Biomarkers For Age-related Macular Degeneration Research Articles

MicroRNAs as diagnostic and prognostic biomarkers of age-related macular degeneration: advances and limitations.

A main cause of vision loss in the elderly is age-related macular degeneration (AMD). Among the cellular, biochemical, and molecular changes linked to this disease, inflammation and angiogenesis appear as being crucial in AMD pathogenesis and progression. There are two forms of the disease: dry AMD, accounting for 80–90% of cases, and wet AMD. The disease usually begins as dry AMD associated with retinal pigment epithelium and photoreceptor degeneration, whereas wet AMD is associated with choroidal neovascularization resulting in severe vision impairment. The new vessels are largely malformed, leading to blood and fluid leakage within the disrupted tissue, which provokes inflammation and scar formation and results in retinal damage and detachment. MicroRNAs are dysregulated in AMD and may facilitate the early detection of the disease and monitoring disease progression. Two recent reviews of microRNAs in AMD had indicated weaknesses or limitations in four earlier investigations. Studies in the last three years have shown considerable progress in overcoming some of these concerns and identifying specific microRNAs as biomarkers for AMD. Further large-scale studies are warranted using appropriate statistical methods to take into account gender and age disparity in the study populations and confounding factors such as smoking status.

Association of imaging biomarkers and local activation of complement in aqueous humor of patients with early forms of age-related macular degeneration

PurposeTo investigate a possible correlation between established imaging biomarkers for age-related macular degeneration and local complement system activation, measured in aqueous humor (AH) of patients with early stages of age-related macular degeneration (AMD) and controls.MethodsThis analysis included prospectively acquired AH samples of 106 eyes (35 with early/intermediate AMD, 71 controls). The levels of complement protein 3 (C3), 4 (C4), 5 (C5); activation products of complement factor 3a (C3a) and Ba, C3b/iC3b; complement factors B, D, H, I (CFB, CFD, CFH, CFI); and total protein concentration were analyzed. Quantitative levels of complement factors were correlated to the presence of reticular pseudodrusen (RPD), the presence of hyperreflective foci (HRF), and total drusen volume (DV) graded on imaging by spectral-domain optical coherence tomography and using Spearman’s rank correlation test.ResultsDV correlated with C3b/iC3b (r = 0.285; P = 0.034), C3a (r = 0.200; P = 0.047), Ba (r = 0.262; P = 0.009), and C5 (r = 430; P = 0.005), and showed a tendency towards correlation with C3a (r = 0.198; P = 0.057). HRF correlated significantly with C5 (r = 0.388; P = 0.011) and RPD showed a tendency towards correlation with CFB (r = 0.196; P = 0.050).ConclusionIn patients with early AMD, HRF and drusen parameters but not RPD show low to fair levels of correlation with local complement activation in patients’ AH. Better understanding of complement activation could provide some insights into the pathogenesis of AMD. Imaging biomarkers could be useful to identify suitable patients for future clinical trials with complement-modulating therapies.

Macular Pigment Optical Density and Photoreceptor Outer Segment Length as Predisease Biomarkers for Age-Related Macular Degeneration.

To explore predisease biomarkers, which may help screen for the risk of age-related macular degeneration (AMD) at very early stages, macular pigment optical density (MPOD) and photoreceptor outer segment (PROS) length were analyzed. Thirty late AMD fellow eyes, which are at high risk and represent the predisease condition of AMD, were evaluated and compared with 30 age-matched control eyes without retinal diseases; there was no early AMD involvement in the AMD fellow eyes. MPOD was measured using MPS2® (M.E. Technica Co. Ltd., Tokyo, Japan), and PROS length was measured based on optical coherence tomography images. MPOD levels and PROS length in the AMD fellow eyes were significantly lower and shorter, respectively, than in control eyes. MPOD and PROS length were positively correlated in control eyes (R = 0.386; p = 0.035) but not in AMD fellow eyes. Twenty (67%) AMD fellow eyes met the criteria of MPOD < 0.65 and/or PROS length < 35 μm, while only five (17%) control eyes did. After adjusting for age and sex, AMD fellow eyes more frequently satisfied the definition (p < 0.001; 95% confidence interval, 3.50–60.4; odds ratio, 14.6). The combination of MPOD and PROS length may be a useful biomarker for screening predisease AMD patients, although further studies are required in this regard.

