Pericyte dysfunction and loss of interpericyte tunneling nanotubes promote neurovascular deficits in glaucoma.

Reduced blood flow and impaired neurovascular coupling are recognized features of glaucoma, the leading cause of irreversible blindness worldwide, but the mechanisms underlying these defects are unknown. Retinal pericytes regulate microcirculatory blood flow and coordinate neurovascular coupling through interpericyte tunneling nanotubes (IP-TNTs). Using two-photon microscope live imaging of the mouse retina, we found reduced capillary diameter and impaired blood flow at pericyte locations in eyes with high intraocular pressure, the most important risk factor to develop glaucoma. We show that IP-TNTs are structurally and functionally damaged by ocular hypertension, a response that disrupted light-evoked neurovascular coupling. Pericyte-specific inhibition of excessive Ca2+ influx rescued hemodynamic responses, protected IP-TNTs and neurovascular coupling, and enhanced retinal neuronal function as well as survival in glaucomatous retinas. Our study identifies pericytes and IP-TNTs as potential therapeutic targets to counter ocular pressure-related microvascular deficits, and provides preclinical proof of concept that strategies aimed to restore intrapericyte calcium homeostasis rescue autoregulatory blood flow and prevent neuronal dysfunction.

Genetic Risk Stratification of Primary Open-Angle Glaucoma in Japanese Individuals.

PURPOSE: To assess the impact of genetic risk estimation for primary open-angle glaucoma (POAG) in Japanese individuals. DESIGN: Cross-sectional analysis. PARTICIPANTS: Genetic risk scores (GRSs) were constructed based on a genome-wide association study (GWAS) of POAG in Japanese people. A total of 3625 Japanese individuals, including 1191 patients and 2434 controls (Japanese Tohoku), were used for the model selection. We also evaluated the discriminative accuracy of constructed GRSs in a dataset comprising 1034 patients and 1147 controls (the Japan Glaucoma Society Omics Group [JGS-OG] and the Genomic Research Committee of the Japanese Ophthalmological Society [GRC-JOS]) and 1900 participants from a population-based study (Hisayama Study). METHODS: We evaluated 2 types of GRSs: polygenic risk scores using the pruning and thresholding procedure and a GRS using variants associated with POAG in the GWAS of the International Glaucoma Genetics Consortium (IGGC). We selected the model with the highest areas under the receiver operating characteristic curve (AUC). In the population-based study, we evaluated the correlations between GRS and ocular measurements. MAIN OUTCOME MEASURE: Proportion of patients with POAG after stratification according to the GRS. RESULTS: We found that a GRS using 98 variants, which showed genome-wide significance in the IGGC, showed the best discriminative accuracy (AUC, 0.65). In the Japanese Tohoku, the proportion of patients with POAG in the top 10% individuals was significantly higher than that in the lowest 10% (odds ratio [OR], 6.15; 95% confidence interval [CI], 4.35-8.71). In the JGS-OG and GRC-JOS, we confirmed similar impact of POAG GRS (AUC, 0.64; OR [top vs. bottom decile], 5.81; 95% CI, 3.79-9.01). In the population-based study, POAG prevalence was significantly higher in the top 20% individuals of the GRS compared with the bottom 20% (9.2% vs. 5.0%). However, the discriminative accuracy was low (AUC, 0.56). The POAG GRS was correlated positively with intraocular pressure (r = 0.08: P = 4.0 × 10-4) and vertical cup-to-disc ratio (r = 0.11; P = 4.0 × 10-6). CONCLUSIONS: The GRS showed moderate discriminative accuracy for POAG in the Japanese population. However, risk stratification in the general population showed relatively weak discriminative performance. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Vasoreactivity of the optic nerve head, nailfold, and facial skin in response to cold provocation in normal-tension glaucoma patients.

