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BACKGROUND: Computed tomography (CT) derived body composition measurements of sarcopenia are an emerging form of prognostication in many disease processes. Although the L3 vertebral level is commonly used to measure skeletal muscle mass, other studies have suggested the utilisation of other segments. This study was performed to assess the variation and reproducibility of skeletal muscle mass at vertebral levels T4, T12 and L3 in pre-operative rectal cancer patients. If thoracic measurements were equivalent to those at L3, it will allow for body composition comparisons in a larger range of cancers where lumbar CT images are not routinely measured. RESEARCH METHODS: Patients with stage I - III rectal cancer, undergoing curative resection from 2010 - 2014, were assessed. CT based quantification of skeletal muscle was used to determine skeletal muscle cross sectional area (CSA) and skeletal muscle index (SMI). Systematic differences between the measurements at L3 with T4 and T12 vertebral levels were evaluated by percentile rank differences to assess distribution of differences and ordinary least product regression (OLP) to detect and distinguish fixed and proportional bias. RESULTS: Eighty eligible adult patients were included. Distribution of differences between T12 SMI and L3 SMI were more marked than differences between T4 SMI and L3 SMI. There was no fix or proportional bias with T4 SMI, but proportional bias was detected with T12 SMI measurements. T4 CSA duplicate measurements had higher test-retest reliability: coefficient of repeatability was 34.10 cm2 for T4 CSA vs 76.00 cm2 for T12 CSA. Annotation time (minutes) with L3 as reference, the median difference was 0.85 for T4 measurements and -0.03 for T12 measurements. Thirty-seven patients (46%) had evidence of sarcopenia at the L3 vertebral level, with males exhibiting higher rates of sarcopenia. However, there was no association between sarcopenia and post-operative complications, recurrence or hospital LOS (le

Inherited retinal dystrophies (IRDs) as well as genetically complex retinal phenotypes represent a heterogenous group of ocular diseases, both on account of their phenotypic and genotypic characteristics. Therefore, overlaps in clinical features often complicate or even impede their correct clinical diagnosis. Deciphering the molecular basis of retinal diseases has not only aided in their disease classification but also helped in our understanding of how different molecular pathologies may share common pathomechanisms. In particular, these relate to dysregulation of two key processes that contribute to cellular integrity, namely extracellular matrix (ECM) homeostasis and inflammation. Pathological changes in the ECM of Bruch's membrane have been described in both monogenic IRDs, such as Sorsby fundus dystrophy (SFD) and Doyne honeycomb retinal dystrophy (DHRD), as well as in the genetically complex age-related macular degeneration (AMD) or diabetic retinopathy (DR). Additionally, complement system dysfunction and distorted immune regulation may also represent a common connection between some IRDs and complex retinal degenerations. Through highlighting such overlaps in molecular pathology, this review aims to illuminate how inflammatory processes and ECM homeostasis are linked in the healthy retina and how their interplay may be disturbed in aging as well as in disease.

Purpose: This study aimed to stratify eyes with keratoconus (KC) based on longitudinal changes in all Pentacam parameters into clusters using unsupervised machine learning, with the broader objective of more clearly defining the characteristics of KC progression. Methods: A data-driven cluster analysis (hierarchical clustering) was undertaken on a retrospective cohort of 1017 kC eyes and 128 control eyes. Clusters were derived using 6-month tomographical change in individual eyes from analysis of the reduced dimensionality parameter space using all available Pentacam parameters (406 principal components). The optimal number of clusters was determined by the clustering's capacity to discriminate progression between KC and control eyes based on change across parameters. One-way ANOVA was used to compare parameters between inferred clusters. Complete Pentacam data changes at 6, 12 and 18-month time points provided validation datasets to determine the generalizability of the clustering model. Results: We identified three clusters in KC progression patterns. Eyes designated within cluster 3 had the most rapidly changing tomographical parameters compared to eyes in either cluster 1 or 2. Eyes designated within cluster 1 reflected minimal changes in tomographical parameters, closest to the tomographical changes of control (non-KC) eyes. Thirty-nine corneal curvature parameters were identified and associated with these stratified clusters, with each of these parameters changing significantly different between three clusters. Similar clusters were identified at the 6, 12 and 18-month follow-up. Conclusions: The clustering model developed was able to automatically detect and categorize KC tomographical features into fast, slow, or limited change at different time points. This new KC stratification tool may provide an opportunity to provide a precision medicine approach to KC.

CLINICAL RELEVANCE: The Keratoconus International Consortium (KIC) will allow better understanding of keratoconus. BACKGROUND: Keratoconus is a disorder characterised by corneal elevation and thinning, leading to reduced vision. The current gaps in understanding of this disease will be discussed and the need for a multi-pronged and multi-centre engagement to enhance our understanding of keratoconus will be highlighted. DESIGN: KIC has been established to address the gaps in our understanding of keratoconus with the aim of collecting baseline as well as longitudinal data on several fields. PARTICIPANTS: Keratoconus and control (no corneal condition) subjects from different sites globally will be recruited in the study. METHODS: KIC collects data using an online, secure database, which enables standardised data collection at member sites. Data fields collected include medical history, clinical features, quality of life and economic burden questionnaires and possible genetic sample collection from patients of different ethnicities across different geographical locations. RESULTS: There are currently 40 Australian and international clinics or hospital departments who have joined the KIC. Baseline data has so far been collected on 1130 keratoconus patients and indicates a median age of 29.70 years with 61% being male. A total of 15.3% report a positive family history of keratoconus and 57.7% self-report a history of frequent eye rubbing. CONCLUSION: The strength of this consortium is its international, collaborative design and use of a common data collection tool. Inclusion and analyses of cross-sectional and longitudinal data will help answer many questions that remain in keratoconus, including factors affecting progression and treatment outcomes.

