AMD Patients Research Articles

Viewpoint: AMD patients are being failed over dietary supplements

Following new research revealing vision improvement in AMD sufferers taking dietary supplements, Professor John Nolan of the Nutrition Research Centre Ireland, addresses the ‘controversy’ around use of carotenoids to boost eye health, and calls for immediate action to offer hope to patients

Validation of iPS Cell-Derived RPE Tissue in Animal Models.

Previous work suggests that replacing diseased Retinal Pigment Epithelium (RPE) with a healthy autologous RPE sheet can provide vision rescue for AMD patients. We differentiated iPSCs into RPE using a directed differentiation protocol. RPE cells at the immature RPE stage were purified and seeded onto either electrospun poly(lactic-co-glycolic acid) (PLGA) scaffolds or non-biodegradable polyester cell culture inserts and compared the two tissues. In vitro, PLGA and polyester substrates produced functionally similar results. Following in vitro evaluation, we tested RPE tissue in animal models for safety and function. Safety studies were conducted in RNU rats using an injection composed of intact cells and homogenized scaffolds. To assess function and develop surgical procedures, the tissues were implanted into an acute RPE injury model pig eye and evaluated using optical coherence tomography (OCT), multifocal ERG (mfERG), and histology. Subretinal injection studies in rats demonstrated safety of the implant. Biodegradability and biocompatibility data from a pig model demonstrated that PLGA scaffold is safe, with the added benefit of being resorbed by the body over time, leaving no foreign material in the eye. We confirmed that biodegradable substrates provide suitable support for RPE maturation and transplantation.

Association of treatment adherence with real-life VA outcomes in AMD, DME, and BRVO patients.

PurposeReal-life clinical outcomes of patients treated with anti-VEGF drugs for neovascular age-related macular degeneration (nAMD), diabetic macular edema (DME), or macular edema secondary to branch retinal vein occlusion (BRVO) are often inferior to results from randomized clinical trials. This observational cohort study investigates treatment adherence and real-life clinical outcomes within the first year of treatment.Patients and methodsA total of 708 treatment-naïve patients (466 nAMD, 134 DME, and 108 BRVO) were included. Patients were followed with a PRN treatment protocol with three intravitreal injections (IVIs) and a series of 3 monthly injections in case of persistent or recurrent disease activity, as determined by monthly follow-up exams including optical coherence tomographies. Occurrence of gaps of >56 days between treatments or follow-up (nonadherence [NA]) and the reasons for NA (patient- or center-associated) as well as disease activity within the first 12 months of treatment were analyzed. Visual acuity (VA) as well as numbers and dates of optical coherence tomography and IVI were extracted from medical records.ResultsNA occurred significantly more often in patients with DME (44%) than nAMD (32%) or BRVO (25%, p<0.01 between groups). NA was mainly patient-associated (nAMD: 80.0%, DME: 83.1%, BRVO: 70.4%, p=0.38 between groups). Patients with nAMD and DME and appropriate treatment/follow-up adherence had a better chance of significantly gaining or maintaining VA, respectively (19.9% vs 12.0% with 3-line-gain in nAMD and 1.3% vs 15.3% 3-line loss in DME; each p<0.05). NA did not correlate with VA outcomes in BRVO (3-line gain 30.9% vs 48.1% and 3-line loss 8.6% vs 7.4%; p>0.05).ConclusionNA to treatment and follow-up regimens is a common problem in the management of patients with AMD and DME and limits clinical treatment outcomes under real-life conditions. Patients with DME have the highest risk of patient-associated NA, associated with a higher risk for significant VA loss.

Fluorescence Lifetimes of Drusen in Age-Related Macular Degeneration.