BACKGROUND: The dysfunction of optic nerve head (ONH) hemodynamics has been suggested to be involved in the pathogenesis of normal-tension glaucoma (NTG). The aim of this study was to compare vasoreactivity in the ONH, nailfold, and facial skin in response to cold-water provocation in NTG patients and healthy controls. METHODS: We performed cold-water provocation in 14 eyes of 14 NTG patients and 15 eyes of 15 age-matched control subjects. Laser speckle flowgraphy-derived tissue-area mean blur rate (MT), skin blood flowmetry-derived pulse wave amplitude (PA), nailfold capillaroscopy-derived nailfold capillary diameter, and other clinical parameters were recorded at baseline and 4 and 6 min after the cold stimulus. We compared changes (as percentages) in these variables in the NTG and control subjects with a linear mixed-effects model and evaluated correlations between these changes with Spearman's rank correlation coefficient. RESULTS: The interaction term between the NTG group (reference, control group) and the 4-min protocol step (reference, baseline) significantly affected the changes in MT, nailfold capillary diameter and PA (ß = -9.51%, P = 0.017, ß = -20.32%, P = 0.002; ß = + 18.06%, P = 0.017, respectively). The change in MT was positively correlated with the change in nailfold capillary diameter, and negatively correlated with the change in PA (r = 0.39, P = 0.036; r = -0.40, P = 0.031, respectively). CONCLUSION: NTG patients showed abnormal vasoconstriction in the ONH and nailfold and vasodilation in the facial skin in response to cold-water provocation.

Time-Course Changes in Optic Nerve Head Blood Flow and Retinal Nerve Fiber Layer Thickness in Eyes with Open-angle Glaucoma.

PURPOSE: To determine whether decreased optic nerve head (ONH) blood flow (BF) precedes or follows decreased circumpapillary retinal nerve fiber layer thickness (cpRNFLT) in eyes with open-angle glaucoma (OAG). DESIGN: Retrospective, longitudinal study. PARTICIPANTS: This study followed up 350 eyes of 225 OAG patients for at least 2 years and collected data from each patient from at least 5 examinations obtained with laser speckle flowgraphy (LSFG) and OCT. METHODS: In the superior, temporal, and inferior ONH quadrants, tissue area mean blur rate (MT), representing ONH tissue BF, was measured with LSFG, whereas cpRNFLT was measured with OCT. A multivariate linear mixed-effects model was used to identify potential predictors of faster MT decrease, adjusting for possible confounding factors. Based on these results, each quadrant of each patient was assigned a risk point if the quadrant was the superior or temporal, if patient age was older than the median (61 years), and if patient pulse rate was higher than median (74 beats per minute). The quadrants were then compared with a mixed-effects Cox model for MT and cpRNFLT changes, defined as a difference between the baseline value and the values from the latest 2 consecutive follow-up visits of more than 1.96 × the corresponding coefficient of variation. MAIN OUTCOME MEASURES: Ophthalmic and systemic variables and MT and cpRNFLT in the superior, temporal, and inferior quadrants. RESULTS: The multivariate model showed that MT decrease was faster in older patients with higher pulse rate and slower in inferior quadrants (P < 0.05). Quadrants with 0 risk points showed primary cpRNFLT decrease (P = 0.048), 1-risk point quadrants showed simultaneous cpRNFLT and MT decrease (P = 0.260), and 2-risk point and 3-risk point quadrants showed primary MT decrease (P < 0.001). CONCLUSIONS: Older patients with higher pulse rate are at greater risk of a primary reduction in ONH tissue BF, that is, preceding cpRNFLT decrease, in the superior and temporal quadrants.

CHOP deletion and anti-neuroinflammation treatment with hesperidin synergistically attenuate NMDA retinal injury in mice.