(1) Background: The objective of this review was to synthesize available data on the use of machine learning to evaluate its accuracy (as determined by pooled sensitivity and specificity) in detecting keratoconus (KC), and measure reporting completeness of machine learning models in KC based on TRIPOD (the transparent reporting of multivariable prediction models for individual prognosis or diagnosis) statement. (2) Methods: Two independent reviewers searched the electronic databases for all potential articles on machine learning and KC published prior to 2021. The TRIPOD 29-item checklist was used to evaluate the adherence to reporting guidelines of the studies, and the adherence rate to each item was computed. We conducted a meta-analysis to determine the pooled sensitivity and specificity of machine learning models for detecting KC. (3) Results: Thirty-five studies were included in this review. Thirty studies evaluated machine learning models for detecting KC eyes from controls and 14 studies evaluated machine learning models for detecting early KC eyes from controls. The pooled sensitivity for detecting KC was 0.970 (95% CI 0.949-0.982), with a pooled specificity of 0.985 (95% CI 0.971-0.993), whereas the pooled sensitivity of detecting early KC was 0.882 (95% CI 0.822-0.923), with a pooled specificity of 0.947 (95% CI 0.914-0.967). Between 3% and 48% of TRIPOD items were adhered to in studies, and the average (median) adherence rate for a single TRIPOD item was 23% across all studies. (4) Conclusions: Application of machine learning model has the potential to make the diagnosis and monitoring of KC more efficient, resulting in reduced vision loss to the patients. This review provides current information on the machine learning models that have been developed for detecting KC and early KC. Presently, the machine learning models performed poorly in identifying early KC from control eyes and many of these research studies did not follow established reporting standa

Keratoconus is a complex and multifactorial disease and its exact aetiology remains unknown. This current study examined the important environmental risk factors and their association with keratoconus. This study was registered in the PROSPERO International Prospective Register of systematic reviews under registration number CRD42021256792 in 2021. Scopus, Web of Science, PubMed, and Cochrane CENTRAL databases were searched for all relevant articles published from 1 January 1900 to 31 July 2021. National Institutes of Health Quality Assessment Tool was used to assess the methodological quality of the studies. The assessment for statistical heterogeneity was assessed using the Z-statistics on RevMan v5.4. P-value of <0.05 was considered as statistically significant and I2 < 25% as homogenous. Thirty studies were included in this meta-analysis. Pooled odds ratio was calculated with 95% CI. The pooled odds ratio (OR) of eye rubbing, atopy, asthma, and eczema was 3.64 (95% CI, 2.02, 6.57), 1.90 (95% CI, 1.22, 2.94), 1.36 (95% CI, 1.15, 1.61) and 1.90 (95% CI, 1.22, 2.94), respectively. The OR for diabetes was 0.86 (95% CI 0.73, 1.02), and use of sunglasses, contact lens, allergic conjunctivitis, side sleep position and prone sleep position was 0.40 (95% CI, 0.16, 0.99), 1.68 (0.70, 4.00), 2.24 (95% CI, 0.68, 7.36), 3.81 (95% CI, 0.31, 46.23), 12.76 (95% CI, 0.27, 598.58), respectively. Twenty studies were considered to be of high quality, nine to be moderate and one to be low. Environmental risk factors have been identified to play a role in the susceptibility of keratoconus. However, further large-scale longitudinal studies are needed to understand the mechanisms between environmental risk factors and keratoconus.

Age-related macular degeneration (AMD) is the leading cause of severe irreversible central vision loss in individuals over 65 years old. Genome-wide association studies (GWASs) have shown that the region at chromosome 10q26, where the age-related maculopathy susceptibility (ARMS2/LOC387715) and HtrA serine peptidase 1 (HTRA1) genes are located, represents one of the strongest associated loci for AMD. However, the underlying biological mechanism of this genetic association has remained elusive. In this article, we extensively review the literature by us and others regarding the ARMS2/HTRA1 risk alleles and their functional significance. We also review the literature regarding the presumed function of the ARMS2 protein and the molecular processes of the HTRA1 protein in AMD pathogenesis in vitro and in vivo, including those of transgenic mice overexpressing HtrA1/HTRA1 which developed Bruch's membrane (BM) damage, choroidal neovascularization (CNV), and polypoidal choroidal vasculopathy (PCV), similar to human AMD patients. The elucidation of the molecular mechanisms of the ARMS2 and HTRA1 susceptibility loci has begun to untangle the complex biological pathways underlying AMD pathophysiology, pointing to new testable paradigms for treatment.

METHODS: We conducted grounded theory semistructured interviews, purposively inviting participants until themed saturation was met. Sentiment analysis was used to determine opinion. RESULTS: We interviewed n = 92 global eye tissue and eye bank professionals. We determined that corneal tissue, which is exported, costs between US $100 and US $6000 or is provided as gratis. Collectively, interviewees indicated that, globally, there were no fixed fee structures in place, and the fee was influenced by multiple factors on both export and import sides. They indicated that ultimately corneas were allocated based on the importers' ability to pay the price determined by the exporting eye bank. DISCUSSION: Allocation of corneal tissue, which is exported, is influenced by the fees charged by the exporters to meet their bottom line and the funds available to importers. Therefore, export allocation is not equitable, with those who can pay a higher fee, prioritized. Steps to guide and support exporters with the development of fee structures that promote equitable allocation are essential. This will assist both export and import eye bank development, corneal tissue access development, and those awaiting a corneal transplant.

Keratoconus is characterised by reduced rigidity of the cornea with distortion and focal thinning that causes blurred vision, however, the pathogenetic mechanisms are unknown. It can lead to severe visual morbidity in children and young adults and is a common indication for corneal transplantation worldwide. Here we report the first large scale genome-wide association study of keratoconus including 4,669 cases and 116,547 controls. We have identified significant association with 36 genomic loci that, for the first time, implicate both dysregulation of corneal collagen matrix integrity and cell differentiation pathways as primary disease-causing mechanisms. The results also suggest pleiotropy, with some disease mechanisms shared with other corneal diseases, such as Fuchs endothelial corneal dystrophy. The common variants associated with keratoconus explain 12.5% of the genetic variance, which shows potential for the future development of a diagnostic test to detect susceptibility to disease.

BACKGROUND: 12.7 million people await a corneal transplant, but 53% are without access to corneal tissue. Sharing corneal tissue across nations can provide some access, however the willingness of export populations, like Australians, to export their donation on death, has never been evaluated. Our research samples the Australian population, determining their willingness to export. MATERIALS AND METHOD: We conducted e-surveys. N = 1044 Australians participated. The sample represented the Australian population, based on population demographics. Chi-Square and bivariate correlation coefficients examined associations between categorical variables, with a sample size of N = 1044, power of 0.80, and alpha of p = 0.05. Outcome measures were based on population sampling, by exploring willingness export, through the e-survey method. RESULTS: 38% (n = 397) of respondents said yes to exportation, 23.8% (n = 248) said no, and 38.2% (n = 399) were undecided. We found no relationship between willingness to export and general demographics, though those registered on the Donatelife Register (p = < .001), and those already willing to donate their eyes (p = < .001) were significantly more willing to export. DISCUSSION: More Australians are willing to export their corneas than not, though a significant portion remain undecided. The Donatelife Register, and donation awareness, are key components of respondent decision making. Therefore, the provision of information about exportation prior to, and at the point-of-donation, is essential for assisting Australian's to decide to export or not. Further examination and development of consent-for-export systems are necessary before routine exportation is undertaken.

