mild radiation complications such as radiation vasculopathy developed in 17.6% of patients. Conclusions PBI is a safe and efficacious treatment for subfoveal CNV not due to AMD. The data with respect to visual outcomes and radiation complications trend in favour of the 16 CGE group, although differences do not reach statistical significance. PBI may be considered as an alternative to current therapies.

I shall discuss the work of researchers at Harvard Medical School who came together in the early 1990s. Scattered across various Harvard-affiliated hospitals and research centers, these individuals were unified by their interest in ocular neovascularization. Together and separately, they investigated models of ocular neovascularization, exploring tumor angiogenesis in eye development and disease.

To examine the relationship between dark adaptation (DA) and optical coherence tomography (OCT)-based macular morphology in age-related macular degeneration (AMD).Prospective, cross-sectional study.Patients with AMD and a comparison group (50 years) without any vitreoretinal disease.All participants were imaged with spectral-domain OCT and color fundus photographs, and then staged for AMD (Age-related Eye Disease Study system). Both eyes were tested with the AdaptDx (MacuLogix, Middletown, PA) DA extended protocol (20 minutes). A software program was developed to map the DA testing spot (2° circle, 5° superior to the fovea) to the OCT B-scans. Two independent graders evaluated the B-scans within this testing spot, as well as the entire macula, recording the presence of several AMD-associated abnormalities. Multilevel mixed-effects models (accounting for correlated outcomes between 2 eyes) were used for analyses.The primary outcome was rod-intercept time (RIT), defined in minutes, as a continuous variable. For subjects unable to reach RIT within the 20 minutes of testing, the value of 20 was assigned.We included 137 eyes (n = 77 subjects), 72.3% (n = 99 eyes) with AMD and the remainder belonging to the comparison group. Multivariable analysis revealed that even after adjusting for age and AMD stage, the presence of any abnormalities within the DA testing spot (ß = 4.8, P0.001), as well as any abnormalities in the macula (ß = 2.4, P = 0.047), were significantly associated with delayed RITs and therefore impaired DA. In eyes with no structural changes within the DA testing spot (n = 76, 55.5%), the presence of any abnormalities in the remaining macula was still associated with delayed RITs (ß = 2.00, P = 0.046). Presence of subretinal drusenoid deposits and ellipsoid zone disruption were a consistent predictor of RIT, whether located within the DA testing spot (P = 0.001 for both) or anywhere in the macula (P0.001 for both). Within the testing spot, the presence of classic drusen or serous pigment epithelium detachment was also significantly associated with impairments in DA (P ≤ 0.018).Our results suggest a significant association between macular morphology evaluated by OCT and time to dark-adapt. Subretinal drusenoid deposits and ellipsoid zone changes seem to be strongly associated with impaired dark adaptation.

This study evaluated the safety and tolerability of ocular RS1 adeno-associated virus (AAV8-RS1) gene augmentation therapy to the retina of participants with X-linked retinoschisis (XLRS). XLRS is a monogenic trait affecting only males, caused by mutations in the RS1 gene. Retinoschisin protein is secreted principally in the outer retina, and its absence results in retinal cavities, synaptic dysfunction, reduced visual acuity, and susceptibility to retinal detachment. This phase I/IIa single-center, prospective, open-label, three-dose-escalation clinical trial administered vector to nine participants with pathogenic RS1 mutations. The eye of each participant with worse acuity (≤63 letters; Snellen 20/63) received the AAV8-RS1 gene vector by intravitreal injection. Three participants were assigned to each of three dosage groups: 1e9 vector genomes (vg)/eye, 1e10 vg/eye, and 1e11 vg/eye. The investigational product was generally well tolerated in all but one individual. Ocular events included dose-related inflammation that resolved with topical and oral corticosteroids. Systemic antibodies against AAV8 increased in a dose-related fashion, but no antibodies against RS1 were observed. Retinal cavities closed transiently in one participant. Additional doses and immunosuppressive regimens are being explored to pursue evidence of safety and efficacy (ClinicalTrials.gov: NCT02317887).