Identification of Novel Serum MicroRNAs in Age-Related Macular Degeneration.

PurposeTo identify circulating microRNAs (miRNA) associated with age-related macular degeneration (AMD). Thus differentially expressed serum miRNA could be used as AMD biomarkers.MethodsThis study involved total RNA isolation from sera from patients with atrophic AMD (n = 10), neovascular AMD (n = 10), and age- and sex-matched controls (n = 10). A total of 377 miRNAs were coanalyzed using array technologies, and differentially regulated miRNAs were determined. Extensive validation studies (n = 90) of serum from AMD patients and controls confirmed initial results. Total RNA isolation was carried out from sera from patients with atrophic AMD (n = 30), neovascular AMD (n = 30), and controls (n = 30). Fourteen miRNAs from the discovery dataset were coanalyzed using quantitative real-time polymerase chain reaction (qRT-PCR) to validate their presence.ResultsUnsupervised hierarchical clustering indicated that AMD serum specimens have a different miRNA profile to healthy controls. We successfully identified and validated the differentially regulated miRNAs in serum from AMD patients versus controls. The biomarker potential of three miRNAs (miR-126, miR-19a, and miR-410) was confirmed by qRT-PCR, with significantly increased quantities in serum of AMD patients compared with healthy controls.ConclusionsIncreased quantities of miR-126, miR-410, and miR-19a in serum from AMD patients indicate that these miRNAs could potentially serve as diagnostic AMD biomarkers. All three miRNAs significantly correlated with AMD pathogenesis.Translational RelevanceThe discovery of new AMD miRNA may act as biomarkers in evaluating AMD diagnosis and prognosis.

4-Hydroxy-7-oxo-5-heptenoic acid (HOHA) lactone induces apoptosis in retinal pigment epithelial cells

Retinal pigment epithelial (RPE) cell dysfunction and death play vital roles in age-related macular degeneration (AMD) pathogenesis. Previously we showed that oxidative cleavage of docosahexenoate (DHA) phospholipids generates an α,β-unsaturated aldehyde, 4-hydroxy-7-oxohept-4-enoic acid (HOHA) lactone, that forms ω-carboxyethylpyrrole (CEP) derivatives through adduction to proteins and ethanolamine phospholipids. CEP derivatives and autoantibodies accumulate in the retinas and blood plasma of individuals with AMD and are a biomarker of AMD. They promote the choroidal neovascularization of “wet AMD”. Immunization of mice with CEP-modified mouse serum albumin induces “dry AMD”-like lesions in their retinas as well as interferon-gamma and interleukin-17 production by CEP-specific T cells that promote inflammatory M1 polarization of macrophages. The present study confirms that oxidative stress or inflammatory stimulus produces CEP in both the primary human ARPE-19 cell line and hRPE cells. Exposure of these cells to HOHA lactone fosters production of reactive oxygen species. Thus, HOHA lactone participates in a vicious cycle, promoting intracellular oxidative stress leading to oxidative cleavage of DHA to produce more HOHA lactone. We now show that HOHA lactone is cytotoxic, inducing apoptotic cell death through activation of the intrinsic pathway. This suggests that therapeutic interventions targeting HOHA lactone-induced apoptosis may prevent the loss of RPE cells during the early phase of AMD. We also discovered that ARPE-19 cells are more susceptible than hRPE cells to HOHA lactone cytotoxicity. This is consistent with the view that, compared to normal RPE cells, ARPE-19 cells exhibit a diseased RPE phenotype that also includes elevated expression of the mesenchymal indicator vimentin, elevated integrin a5 promotor strength and deficient secretion of the anti-VEGF molecule pigment-epithelium-derived factor fostering weaker tight junctions.