The purpose of this study was to characterize fundus autofluorescence lifetimes of retinal drusen in patients with AMD. Patients with AMD and retinal drusen and healthy controls of similar age were examined. A fluorescence lifetime imaging ophthalmoscope was used. Retinal autofluorescence was excited using a 473-nm pulsed laser, and fundus autofluorescence lifetimes of the central retina (30°) were measured in two distinct spectral channels (short: 498 to 560 nm [SSC]; long: 560 to 720 nm [LSC]). Mean retinal autofluorescence lifetimes, corresponding fundus autofluorescence intensity images, spectral domain optical coherence tomography, color fundus images, and clinical data were investigated. Patients were analyzed in two distinct groups (soft drusen and reticular pseudodrusen) and compared with control subjects. Sixty-four eyes of 64 patients with AMD and retinal drusen (age: mean ± SD, 78 ± 8.5 years; range, 59 to 94 years) were investigated and compared with a control group of 20 age-matched healthy subjects. Mean retinal autofluorescence lifetimes in patients with AMD was significantly prolonged compared with the healthy control eyes (mean ± SEM: SSC, 486 ± 18 vs. 332 ± 11 ps, P < 0.0001; LSC: 493 ± 9 vs. 382 ± 17 ps, P < 0.0001). Areas of drusen featured a wide range of fluorescence lifetime values. Long lifetimes were identified in areas of atrophy and in areas of intraretinal hyperreflective deposits. Short lifetimes corresponded to deposits within the photoreceptor outer segment band. Mean retinal autofluorescence lifetimes in AMD patients are significantly prolonged. Intraretinal deposits cause prolonged lifetimes, whereas deposits in the area of the outer photoreceptor segments lead to short fluorescence lifetimes.

Plasma metabolomic study in Chinese patients with wet age-related macular degeneration

BackgroundAge-related macular degeneration (AMD) is a leading disease associated with blindness. It has a high incidence and complex pathogenesis. We aimed to study the metabolomic characteristics in Chinese patients with wet AMD by analyzing the morning plasma of 20 healthy controls and 20 wet AMD patients for metabolic differences.MethodsWe used ultra-high-pressure liquid chromatography and quadrupole-time-of-flight mass spectrometry for this analysis. The relationship of these differences with AMD pathophysiology was also assessed. Remaining data were normalized using Pareto scaling, and then valid data were handled using multivariate data analysis with MetaboAnalysis software, including unsupervised principal component analysis and supervised partial least squares-discriminate analysis. The purpose of the present work was to identify significant metabolites for the analyses. Hierarchical clustering was conducted to identify metabolites that differed between the two groups. Significant metabolites were then identified using the established database, and features were mapped on the Kyoto Encyclopedia of Genes and Genomes.ResultsA total of 5443 ion peaks were detected, all of them attributable to the same 10 metabolites. These included some amino acids, isomaltose, hydrocortisone, and biliverdin. The heights of these peaks differed significantly between the two groups. The biosynthesis of amino acids pathways also differed profoundly between patients with wet AMD and controls.ConclusionsThese findings suggested that metabolic profiles and and pathways differed between wet AMD and controls and may provide promising new targets for AMD-directed therapeutics and diagnostics.

Proteinics study: Relevance and interest for screening, follow‐up, etiopathogenesis of AMD

PurposeTo evaluate Proteinics study impact on AMD diagnosis, screening, follow‐up, etiopathogenesis.MethodsAMD: 40 patients,16 men,24 women,30 with AMD.4 Groups A,B,C,T. A:10 first stage AMD patients,B:10 Atrophy AMD patients(predominant atrophic areas),C:10 patients with Neovascular AMD,T:10 normal patients,control group. Ophthalmologic exam included ETDRS visual acuity(VA),complete ophthalmic examination,Fundus examination,autofluorescence imaging(FAF),(RegionFinderSoftware),optical coherence tomography(HRA Spectral Domain OCT)(OCTenFace software, M‐S software),Fluorescein angiography(FA)and ICG when Neovascular complication. Proteinics Study: Blood tests and analysis, proteinics qualitative and quantitative analysis,all the same for all patients,whatever group. Blood test is done during ophthalmologic exam. Plasma congelation,then Elisa technic,3 stages(SimpleStep Elisa kits)for detection and quantification of IL1β,IL6,IL10,TNFα,tau protein. Classic Elisa technic,2stages,for Amyloid peptid Aß1‐40.Each sample quadruplicate; MicroBCA analysis.ResultsAnalysis will determine proteinics quantitative values in each group of patients, and proportion, characterization, singularity of each of them: group A and C:similar results, higher for C; group T and B:similar results, lower for B. So, characterization, prevalence, specifics of and for each group, therefore a proteinics profile. Proteinics study'evaluation, identification, classification in AMD patients, and especially A, B, C groups allow AMD screening, follow‐up, particularly according to AMD type and stage. Proteinics study may have biomarker feature and let AMD preventionConclusionsProteinics study enhance AMD characterization, allow better diagnosis, follow‐up, screening. Interrelations, correlations between AMD and Proteinics lead to better etiopathogenesis understanding and therapeutics prospects.