Glaucoma is a leading cause of blindness worldwide and is characterized by degeneration associated with the death of retinal ganglion cells (RGCs). It is believed that glaucoma is a group of heterogeneous diseases with multifactorial pathomechanisms. Here, we investigate whether anti-inflammation treatment with an ER stress blockade can selectively promote neuroprotection against NMDA injury in the RGCs. Retinal excitotoxicity was induced with an intravitreal NMDA injection. Microglial activation and neuroinflammation were evaluated with Iba1 immunostaining and cytokine gene expression. A stable HT22 cell line transfected with an NF-kB reporter was used to assess NF-kB activity after hesperidin treatment. CHOP-deficient mice were used as a model of ER stress blockade. Retinal cell death was evaluated with a TUNEL assay. As results, in the NMDA injury group, Iba1-positive microglia increased 6 h after NMDA injection. Also at 6 h, pro-inflammatory cytokines and chemokine increased, including TNFα, IL-1b, IL-6 and MCP-1. In addition, the MCP-1 promoter-driven EGFP signal, which we previously identified as a stress signal in injured RGCs, also increased; hesperidin treatment suppressed this inflammatory response and reduced stressed RGCs. In CHOP-deficient mice that received an NMDA injection, the gene expression of pro-inflammatory cytokines, chemokines, markers of active microglia, and inflammatory regulators was greater than in WT mice. In WT mice, hesperidin treatment partially prevented retinal cell death after NMDA injury; this neuroprotective effect was enhanced in CHOP-deficient mice. These findings demonstrate that ER stress blockade is not enough by itself to prevent RGC loss due to neuroinflammation in the retina, but it has a synergistic neuroprotective effect after NMDA injury when combined with an anti-inflammatory treatment based on hesperidin.

Genome-wide association study identifies seven novel susceptibility loci for primary open-angle glaucoma.

Primary open-angle glaucoma (POAG) is the leading cause of irreversible blindness worldwide for which 15 disease-associated loci had been discovered. Among them, only 5 loci have been associated with POAG in Asians. We carried out a genome-wide association study and a replication study that included a total of 7378 POAG cases and 36 385 controls from a Japanese population. After combining the genome-wide association study and the two replication sets, we identified 11 POAG-associated loci, including 4 known (CDKN2B-AS1, ABCA1, SIX6 and AFAP1) and 7 novel loci (FNDC3B, ANKRD55-MAP3K1, LMX1B, LHPP, HMGA2, MEIS2 and LOXL1) at a genome-wide significance level (P < 5.0×10-8), bringing the total number of POAG-susceptibility loci to 22. The 7 novel variants were subsequently evaluated in a multiethnic population comprising non-Japanese East Asians (1008 cases, 591 controls), Europeans (5008 cases, 35 472 controls) and Africans (2341 cases, 2037 controls). The candidate genes located within the new loci were related to ocular development (LMX1B, HMGA2 and MAP3K1) and glaucoma-related phenotypes (FNDC3B, LMX1B and LOXL1). Pathway analysis suggested epidermal growth factor receptor signaling might be involved in POAG pathogenesis. Genetic correlation analysis revealed the relationships between POAG and systemic diseases, including type 2 diabetes and cardiovascular diseases. These results improve our understanding of the genetic factors that affect the risk of developing POAG and provide new insight into the genetic architecture of POAG in Asians.

A pyruvate dehydrogenase kinase inhibitor prevents retinal cell death and improves energy metabolism in rat retinas after ischemia/reperfusion injury.

We aimed to assess the neuroprotective effect of a pyruvate dehydrogenase kinase (PDK) inhibitor, Nov3r after ischemia/reperfusion (IR) injury in rats. IR injury was induced by applying 150 mmHg of intraocular pressure for 50 min. Nov3r was orally administered (100 mg/kg) 3 h before and 24 h after IR injury. TUNEL-positive cells increased and immunoreactive RBPMS-positive cells decreased in the rat retinas after IR injury. Administration of Nov3r significantly ameliorated the increase in TUNEL-positive cells and prevented the RBPMS-positive cell decrease. Similarly, the number of IR-induced Iba1-positive microglial cells was significantly reduced with Nov3r treatment. Among metabolic parameters, IR damage induced the elevation of lactate and pyruvate, and the reduction of ATP. Oral administration of Nov3r ameliorated these changes. Our data suggest that the Nov3r had a retinal neuroprotective effect in IR injury in rats. This finding suggests that the regulation of pyruvate dehydrogenase (PDH) activity has potential therapeutic value by enabling metabolic reprograming in diseases associated with ischemic retinal damage, such as diabetic retinopathy, retinopathy of prematurity, retinal vein occlusion, ischemic optic neuropathy and glaucoma.