Selecting likely causal risk factors from high-throughput experiments using multivariable Mendelian randomization

Modern high-throughput experiments provide a rich resource to investigate causal determinants of disease risk. Mendelian randomization (MR) is the use of genetic variants as instrumental variables to infer the causal effect of a specific risk factor on an outcome. Multivariable MR is an extension of the standard MR framework to consider multiple potential risk factors in a single model. However, current implementations of multivariable MR use standard linear regression and hence perform poorly with many risk factors. Here, we propose a two-sample multivariable MR approach based on Bayesian model averaging (MR-BMA) that scales to high-throughput experiments. In a realistic simulation study, we show that MR-BMA can detect true causal risk factors even when the candidate risk factors are highly correlated. We illustrate MR-BMA by analysing publicly-available summarized data on metabolites to prioritise likely causal biomarkers for age-related macular degeneration.

Associations of microRNAs, Angiogenesis-Regulating Factors and CFH Y402H Polymorphism-An Attempt to Search for Systemic Biomarkers in Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) remains the leading cause of blindness in elderly people, but the pathophysiology of this disease is still largely unknown. We investigated the systemic expression of angiogenesis-regulating growth factors and selected miRNAs known to regulate angiogenesis in AMD patients. We also focused on possible correlations of their expression with the presence of CFH Y402H or ARMS A69S risk variants. A total of 354 AMD patients and 121 controls were enrolled in this study. The levels of angiogenesis-regulating factors were analyzed in plasma samples using Luminex technology. The expression of selected miRNAs was analyzed in peripheral blood plasma using real-time qPCR. The genetic analysis was performed with an Illumina NextSeq500 system. AMD was an independent factor associated with lower levels of angiogenin (β = −0.29, p < 0.001), endostatin (β = −0.18, p < 0.001), FGF-basic (β = −0.18, p < 0.001), PlGF (β = −0.24, p < 0.001), miRNA-21-3p (β = −0.13, p = 0.01) and miRNA-155-5p (β = −0.16, p = 0.002); and with higher levels of FGF-acidic (β = 0.11, p = 0.03), miRNA-23a-3p (β = 0.17, p < 0.001), miRNA-126-5p (β = 0.13, p = 0.009), miRNA-16-5p (β = 0.40, p < 0.001), miRNA-17-3p (β = 0.13, p = 0.01), miRNA-17-5p (β = 0.17, p < 0.001), miRNA-223-3p (β = 0.15, p = 0.004), and miRNA-93 (β = 0.11, p = 0.04). The expression of analyzed miRNA molecules significantly correlated with the levels of tested angiogenesis-regulating factors and clinical parameters in AMD patients, whereas such correlations were not observed in controls. We also found an association between the CFH Y402H polymorphism and miRNA profiles, whereby TT homozygotes showed evidently higher expression of miRNA-16-5p than CC homozygotes or TC heterozygotes (p = 0.0007). Our results suggest that the balance between systemic pro- and anti-angiogenic factors and miRNAs is vital in multifactorial AMD pathogenesis.

Automated quantification of choroidal neovascularization on Optical Coherence Tomography Angiography images

ObjectivesTo report the design of an automated quantification algorithm for choroidal neovascularization (CNV) in the context of neovascular age-related macular degeneration (AMD), based on Optical Coherence Tomography Angiography (OCTA) images. Material and methodsIn this study, 54 patients (mean age 75.80 ± 14.29 years) with neovascular AMD (type 1 and type 2 CNV) were included retrospectively and separated into two groups (Group 1–24 images; Group 2–30 images), according to the lesion topology. All patients underwent a 3 × 3 mm OCTA examination (AngioVue, Optovue, Freemont, California). The proposed algorithm is based on segmentation and enhancement methods including Frangi filter, Gabor wavelets and Fuzzy-C-Means Classification. Our results were compared to the manual quantifications given by the embedded quantification software “AngioAnalytics”. ResultsAutomated CNV segmentation and quantification of three neovascular AMD biomarkers: the total vascular area (TVA), the total area (TA) and the vascular density (VD) were possible in all cases. Automated versus manual quantification comparison revealed a statistically significant difference for TVA and VD measurements for both groups (p = 0.00036 for Group 1 TVA, p < 0.0001 for Group 1 VD and Group 2 TVA and VD). The difference in TA measurements was not significant in Group 2 (p = 0.143). Bland-Altman analysis revealed low inter-method bias for TA measurements and higher bias for TVA and VD. ConclusionThis paper presents a method for segmenting and quantifying CNV that constitutes a valid option for clinicians. Complementary validations have to be carried out to compare our method's accuracy to “AngioAnalytics”.