Is macular pigment spatial profile a clinical biomarker in children of AMD parents?

SummaryA central dip in macular pigment (MP) has shown to be more prevalent with age, in AMD patients, and in smokers. We investigated clinical and genetic biomarkers in non‐smoking children of AMD parents (n = 131) over 40 years of age without AMD pathology in relation to their MP spatial profile. MP peak and volume were obtained with ARIS (Visual Pathways, Inc, Prescott, AZ USA) while spatial profiles for both eyes were classified visually and objectively. We explored risk factors including serum carotenoids and in‐vivo skin carotenoids, and cardiovascular biomarkers including omega‐3 fatty acids EPA and DHA, methylenetetrahydrofolate reductase C677T and A1298C in this well‐nourished Caucasian study group. Objectively, a central dip appeared more prevalent in children of AMD patients (41%) when compared to a healthy population. Those with central dip showed increased mean MP peak and volume, decreased serum L, Z, EPA and DHA, and decreased skin carotenoids in comparison to no dip. Identifying biomarkers in children genetically susceptible to AMD and introducing lifestyle changes such as nutrient repletion could provide invaluable advice to those associated with increased risk of AMD.

Anti‐VEGF intravitreal injections in the management of radiation maculopathy – Against

SummaryRadiation maculopathy (ischaemic or oedematous) is a dose‐dependent complication after radiation therapy associated with vascular decompensation caused by radiation damage. It develops primarily in patients treated for choroidal melanoma or other intraocular tumours following plaque brachytherapy or proton beam radiotherapy though it has been reported following epimacular brachytherapy for AMD patients or regional radiotherapy in other cases of malignant neoplasia. Though anti‐VEGF treatment has been proposed in the management of radiation maculopathy there are inherent problems in its use. The pharmacokinetics of anti‐VEGF necessitate repeated, and an increased number of intravitreal injections. The efficacy of anti‐VEGF has been most robust when used preventively. Alternative treatments include macular laser, triamcinolone acetonide injections and more recently dexamethasone implants. The former had good anatomic results and the latter have comparable results with anti‐VEGF agents in smaller series with less injections. Dexamethasone implants also had a more favourable outcome in cases non‐responsive to bevacizumab. Laser could still have an important preventive role especially in the treatment of ischemic areas.

Structural Bscan OCT correlation with OCT angiography biomarkers of activity in neovascular age related macular degeneration

PurposeFluid on structural B scan OCT is currently the most important sign among all imaging modalities of neovascular AMD. Recently, OCT angiography biomarkers of activity of nAMD have been described. The aim of the study is to compare the appearance of OCT angiography of eyes with wAMD with intraretinal or subretinal fluid against those without fluid on Bscan OCT.MethodsRetrospective cross sectional study.Fluid on Bscan OCT was classified as intraretinal fluid such as cysts and subretinal fluid as fluid under the retina. OCT angiography biomarkers of activity of nAMD were presence of secondary small vessel branching, peripheral arcades with anastomoses and presence of a dense net (DN). Inclusion criteria were: presence of neovessels on OCT angiography and sufficent images quality on both modalities. Presence of degenerative cysts and/or PED was an exclusion criteria.Results59 eyes of 55 patients naive or treated for wet AMD were included (average age: 80 (SD ±7), min‐max: 63‐95). Subretinal fluid was present in 20% eyes (12/59), intraretinal fluid in 25% (15/59), a combination of both in 10% (6/59) and absence of subretinal or intraretinal fluid in 44% (26/59). DN was present in 30% (18/59), LN in 69% (41/59). Dense net was present in statistically significantly more of the uncontrolled eyes 45% (15/33) (p=0.009, fisher exact test.) Peripheral arcades with anastomoses were evident in 46% (12/26) of controlled eyes and 61% (20/33) of uncontrolled eyes. Small vessel branching was evident in 0% (0/26) of the controlled eyes and in 36% (12/33) uncontrolled eyes.ConclusionsDense net are significantly more prevalent in eyes with uncontrolled AMD; small vessel branching and peripheral loops are also more prevalent in these eyes. This suggest that OCT‐A provides imaging of the morphology of choroidal CNV in AMD patients associated with disease activity.