Relationship between laser speckle flowgraphy and optical coherence tomography angiography measurements of ocular microcirculation.

PURPOSE: The purpose of this study was to investigate the relationship between laser speckle flowgraphy (LSFG) and optical coherence tomography angiography (OCTA) measurements of ocular microcirculation in normal and open-angle glaucoma (OAG) subjects. METHODS: This study included 18 eyes of 18 OAG patients and ten eyes of ten age-matched healthy controls. LSFG was used to measure mean blur rate (MBR) in the optic nerve head (ONH) vessel area (MV) and tissue area (MT). OCTA was used to measure a new parameter, peripapillary relative intensity (PRI), in the superficial retina, superficial choroid, and deep choroid. Statistical associations were then determined. RESULTS: MV, MT, superficial-retinal PRI, and superficial-choroidal PRI were lower in the OAG subjects than the controls (P = 0.02, P < 0.001, P = 0.02 and P = 0.008, respectively). Superficial-retinal PRI was correlated with MV and MT (R = 0.68, P < 0.001 and R = 0.63, P < 0.001, respectively). Superficial-choroidal PRI was also correlated with MV and MT (R = 0.45, P = 0.02 and R = 0.57, P = 0.002, respectively). Multiple regression analysis revealed that MV and MT independently contributed to superficial-retinal PRI (P = 0.008 and P = 0.04, respectively), while only MT contributed to superficial-choroidal PRI (P = 0.03). CONCLUSIONS: Our finding that OCTA-measured PRI was related to LSFG-measured MBR was reasonable, considering the vascular anatomy of the eye. Thus, PRI, like MBR, may be a promising biomarker of ocular microcirculation that can reveal the presence of ocular diseases such as OAG.

The association between systemic oxidative stress and ocular blood flow in patients with normal-tension glaucoma.

PURPOSE: To evaluate the association between ocular blood flow and biomarkers of systemic oxidative stress, as well as the potential of these biomarkers to assess normal-tension glaucoma (NTG). METHODS: This study included 73 eyes of 73 patients with NTG. We assessed ocular blood flow by measuring mean blur rate (MBR) in the optic nerve head using laser speckle flowgraphy, both overall and separately in the vessel and tissue areas. We also measured urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) and skin autofluorescence (SAF), and lastly, determined correlations between these measurements and with other clinical parameters. RESULTS: SAF was correlated with age, circumpapillary retinal nerve fiber layer thickness (cpRNFLT), mean deviation (MD), and overall MBR (P = 0.003, P = 0.013, P = 0.015 and P = 0.006, respectively). SAF and 8-OHdG were both correlated with tissue-area MBR (P = 0.006 and P = 0.010, respectively). Visual acuity, cpRNFLT, mean deviation and tissue-area MBR had a significant tendency to change with NTG severity (P = 0.014, P < 0.001, P < 0.001 and P = 0.006, respectively). Multiple regression analysis revealed that cpRNFLT and 8-OHdG were independent contributing factors to MD (P < 0.001 and P = 0.040, respectively), and that cpRNFLT and 8-OHdG were independent contributing factors to tissue-area MBR (P = 0.005 and P = 0.028, respectively). CONCLUSIONS: We found a close relationship between cpRNFLT, MD, tissue MBR, SAF and 8-OHdG, suggesting that systemic oxidative stress is associated with decreased ocular blood flow and may be involved in the pathogenesis of NTG.

Critical neuroprotective roles of heme oxygenase-1 induction against axonal injury-induced retinal ganglion cell death.