PURPOSE: To investigate the performance of a machine learning model based on a reduced dimensionality parameter space derived from complete Pentacam parameters to identify subclinical keratoconus (KC). METHODS: All 1692 available parameters were obtained from the Pentacam imaging machine on 145 subclinical KC and 122 control eyes. We applied a principal component analysis (PCA) to the complete Pentacam dataset to reduce its parameter dimensionality. Subsequently, we investigated machine learning performance of the random forest algorithm with increasing numbers of components to identify their optimal number for detecting subclinical KC from control eyes. RESULTS: The dimensionality of the complete set of 1692 Pentacam parameters was reduced to 267 principal components using PCA. Subsequent selection of 15 of these principal components explained over 85% of the variance of the original Pentacam-derived parameters and input to train a random forest machine learning model to achieve the best accuracy of 98% in detecting subclinical KC eyes. The model established also reached a high sensitivity of 97% in identification of subclinical KC and a specificity of 98% in recognizing control eyes. CONCLUSIONS: A random forest-based model trained using a modest number of components derived from a reduced dimensionality representation of complete Pentacam system parameters allowed for high accuracy of subclinical KC identification.

PURPOSE: This study describes the development of a deep learning algorithm based on the U-Net architecture for automated segmentation of geographic atrophy (GA) lesions in fundus autofluorescence (FAF) images. METHODS: Image preprocessing and normalization by modified adaptive histogram equalization were used for image standardization to improve effectiveness of deep learning. A U-Net-based deep learning algorithm was developed and trained and tested by fivefold cross-validation using FAF images from clinical datasets. The following metrics were used for evaluating the performance for lesion segmentation in GA: dice similarity coefficient (DSC), DSC loss, sensitivity, specificity, mean absolute error (MAE), accuracy, recall, and precision. RESULTS: In total, 702 FAF images from 51 patients were analyzed. After fivefold cross-validation for lesion segmentation, the average training and validation scores were found for the most important metric, DSC (0.9874 and 0.9779), for accuracy (0.9912 and 0.9815), for sensitivity (0.9955 and 0.9928), and for specificity (0.8686 and 0.7261). Scores for testing were all similar to the validation scores. The algorithm segmented GA lesions six times more quickly than human performance. CONCLUSIONS: The deep learning algorithm can be implemented using clinical data with a very high level of performance for lesion segmentation. Automation of diagnostics for GA assessment has the potential to provide savings with respect to patient visit duration, operational cost and measurement reliability in routine GA assessments. TRANSLATIONAL RELEVANCE: A deep learning algorithm based on the U-Net architecture and image preprocessing appears to be suitable for automated segmentation of GA lesions on clinical data, producing fast and accurate results.

Recovery and access to end-of-life corneal tissue for corneal transplantation, training and research is globally maldistributed. The reasons for the maldistribution are complex and multifaceted, and not well defined or understood. Currently there are few solutions available to effectively address these issues. This review provides an overview of the system, key issues impacting recovery and allocation and emphasises how end-user ophthalmologists and researchers, with support from administrators and the wider sector, can assist in increasing access long-term through sustaining eye banks nationally and globally. We posit that prevention measures and improved surgical techniques, together with the development of novel therapies will play a significant role in reducing demand and enhance the equitable allocation of corneas.

PURPOSE: To identify the most suitable model for assessing the rate of growth of total geographic atrophy (GA) by analysis of model structure uncertainty. METHODS: Model structure uncertainty refers to unexplained variability arising from the choice of mathematical model and represents an example of epistemic uncertainty. In this study, we quantified this uncertainty to help identify a model most representative of GA progression. Fundus autofluorescence (FAF) images and GA progression data (i.e., total GA area estimation at each presentation) were acquired using Spectralis HRA+OCT instrumentation and RegionFinder software. Six regression models were evaluated. Models were compared using various statistical tests, [i.e., coefficient of determination (r2), uncertainty metric (U), and test of significance for the correlation coefficient, r], as well as adherence to expected physical and clinical assumptions of GA growth. RESULTS: Analysis was carried out for 81 GA-affected eyes, 531 FAF images (range: 3-17 images per eye), over median of 57 months (IQR: 42, 74), with a mean baseline lesion size of 2.62 ± 4.49 mm2 (range: 0.11-20.69 mm2). The linear model proved to be the most representative of total GA growth, with lowest average uncertainty (original scale: U = 0.025, square root scale: U = 0.014), high average r2 (original scale: 0.92, square root scale: 0.93), and applicability of the model was supported by a high correlation coefficient, r, with statistical significance (P = 0.01). CONCLUSIONS: Statistical analysis of uncertainty suggests that the linear model provides an effective and practical representation of the rate and progression of total GA growth based on data from patient presentations in clinical settings. TRANSLATIONAL RELEVANCE: Identification of correct model structure to characterize rate of growth of total GA in the retina using FAF images provides an objective metric for comparing interventions and charting GA progression in clinical presentations

Corneal curvature, a highly heritable trait, is a key clinical endophenotype for myopia - a major cause of visual impairment and blindness in the world. Here we present a trans-ethnic meta-analysis of corneal curvature GWAS in 44,042 individuals of Caucasian and Asian with replication in 88,218 UK Biobank data. We identified 47 loci (of which 26 are novel), with population-specific signals as well as shared signals across ethnicities. Some identified variants showed precise scaling in corneal curvature and eye elongation (i.e. axial length) to maintain eyes in emmetropia (i.e. HDAC11/FBLN2 rs2630445, RBP3 rs11204213); others exhibited association with myopia with little pleiotropic effects on eye elongation. Implicated genes are involved in extracellular matrix organization, developmental process for body and eye, connective tissue cartilage and glycosylation protein activities. Our study provides insights into population-specific novel genes for corneal curvature, and their pleiotropic effect in regulating eye size or conferring susceptibility to myopia.

Purpose: Keratoconus (KC) represents one of the leading causes of corneal transplantation worldwide. Detecting subclinical KC would lead to better management to avoid the need for corneal grafts, but the condition is clinically challenging to diagnose. We wished to compare eight commonly used machine learning algorithms using a range of parameter combinations by applying them to our KC dataset and build models to better differentiate subclinical KC from non-KC eyes. Methods: Oculus Pentacam was used to obtain corneal parameters on 49 subclinical KC and 39 control eyes, along with clinical and demographic parameters. Eight machine learning methods were applied to build models to differentiate subclinical KC from control eyes. Dominant algorithms were trained with all combinations of the considered parameters to select important parameter combinations. The performance of each model was evaluated and compared. Results: Using a total of eleven parameters, random forest, support vector machine and k-nearest neighbors had better performance in detecting subclinical KC. The highest area under the curve of 0.97 for detecting subclinical KC was achieved using five parameters by the random forest method. The highest sensitivity (0.94) and specificity (0.90) were obtained by the support vector machine and the k-nearest neighbor model, respectively. Conclusions: This study showed machine learning algorithms can be applied to identify subclinical KC using a minimal parameter set that are routinely collected during clinical eye examination. Translational Relevance: Machine learning algorithms can be built using routinely collected clinical parameters that will assist in the objective detection of subclinical KC.