Vision improvement in dry and wet Age‐Related Macular Degeneration (AMD) patients after treatment with new corneal CPV procedure for light redirections onto the retina

PurposeTo evaluate in two clinics vision improvement in AMD patients treated with a new corneal elastic modulus‐altering procedure (CPV) for light redirections onto multiple retinal areas.MethodsTen eyes (6 wet, 4 dry), with best‐corrected near and distance visual acuities (CNVA and CDVA) between 20/80 and 20/400, without pathological retinal morphology completely affecting the central 10° (3 mm) on OCT, were given a single treatment of CPV and were examined for 12 months post‐CPV.ResultsLogMAR Mean CNVA and CDVA in treated eyes improved (p &lt; 0.01) from pre‐CPV 0.957 and 0.942 to 0.512 and 0.638, 0.544 and 0.602, 0.531 and 0.672 at 1, 6 and 12 m, respectively. The number of ETDRS letters gained in mean binocular CNVA/CDVA were 22/15, 25/19, 25/18 at 1, 6, and 12 m, respectively. Administered NEI VFQ‐25 at 3 m improved from pre‐CPV. No CPV complications or adverse events occurred.ConclusionsCPV effectively, safely, rapidly and comfortably redirects light onto functional retinal areas after a single treatment to provide, without visual training, significantly improved monocular and binocular near and distance vision without causing peripheral field restriction or diplopia in patients with atrophic or neovascular AMD with poor vision, including after anti‐VEGF therapy to which CPV is complementary and for which CPV may improve compliance because of improved vision restoration.

Serum adiponectin associates with aging rather than neovascular AMD

PurposeTo study whether adiponectin serum levels are changed in neovascular AMD. Adiponectin, secreted primarily by adipocytes, modulates cellular energy metabolism. Adiponectin participates also in retinal homeostasis and autophagy. Adiponectin receptors are abundant in RPE‐choroid level. We have previously reported that ADIPOR1‐gene variant associates with advanced AMD in Finnish population. Here, we evaluate the role of peripheral adiponectin in AMD.MethodsAfter obtaining written informed consent, venous blood samples were collected from 145 neovascular AMD patients (mean age: 79.4 ± 6.1 years) and 90 controls (mean age: 74.6 ± 6.3 years) in a cross‐sectional design. Other causes of macular oedema were excluded. Serum adiponectin levels were quantified using a commercial ELISA kit.ResultsSerum levels of adiponectin in neovascular AMD group (12.0 ± 9.5 μg/mL) did not significantly differ from levels in control group (11.1 ± 7.8 μg/mL; t‐test p = 0.423). Adiponectin levels increased with aging (rs = 0.237, p = 0.01). Women had higher mean adiponectin levels (13.2 ± 9.4 μg/mL, n = 175) compared to men (7.8 ± 6.2 μg/mL, n = 70; t‐test p = 0.001), but no difference was detected between AMD‐patients and controls when analyzed separately by gender (women: 13.2 ± 8.6 μg/mL vs. 13.2 ± 9.9 μg/mL, respectively; t‐test p = 0.959; men: 8.3 ± 7.5 μg/mL vs. 7.2 ± 4.0 μg/mL, respectively; t‐test p = 0.439).ConclusionsOur findings suggest that serum adiponectin levels are not altered in neovascular AMD. However, adiponectin may be considered more like a general serum biomarker of aging. As already known, gender can influence the adiponectin level in elderly.