Although axonal damage induces significant retinal ganglion cell (RGC) death, small numbers of RGCs are able to survive up to 7 days after optic nerve crush (NC) injury. To develop new treatments, we set out to identify patterns of change in the gene expression of axonal damage-resistant RGCs. To compensate for the low density of RGCs in the retina, we performed retrograde labeling of these cells with 4Di-10ASP in adult mice and 7 days after NC purified the RGCs with fluorescence-activated cell sorting. Gene expression in the cells was determined with a microarray, and the expression of Ho-1 was determined with quantitative PCR (qPCR). Changes in protein expression were assessed with immunohistochemistry and immunoblotting. Additionally, the density of Fluoro-gold-labeled RGCs was counted in retinas from mice pretreated with CoPP, a potent HO-1 inducer. The microarray and qPCR analyses showed increased expression of Ho-1 in the post-NC RGCs. Immunohistochemistry also showed that HO-1-positive cells were present in the ganglion cell layer (GCL), and cell counting showed that the proportion of HO-1-positive cells in the GCL rose significantly after NC. Seven days after NC, the number of RGCs in the CoPP-treated mice was significantly higher than in the control mice. Combined pretreatment with SnPP, an HO-1 inhibitor, suppressed the neuroprotective effect of CoPP. These results reflect changes in HO-1 activity to RGCs that are a key part of RGC survival. Upregulation of HO-1 signaling may therefore be a novel therapeutic strategy for glaucoma.

Correlation between structure/function and optic disc microcirculation in myopic glaucoma, measured with laser speckle flowgraphy.

BACKGROUND: It is difficult to identify glaucoma in myopic eyes because the configuration of the optic disc varies; yet it is important clinically. Here, we used laser speckle flowgraphy (LSFG) to measure mean blur rate (MBR), representing optic disc microcirculation, and assessed its ability to identify glaucoma in eyes with myopic optic discs. METHODS: 129 eyes (normal disc: 21 eyes; myopic disc: 108 eyes) were enrolled. The eyes were classified as normal or mildly, moderately, or severely glaucomatous with standard automated perimetry (SAP). We determined the relationship between optic nerve head (ONH) MBR, measured with LSFG, mean deviation (MD), measured with SAP, and circumpapillary retinal nerve fiber layer thickness (cpRNFLT), measured with optical coherence tomography (OCT). RESULTS: ONH MBR and cpRNFLT decreased significantly with the severity of glaucoma. MBR was significantly correlated with cpRNFLT and MD (r =0.65 and r =0.63, respectively). A multiple regression analysis revealed that MBR and cpRNFLT were independent factors indicating glaucoma severity. A logistic regression analysis revealed that MBR and cpRNFLT were also independent factors indicating the presence of glaucoma. In a receiver operating characteristic (ROC) analysis, MBR and cpRNFLT could both differentiate between normal and glaucomatous eyes (MBR area under the ROC curve: 0.86, with a cut-off score of 24.0 AU). CONCLUSION: These results suggest that in addition to cpRNFLT, non-invasive and objective LSFG measurements of MBR may enable the identification of glaucoma and the classification of its severity in eyes with myopic optic discs.

The Effect of ß-Blocker Eye Drops on Pulse Rate, Ocular Blood Flow, and Glaucoma Progression: A Retrospective Longitudinal Study.

INTRODUCTION: Our study was conducted to determine factors associated with the effectiveness of a ß-blocker eye drop add-on in altering pulse rate (PR) in glaucoma patients. METHODS: This retrospective study examined 236 eyes of 138 patients who received a ß-blocker eye drop add-on during follow-up. Patients were included if at least one PR measurement was available both before and after the add-on was started. We collected data on ophthalmic parameters: longitudinal PR; longitudinal choroidal blood flow, represented by laser speckle flowgraphy-measured mean blur rate (MBR); and diacron-reactive oxygen metabolites (d-ROMs). We used a multivariable linear mixed-effects model to investigate the effectiveness of the ß-blocker eye drop add-on in altering PR and examined factors contributing to a larger PR alteration after the add-on was started by analyzing the effect on PR of the interaction term between the add-on and clinical factors. We used the k-means method to classify the patients. RESULTS: The ß-blocker eye drop add-on reduced PR (- 7.61 bpm, P < 0.001). Female gender, higher PR when the add-on was started, lower central corneal thickness, and a higher d-ROM level were associated with greater reduction in PR (P < 0.05). In a cluster of patients with these clinical features, choroidal MBR increased by + 3.42% when we adjusted for change over time; MD slope, which represents the speed of glaucoma progression, improved by + 0.64 dB/year (P < 0.05). CONCLUSIONS: We identified a glaucoma subgroup in which PR decreased, choroidal blood flow increased, and glaucoma progression slowed after a ß-blocker eye drop add-on was started.