Automated detection and classification of early AMD biomarkers using deep learning

Age-related macular degeneration (AMD) affects millions of people and is a leading cause of blindness throughout the world. Ideally, affected individuals would be identified at an early stage before late sequelae such as outer retinal atrophy or exudative neovascular membranes develop, which could produce irreversible visual loss. Early identification could allow patients to be staged and appropriate monitoring intervals to be established. Accurate staging of earlier AMD stages could also facilitate the development of new preventative therapeutics. However, accurate and precise staging of AMD, particularly using newer optical coherence tomography (OCT)-based biomarkers may be time-intensive and requires expert training which may not feasible in many circumstances, particularly in screening settings. In this work we develop deep learning method for automated detection and classification of early AMD OCT biomarker. Deep convolution neural networks (CNN) were explicitly trained for performing automated detection and classification of hyperreflective foci, hyporeflective foci within the drusen, and subretinal drusenoid deposits from OCT B-scans. Numerous experiments were conducted to evaluate the performance of several state-of-the-art CNNs and different transfer learning protocols on an image dataset containing approximately 20000 OCT B-scans from 153 patients. An overall accuracy of 87% for identifying the presence of early AMD biomarkers was achieved.

Metabolomics in serum of patients with non-advanced age-related macular degeneration reveals aberrations in the glutamine pathway.

Age-related macular degeneration (AMD) is a common, progressive multifactorial vision-threatening disease and many genetic and environmental risk factors have been identified. The risk of AMD is influenced by lifestyle and diet, which may be reflected by an altered metabolic profile. Therefore, measurements of metabolites could identify biomarkers for AMD, and could aid in identifying high-risk individuals. Hypothesis-free technologies such as metabolomics have a great potential to uncover biomarkers or pathways that contribute to disease pathophysiology. To date, only a limited number of metabolomic studies have been performed in AMD. Here, we aim to contribute to the discovery of novel biomarkers and metabolic pathways for AMD using a targeted metabolomics approach of 188 metabolites. This study focuses on non-advanced AMD, since there is a need for biomarkers for the early stages of disease before severe visual loss has occurred. Targeted metabolomics was performed in 72 patients with early or intermediate AMD and 72 control individuals, and metabolites predictive for AMD were identified by a sparse partial least squares discriminant analysis. In our cohort, we identified four metabolite variables that were most predictive for early and intermediate stages of AMD. Increased glutamine and phosphatidylcholine diacyl C28:1 levels were detected in non-advanced AMD cases compared to controls, while the rate of glutaminolysis and the glutamine to glutamate ratio were reduced in non-advanced AMD. The association of glutamine with non-advanced AMD corroborates a recent report demonstrating an elevated glutamine level in early AMD using a different metabolomics technique. In conclusion, this study indicates that metabolomics is a suitable method for the discovery of biomarker candidates for AMD. In the future, larger metabolomics studies could add to the discovery of novel biomarkers in yet unknown AMD pathways and expand our insights in AMD pathophysiology.

Identification of aberrantly methylated differentially expressed genes in age-related macular degeneration

DNA methylation plays a significant role in many diseases. Age-related macular degeneration (AMD) is a leading cause of vision loss for people aged 50 years and above, but the etiology and pathogenesis are largely unknown. This study aimed to identify the aberrantly methylated differentially expressed genes (DEGs) in AMD and predict the related pathways on the basis of public data.Aberrant methylation can influence the functions of key genes by altering their expression. Here, we found out DEGs by overlapping public microarray data (GSE29801 and GSE102952). Functional and enrichment analyses of selected genes were performed using the DAVID database. Subsequently, protein–protein interaction (PPI) networks were constructed by using STRING and visualized in cytoscape to determine hub genes. Finally, we collected AMD patients’ blood samples to identify the methylation statuses of these hub genes by using methylated DNA immunoprecipitation.In total, 156 hypermethylation-low expression genes and 127 hypomethylation-high expression genes were predicted. The hypermethylation-low expression genes were enriched in biological processes of response to cardiac conduction, ATP binding, and cell–cell junction assembly. The top 5 hub genes of the PPI network were HSP90AA1, HSPA1L, HSPE1, HSP90B1, and NOP56. Meanwhile, the hypomethylation-high expression genes were enriched in the biological processes of response to positive regulation of the MAPK cascade, actin cytoskeleton reorganization, dentate gyrus development, and cell migration. The top 5 hub genes of this PPI network were PIK3R1, EZR, IGF2, SLC2A1, and CDKN1C. Moreover, the methylation statuses of NOP56, EZR, IGF2, SLC2A1, CDKN1C were confirmed to be altered in the blood of AMD patients.This study indicated possible aberrantly methylated DEGs and differentially expressed pathways in AMD by bioinformatics analysis, providing novel insights for unraveling the pathogenesis of AMD. Hub genes, including NOP56, EZR, IGF2, SLC2A1, CDKN1C, might serve as aberrant methylation-based candidate biomarkers for AMD in future applications.