IMPORTANCE: This is the first study to estimate the lifetime costs associated with keratoconus based on a questionnaire completed by patients and highlights the significant economic burden of the disease. As keratoconus affects individuals from a young age, the study highlights keratoconus as a public health concern. BACKGROUND: Keratoconus is a disorder characterized by corneal steepening and thinning, leading to reduced visual acuity. To date, there have been no studies evaluating the economic costs of keratoconus from a patient's perspective. DESIGN: A randomized cross-sectional study undertaken in Australia where keratoconus subjects were recruited from public and private ophthalmology and optometry clinics. PARTICIPANTS: A total of 100 participants completed the questionnaire: median age was 31 years and 57% were males. METHODS: A keratoconus health expenditure questionnaire was designed to assess direct and indirect expenditures for each individual. MAIN OUTCOME MEASURES: Total direct and indirect costs associated with the condition were calculated along with the estimated lifetime per capita costs. RESULTS: The total cost related to direct and indirect care was estimated to be AUD 3365. By applying our cost data to keratoconus prevalence data for the Australian population, the total cost is estimated to be approximately AUD 44.7 million per year in Australia. CONCLUSIONS AND RELEVANCE: Our results show that the costs associated with the diagnosis and management of keratoconus represent a significant cost to patients. An understanding of this is important not only to individuals and their families, but also health care providers, health insurers and the wider health system.

PURPOSE: The purpose of this study was to summarize and evaluate artificial intelligence (AI) algorithms used in geographic atrophy (GA) diagnostic processes (e.g. isolating lesions or disease progression). METHODS: The search strategy and selection of publications were both conducted in accordance with the Preferred of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. PubMed and Web of Science were used to extract literary data. The algorithms were summarized by objective, performance, and scope of coverage of GA diagnosis (e.g. lesion automation and GA progression). RESULTS: Twenty-seven studies were identified for this review. A total of 18 publications focused on lesion segmentation only, 2 were designed to detect and classify GA, 2 were designed to predict future overall GA progression, 3 focused on prediction of future spatial GA progression, and 2 focused on prediction of visual function in GA. GA-related algorithms reported sensitivities from 0.47 to 0.98, specificities from 0.73 to 0.99, accuracies from 0.42 to 0.995, and Dice coefficients from 0.66 to 0.89. CONCLUSIONS: Current GA-AI publications have a predominant focus on lesion segmentation and a minor focus on classification and progression analysis. AI could be applied to other facets of GA diagnoses, such as understanding the role of hyperfluorescent areas in GA. Using AI for GA has several advantages, including improved diagnostic accuracy and faster processing speeds. TRANSLATIONAL RELEVANCE: AI can be used to quantify GA lesions and therefore allows one to impute visual function and quality-of-life. However, there is a need for the development of reliable and objective models and software to predict the rate of GA progression and to quantify improvements due to interventions.

The knowledge on the genetic background of refractive error and myopia has expanded dramatically in the past few years. This white paper aims to provide a concise summary of current genetic findings and defines the direction where development is needed. We performed an extensive literature search and conducted informal discussions with key stakeholders. Specific topics reviewed included common refractive error, any and high myopia, and myopia related to syndromes. To date, almost 200 genetic loci have been identified for refractive error and myopia, and risk variants mostly carry low risk but are highly prevalent in the general population. Several genes for secondary syndromic myopia overlap with those for common myopia. Polygenic risk scores show overrepresentation of high myopia in the higher deciles of risk. Annotated genes have a wide variety of functions, and all retinal layers appear to be sites of expression. The current genetic findings offer a world of new molecules involved in myopiagenesis. As the missing heritability is still large, further genetic advances are needed. This Committee recommends expanding large-scale, in-depth genetic studies using complementary big data analytics, consideration of gene-environment effects by thorough measurement of environmental exposures, and focus on subgroups with extreme phenotypes and high familial occurrence. Functional characterization of associated variants is simultaneously needed to bridge the knowledge gap between sequence variance and consequence for eye growth.

BACKGROUND: Keratoconus (KC) is a common corneal condition with an unknown gender predominance. Although numerous studies have investigated the genetic component of KC, no specific genes have yet been attributed to the condition. We recently reported posterior segment changes occurring in the eyes of KC patients. However, it is not clear whether these changes are part of KC pathogenesis or reflect changes in anatomical features of the eye manifested by changes at the cornea. Given retinal changes represent the main characteristics observed in age-related macular degeneration (AMD) and that pleiotropy has been demonstrated between different eye diseases, we wished to assess if known AMD associated genes were also associated with KC. METHODS: A total of 248 KC subjects and 366 non-KC (control) subjects were recruited from public and private clinics in Melbourne for this analysis. Nineteen single nucleotide polymorphisms (SNPs) previously associated with AMD, including rs10490924 (ARMS2/HTRA1), rs10737680 (CFH), rs13278062 (TNFRSF10A), rs1864163 (CETP), rs2230199 (C3), rs3130783 (IER3/DDR1), rs334353 (TGFBR1), rs3812111 (COL10A1), rs429608 (C2/CFB), rs4420638 (APOE), rs4698775 (CFI), rs5749482 (TIMP3), rs6795735 (ADAMTS9), rs8017304 (RAD51B), rs8135665 (SLC16A8), rs920915 (LIPC), rs943080 (VEGFA), rs9542236 (B3GALTL) and rs13081855 (COL8A1/FILIP1L), were genotyped in this cohort. Logistic regression was applied to evaluate the association between these SNPs and KC on both genders together, as well as each gender separately. Linear regression was also applied to assess the association between SNPs and corneal curvature. Bonferroni correction was applied to adjust for multiple testing. RESULTS: Genotyping data were available for 18 SNPs. The SNP, rs6795735 (ADAMTS9) was significantly associated with KC (p = 3.5 × 10- 4) when both genders were assessed, whereas rs5749482 (TIMP3) was only associated in males (p = 7.7 × 10- 4) following Bonferroni multiple correction. Howev