Optical Coherence Tomography and Optical Coherence Tomography Angiography: Essential Tools for Detecting Glaucoma and Disease Progression.

Early diagnosis and detection of disease progression are critical to successful therapeutic intervention in glaucoma, the leading cause of irreversible blindness worldwide. Optical coherence tomography (OCT) is a non-invasive imaging technique that allows objective quantification in vivo of key glaucomatous structural changes in the retina and the optic nerve head (ONH). Advances in OCT technology have increased the scan speed and enhanced image quality, contributing to early glaucoma diagnosis and monitoring, as well as the visualization of critically important structures deep within the ONH, such as the lamina cribrosa. OCT angiography (OCTA) is a dye-free technique for noninvasively assessing ocular microvasculature, including capillaries within each plexus serving the macula, peripapillary retina and ONH regions, as well as the deeper vessels of the choroid. This layer-specific assessment of the microvasculature has provided evidence that retinal and choroidal vascular impairments can occur during early stages of glaucoma, suggesting that OCTA-derived measurements could be used as biomarkers for enhancing detection of glaucoma and its progression, as well as to reveal novel insights about pathophysiology. Moreover, these innovations have demonstrated that damage to the macula, a critical region for the vision-related quality of life, can be observed in the early stages of glaucomatous eyes, leading to a paradigm shift in glaucoma monitoring. Other advances in software and hardware, such as artificial intelligence-based algorithms, adaptive optics, and visible-light OCT, may further benefit clinical management of glaucoma in the future. This article reviews the utility of OCT and OCTA for glaucoma diagnosis and disease progression detection, emphasizes the importance of detecting macula damage in glaucoma, and highlights the future perspective of OCT and OCTA. We conclude that the OCT and OCTA are essential glaucoma detection and monitoring tools, leading to clinical and economic benefits for patients and society.

Autophagy disruption reduces mTORC1 activation leading to retinal ganglion cell neurodegeneration associated with glaucoma.

Autophagy dysfunction has been associated with several neurodegenerative diseases including glaucoma, characterized by the degeneration of retinal ganglion cells (RGCs). However, the mechanisms by which autophagy dysfunction promotes RGC damage remain unclear. Here, we hypothesized that perturbation of the autophagy pathway results in increased autophagic demand, thereby downregulating signaling through mammalian target of rapamycin complex 1 (mTORC1), a negative regulator of autophagy, contributing to the degeneration of RGCs. We identified an impairment of autophagic-lysosomal degradation and decreased mTORC1 signaling via activation of the stress sensor adenosine monophosphate-activated protein kinase (AMPK), along with subsequent neurodegeneration in RGCs differentiated from human pluripotent stem cells (hPSCs) with a glaucoma-associated variant of Optineurin (OPTN-E50K). Similarly, the microbead occlusion model of glaucoma resulting in ocular hypertension also exhibited autophagy disruption and mTORC1 downregulation. Pharmacological inhibition of mTORC1 in hPSC-derived RGCs recapitulated disease-related neurodegenerative phenotypes in otherwise healthy RGCs, while the mTOR-independent induction of autophagy reduced protein accumulation and restored neurite outgrowth in diseased OPTN-E50K RGCs. Taken together, these results highlight an important balance between autophagy and mTORC1 signaling essential for RGC homeostasis, while disruption to these pathways contributes to neurodegenerative features in glaucoma, providing a potential therapeutic target to prevent neurodegeneration.

Neurovascular dysfunction in glaucoma.