Characterization of SIRT1/DNMTs Functions and LINE-1 Methylation in Patients with Age-Related Macular Degeneration.

Previous studies proposed the application of DNA methylation signatures as clinical biomarkers of age-related macular degeneration (AMD). However, the characterization of Long Interspersed Nuclear Element-1 (LINE-1) methylation levels—a surrogate marker of global DNA methylation—in AMD patients has not been investigated so far. In the present study, we first characterized DNA methyltransferases (DNMTs) and Sirtuin 1 (SIRT1) functions in blood samples of 40 AMD patients and 10 age- and sex-matched controls. Then, we evaluated whether changes in DNMTs functions were associated with different LINE-1 methylation levels in leukocyte DNA. We demonstrated that total DNMTs activity was 48% higher in AMD patients than in controls (p = 0.005). AMD patients also exhibited up-regulation of DNMT1 and DNMT3B expression (FC = 2.6; p = 0.003 and FC = 2.4; p = 0.018, respectively). In line with increased DNMTs functions, the LINE-1 methylation level was higher in AMD patients than in controls (mean = 69.10%; SE = 0.68 vs. mean = 65.73%; SE = 0.59; p = 0.020). All p-values were adjusted by Bonferroni correction. In AMD patients, LINE-1 methylation level was positively associated with total DNMTs activity (r = 0.694; p < 0.001), DNMT1 (r = 0.579; p < 0.001), and DNMT3B (r = 0.521; p = 0.001) expression. Our results encourage further large-size prospective research to understand the relationship between LINE-1 methylation and AMD aetiology, and its usefulness in the clinical setting.

Fluorescence Lifetime Imaging of Tetracycline-Stained Retinal Hydroxyapatite: An Early Biomarker for Age-Related Macular Degeneration?

Fluorescence Lifetime Imaging of Tetracycline-Stained Retinal Hydroxyapatite: An Early Biomarker for Age-Related Macular Degeneration?

Urine Nuclear Magnetic Resonance (NMR) Metabolomics in Age-Related Macular Degeneration.

Biofluid biomarkers of age-related macular degeneration (AMD) are still lacking, and their identification is challenging. Metabolomics is well-suited to address this need, and urine is a valuable accessible biofluid. This study aimed to characterize the urinary metabolomic signatures of patients with different stages of AMD and a control group (>50 years). It was a prospective, cross-sectional study, where subjects from two cohorts were included: 305 from Coimbra, Portugal (AMD patients n = 252; controls n = 53) and 194 from Boston, United States (AMD patients n = 147; controls n = 47). For all participants, we obtained color fundus photographs (for AMD staging) and fasting urine samples, which were analyzed using 1H nuclear magnetic resonance (NMR) spectroscopy. Our results revealed that in both cohorts, urinary metabolomic profiles differed mostly between controls and late AMD patients, but important differences were also found between controls and subjects with early AMD. Analysis of the metabolites responsible for these separations revealed that, even though distinct features were observed for each cohort, AMD was in general associated with depletion of excreted citrate and selected amino acids at some stage of the disease, suggesting enhanced energy requirements. In conclusion, NMR metabolomics enabled the identification of urinary signals of AMD and its severity stages, which might represent potential metabolomic biomarkers of the disease.

Isolation of Retinal Exosome Biomarkers from Blood byTargeted Immunocapture.