PURPOSE: Age-related macular degeneration (AMD) is the worldwide leading cause of blindness among the elderly. Although genome-wide association studies (GWAS) have identified AMD risk variants, their roles in disease etiology are not well-characterized, and they only explain a portion of AMD heritability. METHODS: We performed pathway analyses using summary statistics from the International AMD Genomics Consortium's 2016 GWAS and multiple pathway databases to identify biological pathways wherein genetic association signals for AMD may be aggregating. We determined which genes contributed most to significant pathway signals across the databases. We characterized these genes by constructing protein-protein interaction networks and performing motif analysis. RESULTS: We determined that eight genes (C2, C3, LIPC, MICA, NOTCH4, PLCG2, PPARA, and RAD51B) "drive" the statistical signals observed across pathways curated in the Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome, and Gene Ontology (GO) databases. We further refined our definition of statistical driver gene to identify PLCG2 as a candidate gene for AMD due to its significant gene-level signals (P < 0.0001) across KEGG, Reactome, GO, and NetPath pathways. CONCLUSIONS: We performed pathway analyses on the largest available collection of advanced AMD cases and controls in the world. Eight genes strongly contributed to significant pathways from the three larger databases, and one gene (PLCG2) was central to significant pathways from all four databases. This is, to our knowledge, the first study to identify PLCG2 as a candidate gene for AMD based solely on genetic burden. Our findings reinforce the utility of integrating in silico genetic and biological pathway data to investigate the genetic architecture of AMD.

Emmanuelle Souzeau, who contributed to analysis of data, was inadvertently omitted from the author list in the originally published version of this Article. This has now been corrected in both the PDF and HTML versions of the Article.

PURPOSE: To identify genes and genetic markers associated with corneal astigmatism. METHODS: A meta-analysis of genome-wide association studies (GWASs) of corneal astigmatism undertaken for 14 European ancestry (n=22,250) and 8 Asian ancestry (n=9,120) cohorts was performed by the Consortium for Refractive Error and Myopia. Cases were defined as having >0.75 diopters of corneal astigmatism. Subsequent gene-based and gene-set analyses of the meta-analyzed results of European ancestry cohorts were performed using VEGAS2 and MAGMA software. Additionally, estimates of single nucleotide polymorphism (SNP)-based heritability for corneal and refractive astigmatism and the spherical equivalent were calculated for Europeans using LD score regression. RESULTS: The meta-analysis of all cohorts identified a genome-wide significant locus near the platelet-derived growth factor receptor alpha (PDGFRA) gene: top SNP: rs7673984, odds ratio=1.12 (95% CI:1.08-1.16), p=5.55×10-9. No other genome-wide significant loci were identified in the combined analysis or European/Asian ancestry-specific analyses. Gene-based analysis identified three novel candidate genes for corneal astigmatism in Europeans-claudin-7 (CLDN7), acid phosphatase 2, lysosomal (ACP2), and TNF alpha-induced protein 8 like 3 (TNFAIP8L3). CONCLUSIONS: In addition to replicating a previously identified genome-wide significant locus for corneal astigmatism near the PDGFRA gene, gene-based analysis identified three novel candidate genes, CLDN7, ACP2, and TNFAIP8L3, that warrant further investigation to understand their role in the pathogenesis of corneal astigmatism. The much lower number of genetic variants and genes demonstrating an association with corneal astigmatism compared to published spherical equivalent GWAS analyses suggest a greater influence of rare genetic variants, non-additive genetic effects, or environmental factors in the development of astigmatism.

PURPOSE: Presently, 52 independent nuclear single nucleotide polymorphisms (nSNPs) have been associated with age-related macular degeneration (AMD) but their effects do not explain all its variance. Genetic interactions between the nuclear and mitochondrial (mt) genome may unearth additional genetic loci previously unassociated with AMD risk. METHODS: Joint effects of nSNPs and selected mtSNPs were analyzed by two degree of freedom (2df) joint tests of association in the International AMD Genomics Consortium (IAMDGC) dataset (17,832 controls and 16,144 advanced AMD cases of European ancestry). Subjects were genotyped on the Illumina HumanCoreExome array. After imputation using MINIMAC and the 1000 Genomes Project Phase I reference panel, pairwise linkage disequilibrium pruning, and quality control, 3.9 million nSNPs were analyzed for interaction with mtSNPs chosen based on association in this dataset or publications: A4917G, T5004C, G12771A, and C16069T. RESULTS: Novel locus TRPM1 was identified with genome-wide significant joint effects (P < 5.0 × 10-8) of two intronic TRPM1 nSNPs and AMD-associated nonsynonymous MT-ND2 mtSNP A4917G. Stratified analysis by mt allele identified an association only in 4917A (major allele) carriers (P = 4.4 × 10-9, odds ratio [OR] = 0.90, 95% confidence interval [CI] = 0.87-0.93). Intronic and intergenic ABHD2/RLBP1 nSNPs demonstrated genome-wide significant joint effects (2df joint test P values from 1.8 × 10-8 to 4.9 × 10-8) and nominally statistically significant interaction effects with MT-ND5 synonymous mtSNP G12771A. Although a positive association was detected in both strata, the association was stronger in 12771A subjects (P = 0.0020, OR = 2.17, 95% CI = 1.34-3.60). CONCLUSIONS: These results show that joint tests of main effects and gene-gene interaction reveal associations at some novel loci that were missed when considering main effects alone.

BACKGROUND: Age-related macular degeneration (AMD) is the leading cause of vision loss in Western societies with a strong genetic component. Candidate gene studies as well as genome-wide association studies strongly implicated genetic variations in complement genes to be involved in disease risk. So far, no association of AMD with complement component 4 (C4) was reported probably due to the complex nature of the C4 locus on chromosome 6. METHODS: We used multiplex ligation-dependent probe amplification (MLPA) to determine the copy number of the C4 gene as well as of both relevant isoforms, C4A and C4B, and assessed their association with AMD using logistic regression models. RESULTS: Here, we report on the analysis of 2645 individuals (1536 probands and 1109 unaffected controls), across three different centers, for multiallelic copy number variation (CNV) at the C4 locus. We find strong statistical significance for association of increased copy number of C4A (OR 0.81 (0.73; 0.89);P = 4.4 × 10(-5)), with the effect most pronounced in individuals over 78 years (OR 0.67 (0.55; 0.81)) and females (OR 0.77 (0.68; 0.87)). Furthermore, this association is independent of known AMD-associated risk variants in the nearby CFB/C2 locus, particularly in females and in individuals over 78 years. CONCLUSIONS: Our data strengthen the notion that complement dysregulation plays a crucial role in AMD etiology, an important finding for early intervention strategies and future therapeutics. In addition, for the first time, we provide evidence that multiallelic CNVs are associated with AMD pathology.