Retinal ganglion cells, the neurons that die in glaucoma, are endowed with a high metabolism requiring optimal provision of oxygen and nutrients to sustain their activity. The timely regulation of blood flow is, therefore, essential to supply firing neurons in active areas with the oxygen and glucose they need for energy. Many glaucoma patients suffer from vascular deficits including reduced blood flow, impaired autoregulation, neurovascular coupling dysfunction, and blood-retina/brain-barrier breakdown. These processes are tightly regulated by a community of cells known as the neurovascular unit comprising neurons, endothelial cells, pericytes, Müller cells, astrocytes, and microglia. In this review, the neurovascular unit takes center stage as we examine the ability of its members to regulate neurovascular interactions and how their function might be altered during glaucomatous stress. Pericytes receive special attention based on recent data demonstrating their key role in the regulation of neurovascular coupling in physiological and pathological conditions. Of particular interest is the discovery and characterization of tunneling nanotubes, thin actin-based conduits that connect distal pericytes, which play essential roles in the complex spatial and temporal distribution of blood within the retinal capillary network. We discuss cellular and molecular mechanisms of neurovascular interactions and their pathophysiological implications, while highlighting opportunities to develop strategies for vascular protection and regeneration to improve functional outcomes in glaucoma.

Large-scale multitrait genome-wide association analyses identify hundreds of glaucoma risk loci.

Glaucoma, a leading cause of irreversible blindness, is a highly heritable human disease. Previous genome-wide association studies have identified over 100 loci for the most common form, primary open-angle glaucoma. Two key glaucoma-associated traits also show high heritability: intraocular pressure and optic nerve head excavation damage quantified as the vertical cup-to-disc ratio. Here, since much of glaucoma heritability remains unexplained, we conducted a large-scale multitrait genome-wide association study in participants of European ancestry combining primary open-angle glaucoma and its two associated traits (total sample size over 600,000) to substantially improve genetic discovery power (263 loci). We further increased our power by then employing a multiancestry approach, which increased the number of independent risk loci to 312, with the vast majority replicating in a large independent cohort from 23andMe, Inc. (total sample size over 2.8 million; 296 loci replicated at P < 0.05, 240 after Bonferroni correction). Leveraging multiomics datasets, we identified many potential druggable genes, including neuro-protection targets likely to act via the optic nerve, a key advance for glaucoma because all existing drugs only target intraocular pressure. We further used Mendelian randomization and genetic correlation-based approaches to identify novel links to other complex traits, including immune-related diseases such as multiple sclerosis and systemic lupus erythematosus.

The relationship between choroidal blood flow and glaucoma progression in a Japanese study population.

PURPOSE: To determine whether choroidal blood flow (BF) is related to visual field (VF) defect severity and progression in eyes with open-angle glaucoma (OAG). STUDY DESIGN: Retrospective and longitudinal. METHODS: This study comprised 443 eyes of 285 OAG patients who underwent laser speckle flowgraphy (LSFG), optical coherence tomography, and visual-field (VF) testing at baseline. The patients were then observed for at least 2 years and at least 5 reliable VF tests were performed. In the LSFG images, we set regions of interest at the optic nerve head (ONH) and the parapapillary choroid to obtain ONH-tissue mean blur rate (MBR) and choroidal MBR, respectively. We used univariable and multivariable linear mixed-effects models to determine clinical factors associated with choroidal MBR at baseline. We also used a linear mixed-effects model to determine the contribution of ONH-tissue MBR and choroidal MBR to baseline mean deviation (MD) and to MD slope during follow-up, adjusting for potential confounding factors, including circumpapillary retinal nerve fiber layer thickness. RESULTS: Choroidal MBR was associated with age, MD slope, and ONH-tissue MBR (ß = -0.181, P = 0.001; ß = 0.134, P = 0.002; ß = 0.096, P = 0.049, respectively). ONH-tissue MBR was associated with both MD and MD slope (ß = 0.146, P = 0.004; ß = 0.152, P = 0.009, respectively), whereas choroidal MBR was associated only with MD slope (ß = 0.147, P = 0.005). CONCLUSION: LSFG-derived choroidal MBR might be a useful biomarker to predict VF defect progression in a Japanese population.