The molecular mechanisms involved in induced pluripotent stem cells (iPSCs) generation are poorly understood. The cell death machinery of apoptosis-inducing caspases have been shown to facilitate the process of iPSCs reprogramming. However, the effect of other cell death processes, such as programmed necrosis (necroptosis), on iPSCs induction has not been studied. In this study, we investigated the role of receptor-interacting protein kinase 3 (RIP3), an essential regulator of necroptosis, in reprogramming mouse embryonic fibroblast cells (MEFs) into iPSCs. RIP3 was found to be upregulated in iPSCs compared to MEFs. Deletion of RIP3 dramatically suppressed the reprogramming of iPSCs (~82%). RNA-seq analysis and qRT-PCR showed that RIP3 KO MEFs expressed lower levels of genes that control cell cycle progression and cell division and higher levels of extracellular matrix-regulating genes. The growth rate of RIP3 KO MEFs was significantly slower than WT MEFs. These findings can partially explain the inhibitory effects of RIP3 deletion on iPSCs generation and show for the first time that the necroptosis kinase RIP3 plays an important role in iPSC reprogramming. In contrast to RIP3, the kinase and scaffolding functions of RIPK1 appeared to have distinct effects on reprogramming. Keywords: Reprogramming, Programmed necrosis, Necroptosis, Cell death, RIPK3, RIPK, RIP

Background/aimsMicroperimetry is a technique that is increasingly used to assess visual function in age-related macular degeneration (AMD). In this study, we aimed to evaluate the relationship between retinal sensitivity measured with macular integrity assessment (MAIA) microperimetry and optical coherence tomography (OCT)-based macular morphology in AMD.MethodsProspective, cross-sectional study. All participants were imaged with colour fundus photographs used for AMD staging (Age-Related Eye Disease Study scale), spectral-domain OCT (Spectralis, Heidelberg, Germany) and swept-source OCT (Topcon, Japan). Threshold retinal sensitivity of the central 10° diameter circle was assessed with the full-threshold, 37-point protocol of the MAIA microperimetry device (Centervue, Italy). Univariable and multivariable multilevel mixed-effect linear regression models were used for analysis.ResultsWe included 102 eyes with AMD and 46 control eyes. Multivariable analysis revealed that older age (pConclusionsThere is an association between TRS as determined by MAIA microperimetry and several OCT structural parameters across various stages of AMD. This study highlights the relevance of microperimetry as a functional outcome measure for AMD.

ObjectivesOcular hypertension is a primary risk factor for glaucoma and results in retinal ganglion cell (RGC) degeneration. Current animal models of glaucoma lack severe RGC cell death as seen in glaucoma, making assessment of physiological mediators of cell death difficult. We developed a modified mouse model of ocular hypertension whereby long-lasting elevation of intraocular pressure (IOP) is achieved, resulting in significant reproducible damage to RGCs.ResultsIn this model, microbeads are mixed with hyaluronic acid and injected into the anterior chamber of C57BL/6J mice. The hyaluronic acid allows for a gradual release of microbeads, resulting in sustained blockage of Schlemm's canal. IOP elevation was bimodal during the course of the model's progression. The first peak occurred 1 hours after beads injection, with an IOP value of 44.69 ± 6.00 mmHg, and the second peak occurred 6-12 days post-induction, with an IOP value of 34.91 ± 5.21 mmHg. RGC damage was most severe in the peripheral retina, with a loss of 64.1% compared to that of untreated eyes, while the midperiphery exhibited a 32.4% loss, 4 weeks following disease induction.ConclusionsThese results suggest that sustained IOP elevation causes more RGC damage in the periphery than in the midperiphery of the retina. This model yields significant and reproducible RGC degeneration.

Purpose To compare the visualization of the choriocapillaris and deeper choroidal vessels in healthy eyes in en face swept-source optical coherence tomography (SS-OCT) versus SS-OCT angiography (SS-OCTA). Methods This is a cross-sectional study of consecutive eyes without chorioretinal disease. En face SS-OCT and SS-OCTA images of the choriocapillaris and choroid were assessed for visualization of the vasculature. Choroidal vessel densities (CVD) of the choriocapillaris, inner choroid, midchoroid, and outer choroid were calculated from binarized en face SS-OCT and SS-OCTA images. Paired t-tests and linear regression were used for statistical analysis. Results Twenty-seven eyes of 27 patients were included. There was no statistically significant difference between the CVDs of the midchoroid assessed with en face SS-OCT versus SS-OCTA (P = 0.21). However, there were statistically significant differences between the CVDs for the choriocapillaris (P < 0.001), inner choroid (P < 0.001), and outer choroid (P = 0.006). Qualitative analysis revealed incomplete visualization of vessels in the inner choroid and exaggeration of vessel lumens in the outer choroid with SS-OCTA. Conclusions Visualization of the choriocapillaris is superior with SS-OCTA, but it has numerous limitations that make visualization of deeper choroidal vessels less reliable when compared with en face SS-OCT. It is important to understand such limitations when using these technologies to study the choroidal vasculature in chorioretinal disease. Translational Relevance The presented study of optimal imaging techniques for the choroidal vessels of healthy eyes provides an important foundation for future investigations into the role of the choroidal vasculature in chorioretinal diseases.