Pooled DNA based GWAS to determine genetic association of SNPs with visual acuity (VA) outcome in anti-vascular endothelial growth factor (anti-VEGF) treated neovascular age-related macular degeneration (nAMD) patients. We performed pooled DNA based GWAS on 285 anti-VEGF treated nAMD patients using high density Illumina 4.3 M array. Primary outcome was change in VA in Early Treatment Diabetic Retinopathy Study (ETDRS) letters after 6 months of anti-VEGF treatment (patients who lost ≥5 ETDRS letters classified as non-responders and all remaining classified as responders). GWAS analysis identified 44 SNPs of interest: 37 with strong evidence of association (p < 9 × 10-8), 2 in drug resistance genes (p < 5 × 10-6) and 5 nonsynonymous changes (p < 1 × 10-4). In the validation phase, individual genotyping of 44 variants showed three SNPs (rs4910623 p = 5.6 × 10-5, rs323085 p = 6.5 × 10-4 and rs10198937 p = 1.30 × 10-3) remained associated with VA response at 6 months. SNP rs4910623 also associated with treatment response at 3 months (p = 1.5 × 10-3). Replication of these three SNPs in 376 patients revealed association of rs4910623 with poor VA response after 3 and 6 months of treatment (p = 2.4 × 10-3 and p = 3.5 × 10-2, respectively). Meta-analysis of both cohorts (673 samples) confirmed association of rs4910623 with poor VA response after 3 months (p = 1.2 × 10-5) and 6 months (p = 9.3 × 10-6) of treatment in nAMD patients.

BACKGROUND: Graves' disease is an autoimmune thyroid disease of complex inheritance. Multiple genetic susceptibility loci are thought to be involved in Graves' disease and it is therefore likely that these can be identified by genome wide association studies. This study aimed to determine if a genome wide association study, using a pooling methodology, could detect genomic loci associated with Graves' disease. RESULTS: Nineteen of the top ranking single nucleotide polymorphisms including HLA-DQA1 and C6orf10, were clustered within the Major Histo-compatibility Complex region on chromosome 6p21, with rs1613056 reaching genome wide significance (p = 5 × 10-8). Technical validation of top ranking non-Major Histo-compatablity complex single nucleotide polymorphisms with individual genotyping in the discovery cohort revealed four single nucleotide polymorphisms with p ≤ 10-4. Rs17676303 on chromosome 1q23.1, located upstream of FCRL3, showed evidence of association with Graves' disease across the discovery, replication and combined cohorts. A second single nucleotide polymorphism rs9644119 downstream of DPYSL2 showed some evidence of association supported by finding in the replication cohort that warrants further study. CONCLUSIONS: Pooled genome wide association study identified a genetic variant upstream of FCRL3 as a susceptibility locus for Graves' disease in addition to those identified in the Major Histo-compatibility Complex. A second locus downstream of DPYSL2 is potentially a novel genetic variant in Graves' disease that requires further confirmation.

Myopia is the most common human eye disorder and it results from complex genetic and environmental causes. The rapidly increasing prevalence of myopia poses a major public health challenge. Here, the CREAM consortium performs a joint meta-analysis to test single-nucleotide polymorphism (SNP) main effects and SNP × education interaction effects on refractive error in 40,036 adults from 25 studies of European ancestry and 10,315 adults from 9 studies of Asian ancestry. In European ancestry individuals, we identify six novel loci (FAM150B-ACP1, LINC00340, FBN1, DIS3L-MAP2K1, ARID2-SNAT1 and SLC14A2) associated with refractive error. In Asian populations, three genome-wide significant loci AREG, GABRR1 and PDE10A also exhibit strong interactions with education (P<8.5 × 10(-5)), whereas the interactions are less evident in Europeans. The discovery of these loci represents an important advance in understanding how gene and environment interactions contribute to the heterogeneity of myopia.

Advanced age-related macular degeneration (AMD) is the leading cause of blindness in the elderly, with limited therapeutic options. Here we report on a study of >12 million variants, including 163,714 directly genotyped, mostly rare, protein-altering variants. Analyzing 16,144 patients and 17,832 controls, we identify 52 independently associated common and rare variants (P < 5 × 10(-8)) distributed across 34 loci. Although wet and dry AMD subtypes exhibit predominantly shared genetics, we identify the first genetic association signal specific to wet AMD, near MMP9 (difference P value = 4.1 × 10(-10)). Very rare coding variants (frequency <0.1%) in CFH, CFI and TIMP3 suggest causal roles for these genes, as does a splice variant in SLC16A8. Our results support the hypothesis that rare coding variants can pinpoint causal genes within known genetic loci and illustrate that applying the approach systematically to detect new loci requires extremely large sample sizes.

To identify genetic variants associated with refractive astigmatism in the general population, meta-analyses of genome-wide association studies were performed for: White Europeans aged at least 25 years (20 cohorts, N = 31,968); Asian subjects aged at least 25 years (7 cohorts, N = 9,295); White Europeans aged <25 years (4 cohorts, N = 5,640); and all independent individuals from the above three samples combined with a sample of Chinese subjects aged <25 years (N = 45,931). Participants were classified as cases with refractive astigmatism if the average cylinder power in their two eyes was at least 1.00 diopter and as controls otherwise. Genome-wide association analysis was carried out for each cohort separately using logistic regression. Meta-analysis was conducted using a fixed effects model. In the older European group the most strongly associated marker was downstream of the neurexin-1 (NRXN1) gene (rs1401327, P = 3.92E-8). No other region reached genome-wide significance, and association signals were lower for the younger European group and Asian group. In the meta-analysis of all cohorts, no marker reached genome-wide significance: The most strongly associated regions were, NRXN1 (rs1401327, P = 2.93E-07), TOX (rs7823467, P = 3.47E-07) and LINC00340 (rs12212674, P = 1.49E-06). For 34 markers identified in prior GWAS for spherical equivalent refractive error, the beta coefficients for genotype versus spherical equivalent, and genotype versus refractive astigmatism, were highly correlated (r = -0.59, P = 2.10E-04). This work revealed no consistent or strong genetic signals for refractive astigmatism; however, the TOX gene region previously identified in GWAS for spherical equivalent refractive error was the second most strongly associated region. Analysis of additional markers provided evidence supporting widespread genetic co-susceptibility for spherical and astigmatic refractive errors.

Keratoconus is typically diagnosed through changes at the anterior ocular surface. However, we wished to assess if macular parameter changes might also occur in these patients. We assessed posterior changes through the use of optical coherence tomography and compared to a nonkeratoconus patient group. All subjects underwent clinical examination including macular thickness measurements. The generalized estimation equation model was used to estimate the means and compare the differences in various measurements between keratoconus and nonkeratoconus patients. A total of 129 keratoconus eyes of 67 cases and 174 nonkeratoconus eyes of 87 controls were analysed. Keratoconus individuals presented with a significantly greater mean retinal thickness in the central fovea, inner, and outer macula compared to the nonkeratoconus group (p < 0.05). In addition, individuals presenting with the early signs of keratoconus had significantly greater inner and outer macular volume compared to the nonkeratoconus group (p < 0.05). This study indicates the retina appears to thicken at the fovea and macula and had increased macular volume in keratoconus individuals compared to nonkeratoconus individuals. Thus we posit that structural retinal changes exist in keratoconus eyes that are additional to those typically seen in the anterior segment.