The retinal pigmented epithelium (RPE) forms the outer blood-retinal barrier, provides nutrients, recycles visual pigment, and removes spent discs from the photoreceptors, among many other functions. Because of these critical roles in visual homeostasis, the RPE is a principal location of disease-associated changes in age-related macular degeneration (AMD), emphasizing its importance for study in both visual health and disease. Unfortunately, there are no early indicators of AMD or disease progression, a void that could be filled by the development of early AMD biomarkers. Exosomes are lipid bilayer membrane vesicles of nanoscale sizes that are released in a controlled fashion by cells and carry out a number of extra- and intercellular activities. In the RPE they are released from both the apical and basal sides, and each source has a unique signature/content. Exosomes released from the basolateral side of RPE cells enter the systemic circulation via the choroid and thus represent a potential source of retinal disease biomarkers in blood. Here we discuss the potential of targeted immunocapture of eye-derived exosomes and other small extracellular vesicles from blood for eye disease biomarker discovery.

In vitro drusen model - three-dimensional spheroid culture of retinal pigment epithelial cells.

Age-related macular degeneration (AMD) is a leading cause of blindness in people over 50 years of age in many developed countries. Drusen are yellowish extracellular deposits beneath retinal pigment epithelium (RPE) found in aging eyes and considered as a biomarker of AMD. However, the biogenesis of drusen has not been elucidated. We reported previously that multicellular spheroids of human RPE cells constructed a well-differentiated monolayer of RPE with a Bruch's membrane. We determined that RPE spheroids exhibited drusen formation between the RPE and Bruch's membrane with expression of many drusen-associated proteins, such as amyloid β and complement components, the expression of which was altered by a challenge with oxidative stress. Artificial lipofuscin-loaded RPE spheroids yielded drusen more frequently. In the current study, we showed that drusen originates from the RPE. This culture system is an attractive tool for use as an in vitro drusen model, which might help elucidate the biogenesis of drusen and the pathogenesis of related diseases, such as AMD.

Complete Blood Cell Count–Derived Inflammation Biomarkers in Men with Age-Related Macular Degeneration

ABSTRACTPurpose: To investigate the role of some blood count–derived inflammation biomarkers in age-related macular degeneration (AMD).Methods: Seventy-nine men with late-stage AMD and 79 male age-matched cataract controls without AMD were recruited in March–December, 2016. A blood sample was taken. The following blood cell count–derived indexes were evaluated: neutrophil/lymphocyte ratio (NLR), derived NLR [dNLR = neutrophils/(white blood cells ‒ neutrophils)], platelet/lymphocyte ratio (PLR), monocyte/lymphocyte ratio (MLR), (neutrophils × monocytes)/lymphocyte ratio (SIRI), and (neutrophils × monocytes × platelets)/lymphocyte ratio (AISI).Results: AMD patients had significantly lower median values of white blood cells, monocytes, neutrophils, platelets, and mean platelet volume (MPV). Regarding the combined indexes, only AISI was significantly lower in AMD patients than in controls. Receiver operating characteristics curve analysis revealed that the ability of AISI and MPV to predict AMD is poor.Conclusion: Results suggests that NLR, dNLR, PLR, MLR, SIRI, and AISI are unreliable disease biomarkers in men with AMD. Larger scale studies are necessary to confirm these findings.

Human Plasma Metabolomics Study across All Stages of Age-Related Macular Degeneration Identifies Potential Lipid Biomarkers