This issue of ACTA

The cover illustration shows a prolene thread inserted into Schlemm′s canal as reported by Grieshaber et al. in this issue of Acta. Klemp et al. in Copenhagen describe exudative retinal detachment, cataract formation and melanocytic tumours as the characteristic ocular findings in uveal melanocytic proliferation. Campos et al. in Portugal consider the role of the choroid in diabetic macular oedema and how OCT studies of the choroid advance this field of research. Jørstad et al. in Oslo found positive results with aflibercept in AMD eyes which were resistant to ranibizumab and bevacizumab injections. Many eyes required more than bimonthly aflibercept injections. Vujosevic et al. in Italy emphasize inflammatory biomarkers in diabetic macular oedema such as hyperreflective retinal spots, subfoveal neuroretinal detachment and increased foveal autofluorescence. These biomarkers improved with dexamethasone treatment. Subhi and Lykke Sørensen in Denmark correlated systemic leucocyte activity with onset and size of neovascular lesions in AMD. They suggest that inflammation plays a role in the pathophysiology. Sauer et al. in Germany used fluorescence lifetime imaging to map pigment distribution and evaluate its role in macular hole formation. Hamoudi et al. in Denmark found no difference in corneal endothelial density between combined and separated phacoemulsification and vitrectomy for epiretinal membrane. Makdoumi et al. in Sweden and the USA used blue light and riboflavin to kill resistant staphylococci and suggest this approach may work in corneal infections. Margulis et al. in the Netherlands, Spain and Denmark examined use of nonsteroidal anti-inflammatory eye drops. These were used with cataract surgery and for allergic conjunctivitis. Pärssinen and Kauppinen in Finland found astigmatism to increase with myopia. Corneal astigmatism predominates. Grieshaber et al. in South Africa and Switzerland recommend the Stegmann Schlemm′s canal expander to lower intraocular pressure in patients with moderate to advanced open angle glaucoma. Quérat and Chen in Sweden recommend ICARE® self-tonometry. Day-to-day variations are seen in most patients. Mrowicka et al. in Poland propose that insufficient antioxidant capacity may play a role in age related macular degeneration. This hypothesis is based on study of enzymes and genes related to antioxidant activity. Pilat et al. in UK used hand-held OCT to image anterior segment dysgenesis in children. The following articles are published electronically only and are accessible on www.actaophtalmologica.com, where you can also access the printed articles. Nitta et al. in Japan report an increase in retinal oximetry readings in retinal venules following glaucoma surgery and lowering of intraocular pressure. Tanito et al. in Japan recommend microhook trabeculotomy in glaucoma patients. It lowers IOP and reduces need for glaucoma medications. Mansouri and Shaarawy in the USA and Switzerland fond similar outcomes with scanning laser trabeculoplasty and selective laser trabeculoplasty. Lanza et al. in Italy used light backscattering with in vivo corneal confocal microscopy to evaluate changes in corneal layers in patients with Sjögren syndrome and dry eyes. Shimada et al. in Japan found reduced anterior chamber bacterial contamination with either 0.025% povidone-iodine or 0.0025% polyvinyl alcohol iodine irrigation of ocular surface. Peiretti et al. in Italy, the Netherlands, Saudi Arabia and the USA examined retinal folds following retinal detachment repair. This was not influenced by use of heavy liquids in surgery or postoperative positioning. Chan et al. in Hong Kong, Singapore and UK seek ways to measure and control emulsification of intraocular silicone oil. Kang et al. in Korea measured greater choroidal thickness in eyes with vitreoretinal traction. This was reversed following epiretinal membrane surgery and release of the traction. Jonas et al. in India and Germany found myopic retinopathy to be rare in rural India. It was associated with open angle glaucoma. Carreño et al. in Spain, USA and UK measured several inflammatory cytokines in tear fluid in uveitis patients. Bray et al. in UK suggest that portable electronic vision enhancement systems are cost-effective to improve near vision. However, it may not provide cost-effective improvement in quality of life. Babeau et al. in France comment on an article on using adaptive optics for screening chloroquine retinopathy. Parrovano et al. in Italy used OCT angiography to monitor reduction in neovascular lesions. Van Asten et al. in the Netherlands found positive outcomes with aflibercept in AMD patients unresponsive to bevacizumab. Lytvynchuk in Ukraine surgically removed a retinal branch artery embolus. Dimopoulos et al. in Germany used autologous thrombocyte serum for macular hole surgery. Grewal et al. in USA used intraoperative OCT to guide placement of Argus II retinal prosthesis. Huynh et al. in the USA found no benefit from dilated fundus examination in patients with spontaneous subconjunctival haemorrhage. Qu-Knafo et al. in France suggest that eyes with DME and serous retinal detachment may show greater improvement with ranibizumab than eyes without serous detachment. I hope you are enjoying a pleasant summer and Acta fits into your summer reading schedule.

Associations Between β-Peripapillary Atrophy and Reticular Pseudodrusen in Early Age-Related Macular Degeneration.