PURPOSE: A previous study has indicated suggestive association of the hepatocyte growth factor (HGF) gene with Keratoconus. We wished to assess this association in an independent Caucasian cohort as well as assess its association with corneal curvature. PARTICIPANTS: Keratoconus patients were recruited from private and public clinics in Melbourne, Australia. Non-keratoconic individuals were identified from the Genes in Myopia (GEM) study from Australia. A total of 830 individuals were used for the analysis including 157 keratoconic and 673 non keratoconic subjects. METHODS: Tag single nucleotide polymorphisms (tSNPs) were chosen to encompass the hepatocyte growth factor gene as well as 2 kb upstream of the start codon through to 2 kb downstream of the stop codon. Logistic and linear regression including age and gender as covariates were applied in statistical analysis with subsequent Bonferroni correction. RESULTS: Ten tSNPs were genotyped. Following statistical analysis and multiple testing correction, a statistically significant association was found for the tSNP rs2286194 {p = 1.1×10-(3) Odds Ratio 0.52, 95% CI--0.35, 0.77} for keratoconus. No association was found between the 10 tSNPs and corneal curvature. CONCLUSIONS: These findings provide additional evidence of significant association of the HGF gene with Keratoconus. This association does not appear to act through the corneal curvature route.

BACKGROUND: Intrachromosomal segmental duplications provide the substrate for non-allelic homologous recombination, facilitating extensive copy number variation in the human genome. Many multi-copy gene families are embedded within genomic regions with high levels of sequence identity (>95%) and therefore pose considerable analytical challenges. In some cases, the complexity involved in analyzing such regions is largely underestimated. Rapid, cost effective analysis of multi-copy gene regions have typically implemented quantitative approaches, however quantitative data are not an absolute means of certainty. Therefore any technique prone to degrees of measurement error can produce ambiguous results that may lead to spurious associations with complex disease. RESULTS: In this study we have focused on testing the accuracy and reproducibility of quantitative analysis techniques. With reference to the C-C Chemokine Ligand-3-like-1 (CCL3L1) gene, we performed analysis using real-time Quantitative PCR (QPCR), Multiplex Ligation-dependent Probe Amplification (MLPA) and Paralogue Ratio Test (PRT). After controlling for potential outside variables on assay performance, including DNA concentration, quality, preparation and storage conditions, we find that real-time QPCR produces data that does not cluster tightly around copy number integer values, with variation substantially greater than that of the MLPA or PRT systems. We find that the method of rounding real-time QPCR measurements can potentially lead to mis-scoring of copy number genotypes and suggest caution should be exercised in interpreting QPCR data. CONCLUSIONS: We conclude that real-time QPCR is inherently prone to measurement error, even under conditions that would seem favorable for association studies. Our results indicate that potential variability in the physicochemical properties of the DNA samples cannot solely explain the poor performance exhibited by the real-time QPCR systems. We recommend that more robus

Refractive error (RE) is a complex, multifactorial disorder characterized by a mismatch between the optical power of the eye and its axial length that causes object images to be focused off the retina. The two major subtypes of RE are myopia (nearsightedness) and hyperopia (farsightedness), which represent opposite ends of the distribution of the quantitative measure of spherical refraction. We performed a fixed effects meta-analysis of genome-wide association results of myopia and hyperopia from 9 studies of European-derived populations: AREDS, KORA, FES, OGP-Talana, MESA, RSI, RSII, RSIII and ERF. One genome-wide significant region was observed for myopia, corresponding to a previously identified myopia locus on 8q12 (p = 1.25×10(-8)), which has been reported by Kiefer et al. as significantly associated with myopia age at onset and Verhoeven et al. as significantly associated to mean spherical-equivalent (MSE) refractive error. We observed two genome-wide significant associations with hyperopia. These regions overlapped with loci on 15q14 (minimum p value = 9.11×10(-11)) and 8q12 (minimum p value 1.82×10(-11)) previously reported for MSE and myopia age at onset. We also used an intermarker linkage- disequilibrium-based method for calculating the effective number of tests in targeted regional replication analyses. We analyzed myopia (which represents the closest phenotype in our data to the one used by Kiefer et al.) and showed replication of 10 additional loci associated with myopia previously reported by Kiefer et al. This is the first replication of these loci using myopia as the trait under analysis. "Replication-level" association was also seen between hyperopia and 12 of Kiefer et al.'s published loci. For the loci that show evidence of association to both myopia and hyperopia, the estimated effect of the risk alleles were in opposite directions for the two traits. This suggests that these loci are important contributors to variation of refractive error acros

Refractive error (RE) is a complex, multifactorial disorder characterized by a mismatch between the optical power of the eye and its axial length that causes object images to be focused off the retina. The two major subtypes of RE are myopia (nearsightedness) and hyperopia (farsightedness), which represent opposite ends of the distribution of the quantitative measure of spherical refraction. We performed a fixed effects meta-analysis of genome-wide association results of myopia and hyperopia from 9 studies of European-derived populations: AREDS, KORA, FES, OGP-Talana, MESA, RSI, RSII, RSIII and ERF. One genome-wide significant region was observed for myopia, corresponding to a previously identified myopia locus on 8q12 (p = 1.25x10(-8)), which has been reported by Kiefer et al. as significantly associated with myopia age at onset and Verhoeven et al. as significantly associated to mean spherical-equivalent (MSE) refractive error. We observed two genome-wide significant associations with hyperopia. These regions overlapped with loci on 15q14 (minimum p value = 9.11x10(-11)) and 8q12 (minimum p value 1.82x10(-11)) previously reported for MSE and myopia age at onset. We also used an intermarker linkage-disequilibrium-based method for calculating the effective number of tests in targeted regional replication analyses. We analyzed myopia (which represents the closest phenotype in our data to the one used by Kiefer et al.) and showed replication of 10 additional loci associated with myopia previously reported by Kiefer et al. This is the first replication of these loci using myopia as the trait under analysis. "Replication-level'' association was also seen between hyperopia and 12 of Kiefer et al.'s published loci. For the loci that show evidence of association to both myopia and hyperopia, the estimated effect of the risk alleles were in opposite directions for the two traits. This suggests that these loci are important contributors to variation of refractive error acros