To characterize the plasma metabolomic profile of patients with age-related macular degeneration (AMD) using mass spectrometry (MS). Cross-sectional observational study. We prospectively recruited participants with a diagnosis of AMD and a control group (>50years of age) without any vitreoretinal disease. All participants underwent color fundus photography, used for AMD diagnosis and staging, according to the Age-Related Eye Disease Study classification scheme. Fasting blood samples were collected and plasma was analyzed by Metabolon, Inc. (Durham, NC), using ultrahigh-performance liquid chromatography (UPLC) and high-resolution MS. Metabolon's hardware and software were used to identify peaks and control quality. Principal component analysis and multivariate regression were performed to assess differences in the metabolomic profiles of AMD patients versus controls, while controlling for potential confounders. For biological interpretation, pathway enrichment analysis of significant metabolites was performed using MetaboAnalyst. The primary outcome measures were levels of plasma metabolites in participants with AMD compared with controls and among different AMD severity stages. We included 90 participants with AMD (30 with early AMD, 30 with intermediate AMD, and 30 with late AMD) and 30 controls. Using UPLC and MS, 878 biochemicals were identified. Multivariate logistic regression identified 87 metabolites with levels that differed significantly between AMD patients and controls. Most of these metabolites (82.8%; n= 72), including the most significant metabolites, belonged to the lipid pathways. Analysis of variance revealed that of the 87 metabolites, 48 (55.2%) also were significantly different across thedifferent stages of AMD. A significant enrichment of the glycerophospholipids pathway was identified (P=4.7× 10-9) among these metabolites. Participants with AMD have altered plasma metabolomic profiles compared with controls. Our data suggest that the most significant metabolites map to the glycerophospholipid pathway. These findings have the potential to improve our understanding of AMD pathogenesis, to support the development of plasma-based metabolomics biomarkers of AMD, and to identify novel targets for treatment of this blinding disease.

Algorithms for the Automated Analysis of Age-Related Macular Degeneration Biomarkers on Optical Coherence Tomography: A Systematic Review.

PurposeTo assess the quality of optical coherence tomography (OCT) grading algorithms for retinal biomarkers of age-related macular degeneration (AMD).MethodsFollowing a systematic review of the literature data on detection and quantification of AMD retinal biomarkers by available algorithms were extracted and descriptively synthesized. Algorithm quality was assessed using a modified version of the Quality Assessment of Diagnostic Accuracy Studies 2 checklist with a focus on accuracy against established reference standards and risk of bias.ResultsThirty five studies reporting computer-aided diagnosis (CAD) tools for qualitative analysis or algorithms for quantitative analysis were identified. Compared with manual assessment in reference standards correlation coefficients ranged from 0.54 to 0.97 for drusen, 0.80 to 0.98 for geographic atrophy (GA), and 0.30 to 0.98 for intra- or subretinal fluid and pigment epithelial detachment (PED) detection by automated algorithms. CAD tools achieved area under the curve (AUC) values of 0.94 to 0.99, sensitivity of 0.90 to 1.00, and specificity of 0.89 to 0.92.ConclusionsAutomated analysis of AMD biomarkers on OCT is promising. However, most of the algorithm validation was performed in preselected patients, exhibiting the targeted biomarker only. In addition, type and quality of reported algorithm validation varied substantially.Translational RelevanceThe development of algorithms for combined, simultaneous analysis of multiple AMD biomarkers including AMD staging and the agreement on standardized validation procedures would be of considerable translational value for the clinician and the clinical researcher.

Human plasma metabolomics in age-related macular degeneration (AMD) using nuclear magnetic resonance spectroscopy

PurposeTo differentiate the plasma metabolomic profile of patients with age related macular degeneration (AMD) from that of controls, by Nuclear Magnetic Resonance (NMR) spectroscopy.MethodsTwo cohorts (total of 396 subjects) representative of central Portugal and Boston, USA phenotypes were studied. For each cohort, subjects were grouped according to AMD stage (early, intermediate and late). Multivariate analysis of plasma NMR spectra was performed, followed by signal integration and univariate analysis.ResultsSmall changes were detected in the levels of some amino acids, organic acids, dimethyl sulfone and specific lipid moieties, thus providing some biochemical information on the disease. The possible confounding effects of gender, smoking history and age were assessed in each cohort and found to be minimal when compared to that of the disease. A similar observation was noted in relation to age-related comorbidities. Furthermore, partially distinct putative AMD metabolite fingerprints were noted for the two cohorts studied, reflecting the importance of nutritional and other lifestyle habits in determining AMD metabolic response and potential biomarker fingerprints. Notably, some of the metabolite changes detected were noted as potentially differentiating controls from patients diagnosed with early AMD.ConclusionFor the first time, this study showed metabolite changes in the plasma of patients with AMD as compared to controls, using NMR. Geographical origins were seen to affect AMD patients´ metabolic profile and some metabolites were found to be valuable in potentially differentiating controls from early stage AMD patients. Metabolomics has the potential of identifying biomarkers for AMD, and further work in this area is warranted.