PurposeChoroidal thinning has been associated with reticular pseudodrusen (RPD) and β-peripapillary atrophy (β-PPA), which have been linked to normal-tension glaucoma (NTG). This analysis sought to determine whether RPD are independently associated with β-PPA in early AMD patients. Secondary outcomes included the association of RPD and preexisting diagnosis of glaucoma, cup-to-disc ratio (CDR), subfoveal choroidal thickness (SFCT), and IOP.MethodsThis prospective cross-sectional study examined 78 age- and sex-matched early AMD patients: 43 RPD patients (63 eyes) and 35 non-RPD patients (64 eyes). Exclusion criteria included advanced AMD, high myopia, and vitreoretinal conditions/surgery. RPD and non-RPD groups were identified by confocal scanning laser ophthalmoscopy. β-PPA as well as CDR were graded on digital, nonstereoscopic fundus photos. SFCT was measured on spectral-domain optical coherence tomography for 69 patients (35 RPD and 34 non-RPD). IOP and glaucoma diagnosis were extracted from charts.Resultsβ-PPA had a greater prevalence in RPD than non-RPD (44% vs. 19%, P = 0.002); however, this relationship was not significant when SFCT was added to the model (P = 0.150). A preexisting diagnosis of glaucoma (P = 0.156), CDR (P = 0.176), and IOP (P = 0.98) was not different between groups.ConclusionsRPD in early AMD are associated with presence of β-PPA, but choroidal thickness is a confounder in this relationship. Because β-PPA is a common finding in NTG, focusing on a potential shared pathway between RPD and NTG could improve the understanding of pathophysiology and expand therapies for each condition.

Serum Levels of TIMP-3, LIPC, IER3, and SLC16A8 in CFH-Negative AMD Cases.

AMD is a complex eye disease predominantly occurring in aged population. Till now about 53 genetic loci have been found to be associated with the AMD pathology. AMD pathogenesis is being increasingly known to progress through mechanisms independent of the CFH mediated pathway. Therefore, our aim for current study was to examine the genes by analyzing their expression levels in AMD. We recruited about 50 AMD and same number of age matched controls. We analyzed the CFH duplication and deletion by multiplex ligation probe amplification (MLPA) and found no duplication and deletion in CFH gene in AMD patients. We also estimated the IER-3, SLC16A8, LIPC, and TIMP-3 expression levels in both CFH-negative AMD cases (i.e. no duplication and deletion in CFH gene) besides examining these in AMD and controls. We found that the expression level of LIPC, SLC16A8, and TIMP-3 was significantly associated with AMD pathology in both groups (LIPC: P = 0.008, SLC16A8: P < 0.001, TIMP-3: P < 0.001, respectively). However, we did not find any significant difference in IER-3 levels in AMD and controls. Therefore, the evidence from current study, suggests that AMD pathology may be mediated through mechanistic pathways linked to other genetic loci. J. Cell. Biochem. 118: 2087-2095, 2017. © 2016 Wiley Periodicals, Inc.

Systematic Functional Testing of Rare Variants: Contributions of CFI to Age-Related Macular Degeneration.

PurposeGenome-wide association (GWAS) and sequencing studies for AMD have highlighted the importance of coding variants at loci that encode components of the complement pathway. However, assessing the contribution of such alleles to AMD, especially when they are rare, remains coarse, in part because of the persistent challenge in establishing their functional relevance. Others and we have shown previously that rare alleles in complement factor I (CFI) can be tested functionally using a surrogate in vivo assay of retinal vascularization in zebrafish embryos. Here, we have implemented and scaled these tools to assess the overall contribution of rare alleles in CFI to AMD.MethodsWe performed targeted sequencing of CFI in 731 AMD patients, followed by replication in a second patient cohort of 511 older healthy individuals. Systematic functional testing of all alleles and post-hoc statistical analysis of functional variants was also performed.ResultsWe discovered 20 rare coding nonsynonymous variants, including the previously reported G119R allele. In vivo testing led to the identification of nine variants that alter CFI; six of which are associated with hypoactive complement factor I (FI). Post-hoc analysis in ethnically matched, population controls showed six of these to be present exclusively in cases.ConclusionsTaken together, our data argue that multiple rare and ultra-rare alleles in CFI contribute to AMD pathogenesis; they improve the precision of the assessment of the contribution of CFI to AMD; and they offer a rational route to establishing both causality and direction of allele effect for genes associated with this disorder.