Restoration of mitochondria axonal transport by adaptor Disc1 supplementation prevents neurodegeneration and rescues visual function.

Deficits in mitochondrial transport are a common feature of neurodegenerative diseases. We investigated whether loss of components of the mitochondrial transport machinery impinge directly on metabolic stress, neuronal death, and circuit dysfunction. Using multiphoton microscope live imaging, we showed that ocular hypertension, a major risk factor in glaucoma, disrupts mitochondria anterograde axonal transport leading to energy decline in vulnerable neurons. Gene- and protein-expression analysis revealed loss of the adaptor disrupted in schizophrenia 1 (Disc1) in retinal neurons subjected to high intraocular pressure. Disc1 gene delivery was sufficient to rescue anterograde transport and replenish axonal mitochondria. A genetically encoded ATP sensor combined with longitudinal live imaging showed that Disc1 supplementation increased ATP production in stressed neurons. Disc1 gene therapy promotes neuronal survival, reverses abnormal single-cell calcium dynamics, and restores visual responses. Our study demonstrates that enhancing anterograde mitochondrial transport is an effective strategy to alleviate metabolic stress and neurodegeneration.

METTL23 mutation alters histone H3R17 methylation in normal-tension glaucoma.

Normal-tension glaucoma (NTG) is a heterogeneous disease characterized by retinal ganglion cell (RGC) death leading to cupping of the optic nerve head and visual field loss at normal intraocular pressure (IOP). The pathogenesis of NTG remains unclear. Here, we describe a single nucleotide mutation in exon 2 of the methyltransferase-like 23 (METTL23) gene identified in 3 generations of a Japanese family with NTG. This mutation caused METTL23 mRNA aberrant splicing, which abolished normal protein production and altered subcellular localization. Mettl23-knock-in (Mettl23+/G and Mettl23G/G) and -knockout (Mettl23+/- and Mettl23-/-) mice developed a glaucoma phenotype without elevated IOP. METTL23 is a histone arginine methyltransferase expressed in murine and macaque RGCs. However, the novel mutation reduced METTL23 expression in RGCs of Mettl23G/G mice, which recapitulated both clinical and biological phenotypes. Moreover, our findings demonstrated that METTL23 catalyzed the dimethylation of H3R17 in the retina and was required for the transcription of pS2, an estrogen receptor α target gene that was critical for RGC homeostasis through the negative regulation of NF-κB-mediated TNF-α and IL-1ß feedback. These findings suggest an etiologic role of METTL23 in NTG with tissue-specific pathology.

Genetic variants associated with glaucomatous visual field loss in primary open-angle glaucoma.

Primary open-angle glaucoma (POAG) is characterized by a progressive optic neuropathy with visual field loss. To investigate the genetic variants associated with visual field loss in POAG, Japanese POAG patients (n = 426) and control subjects (n = 246) were genotyped for 22 genetic variants predisposing to POAG that can be classified into those associated with intraocular pressure (IOP) elevation (IOP-related genetic variants) and optic nerve vulnerability independent of IOP (optic nerve-related genetic variants). The genetic risk score (GRS) of the 17 IOP-related and five optic nerve-related genetic variants was calculated, and the associations between the GRS and the mean deviation (MD) of automated static perimetry as an indicator of the severity of visual field loss and pattern standard deviation (PSD) as an indicator of the focal disturbance were evaluated. There was a significant association (Beta = - 0.51, P = 0.0012) between the IOP-related GRS and MD. The severity of visual field loss may depend on the magnitude of IOP elevation induced by additive effects of IOP-related genetic variants. A significant association (n = 135, Beta = 0.65, P = 0.0097) was found between the optic nerve-related, but not IOP-related, GRS and PSD. The optic nerve-related (optic nerve vulnerability) and IOP-related (IOP elevation) genetic variants may play an important role in the focal and diffuse visual field loss respectively. To our knowledge, this is the first report to show an association between additive effects of genetic variants predisposing to POAG and glaucomatous visual field loss, including severity and focal/diffuse disturbance of visual field loss, in POAG.