BACKGROUND: HMG Co-A reductase inhibitors are ubiquitous in our community yet their potential role in age-related macular degeneration (AMD) remains to be determined. OBJECTIVES: To evaluate the effect of simvastatin on AMD progression and the effect modification by polymorphism in apolipoprotein E (ApoE) and complement factor H (CFH) genes. DESIGN: A proof of concept double-masked randomized controlled study. PARTICIPANTS: 114 participants aged 53 to 91 years, with either bilateral intermediate AMD or unilateral non-advanced AMD (with advanced AMD in fellow eye), BCVA ≥ 20/60 in at least one eye, and a normal lipid profile. INTERVENTION: Simvastatin 40 mg/day or placebo, allocated 1:1. MAIN OUTCOME MEASURES: Progression of AMD either to advanced AMD or in severity of non-advanced AMD. Results. The cumulative AMD progression rates were 70% in the placebo and 54% in the simvastatin group. Intent to treat multivariable logistic regression analysis, adjusted for age, sex, smoking and baseline AMD severity, showed a significant 2-fold decrease in the risk of progression in the simvastatin group: OR 0.43 (0.18-0.99), p = 0.047. Post-hoc analysis stratified by baseline AMD severity showed no benefit from treatment in those who had advanced AMD in the fellow eye before enrolment: OR 0.97 (0.27-3.52), p = 0.96, after adjusting for age, sex and smoking. However, there was a significant reduction in the risk of progression in the bilateral intermediate AMD group compared to placebo [adjusted OR 0.23 (0.07-0.75), p = 0.015]. The most prominent effect was observed amongst those who had the CC (Y402H) at risk genotype of the CFH gene [OR 0.08 (0.02-0.45), p = 0.004]. No evidence of harm from simvastatin intervention was detected. CONCLUSION/SIGNIFICANCE: Simvastatin may slow progression of non-advanced AMD, especially for those with the at risk CFH genotype CC (Y402H). Further exploration of the potential use of statins for AMD, with emphasis on genetic subgroups, is warranted.

Macular degeneration is a common cause of blindness in the elderly. To identify rare coding variants associated with a large increase in risk of age-related macular degeneration (AMD), we sequenced 2,335 cases and 789 controls in 10 candidate loci (57 genes). To increase power, we augmented our control set with ancestry-matched exome-sequenced controls. An analysis of coding variation in 2,268 AMD cases and 2,268 ancestry-matched controls identified 2 large-effect rare variants: previously described p.Arg1210Cys encoded in the CFH gene (case frequency (fcase) = 0.51%; control frequency (fcontrol) = 0.02%; odds ratio (OR) = 23.11) and newly identified p.Lys155Gln encoded in the C3 gene (fcase = 1.06%; fcontrol = 0.39%; OR = 2.68). The variants suggest decreased inhibition of C3 by complement factor H, resulting in increased activation of the alternative complement pathway, as a key component of disease biology.

Narrow arterioles in the retina have been shown to predict hypertension as well as other vascular diseases, likely through an increase in the peripheral resistance of the microcirculatory flow. In this study, we performed a genome-wide association study in 18,722 unrelated individuals of European ancestry from the Cohorts for Heart and Aging Research in Genomic Epidemiology consortium and the Blue Mountain Eye Study, to identify genetic determinants associated with variations in retinal arteriolar caliber. Retinal vascular calibers were measured on digitized retinal photographs using a standardized protocol. One variant (rs2194025 on chromosome 5q14 near the myocyte enhancer factor 2C MEF2C gene) was associated with retinal arteriolar caliber in the meta-analysis of the discovery cohorts at genome-wide significance of P-value <5×10(-8). This variant was replicated in an additional 3,939 individuals of European ancestry from the Australian Twins Study and Multi-Ethnic Study of Atherosclerosis (rs2194025, P-value = 2.11×10(-12) in combined meta-analysis of discovery and replication cohorts). In independent studies of modest sample sizes, no significant association was found between this variant and clinical outcomes including coronary artery disease, stroke, myocardial infarction or hypertension. In conclusion, we found one novel loci which underlie genetic variation in microvasculature which may be relevant to vascular disease. The relevance of these findings to clinical outcomes remains to be determined.

Age-related macular degeneration (AMD) is a leading cause of blindness in Western countries and is diagnosed by the clinical appearance of yellow subretinal deposits called drusen. Genetic changes in immune components are clearly implicated in the pathology of this disease. We have previously shown that the purinergic receptor P2X7 can act as a scavenger receptor, mediating phagocytosis of apoptotic cells and insoluble debris. We performed a genetic association study of functional polymorphisms in the P2RX7 and P2RX4 genes in a cohort of 744 patients with AMD and 557 age-matched Caucasian control subjects. The P2X4 Tyr315Cys variant was 2-fold more frequent in patients with AMD compared to control subjects, with the minor allele predicting susceptibility to disease. Pairwise linkage disequilibrium was observed between Tyr315Cys in the P2RX4 gene and Gly150Arg in the P2RX7 gene, and these two minor alleles formed a rare haplotype that was overrepresented in patients with AMD (n=17) compared with control subjects (n=3) (odds ratio 4.05, P=0.026). Expression of P2X7 (wild type or variant 150Arg) in HEK293 cells conferred robust phagocytosis toward latex beads, whereas coexpression of the P2X7 150Arg with P2X4 315Cys variants almost completely inhibited phagocytic capacity. Fresh human monocytes harboring this heterozygous 150Arg-315Cys haplotype showed 40% reduction in bead phagocytosis. In the primate eye, immunohistochemistry indicated that P2X7 and P2X4 receptors were coexpressed on microglia and macrophages, but neither receptor was seen on retinal pigment epithelial cells. These results demonstrate that a haplotype including two rare variants in P2RX7 and P2RX4 confers a functional interaction between these two variant receptors that impairs the normal scavenger function of macrophages and microglia. Failure of this P2X7-mediated phagocytic pathway may impair removal of subretinal deposits and predispose individuals toward AMD.

Age-related macular degeneration (AMD) is a common cause of blindness in older individuals. To accelerate the understanding of AMD biology and help design new therapies, we executed a collaborative genome-wide association study, including >17,100 advanced AMD cases and >60,000 controls of European and Asian ancestry. We identified 19 loci associated at P < 5 × 10(-8). These loci show enrichment for genes involved in the regulation of complement activity, lipid metabolism, extracellular matrix remodeling and angiogenesis. Our results include seven loci with associations reaching P < 5 × 10(-8) for the first time, near the genes COL8A1-FILIP1L, IER3-DDR1, SLC16A8, TGFBR1, RAD51B, ADAMTS9 and B3GALTL. A genetic risk score combining SNP genotypes from all loci showed similar ability to distinguish cases and controls in all samples examined. Our findings provide new directions for biological, genetic and therapeutic studies of AMD.