Aberrant Buildup of All-Trans-Retinal Dimer, a Nonpyridinium Bisretinoid Lipofuscin Fluorophore, Contributes to the Degeneration of the Retinal Pigment Epithelium

Nondegradable fluorophores that accumulate as deleterious lipofuscin of RPE are involved in pathological mechanisms leading to the degeneration of RPE in AMD. A2E, a major component of RPE lipofuscin, could cause damage to RPE cells. Nevertheless, all-trans-retinal dimer (atRAL dimer) was found to be much more abundant than that of A2E in eyes of Abca4-/-Rdh8-/- double-knockout (DKO) mice, a rodent model showing the typical characteristics of retinopathies in AMD patients. Our aim was to elucidate the effect and mechanism of atRAL dimer-induced RPE degeneration. Eyes harvested from C57BL/6J wild-type (WT) and Abca4-/-Rdh8-/- DKO mice were examined by HPLC. Cellular uptake, subcellular localization, 5-bromo-2-deoxyuridine (BrdU), Cdc25C, DNA strand breaks, mitochondrial membrane potential (ΔΨm), and cytochrome c were analyzed by fluorescence microscopy. Cellular toxicity was assayed by lactate dehydrogenase (LDH) assay and dead cell staining. Apoptosis and cell-cycle stages were detected by flow cytometry. Furthermore, in vitro and in vivo expression of proteins associated with cell cycle and apoptosis was measured by immunoblot assays. All-trans-retinal dimer clearly could damage RPE cell membrane and inhibit the proliferation of RPE cells as well as induce DNA damage and cell-cycle arrest at the G2/M phase via activating the ATM/ATR-Chk2-p53 signaling pathway. Moreover, this di-retinal adduct triggered mitochondrion-associated apoptosis in RPE cells. Evidence from the cell-based experiments was also corroborated by a remarkable abnormality in expression of proteins associated with cell cycle (Cyclin B1 and Cdc2) and apoptosis (p53, Bcl-2 and Bax) in the RPE of Abca4-/-Rdh8-/- DKO mice. These findings suggest that atRAL dimer that accumulates beyond a critical level, facilitates age-dependent RPE degeneration.

Age-related macular degeneration associated polymorphism rs10490924 in ARMS2 results in deficiency of a complement activator

BackgroundAge-related macular degeneration (AMD) is the leading cause of blindness in developed countries. The polymorphism rs10490924 in the ARMS2 gene is highly associated with AMD and linked to an indel mutation (del443ins54), the latter inducing mRNA instability. At present, the function of the ARMS2 protein, the exact cellular sources in the retina and the biological consequences of the rs10490924 polymorphism are unclear.MethodsRecombinant ARMS2 was expressed in Pichia pastoris, and protein functions were studied regarding cell surface binding and complement activation in human serum using fluoresence-activated cell sorting (FACS) as well as laser scanning microscopy (LSM). Biolayer interferometry defined protein interactions. Furthermore, endogenous ARMS2 gene expression was studied in human blood derived monocytes and in human induced pluripotent stem cell-derived microglia (iPSdM) by PCR and LSM. The ARMS2 protein was localized in human genotyped retinal sections and in purified monocytes derived from AMD patients without the ARMS2 risk variant by LSM. ARMS2 expression in monocytes under oxidative stress was determined by Western blot analysis.ResultsHere, we demonstrate for the first time that ARMS2 functions as surface complement regulator. Recombinant ARMS2 binds to human apoptotic and necrotic cells and initiates complement activation by recruiting the complement activator properdin. ARMS2-properdin complexes augment C3b surface opsonization for phagocytosis. We also demonstrate for the first time expression of ARMS2 in human monocytes especially under oxidative stress and in microglia cells of the human retina. The ARMS2 protein is absent in monocytes and also in microglia cells, derived from patients homozygous for the ARMS2 AMD risk variant (rs10490924).ConclusionsARMS2 is likely involved in complement-mediated clearance of cellular debris. As AMD patients present with accumulated proteins and lipids on Bruch’s membrane, ARMS2 protein deficiency due to the genetic risk variant might be involved in drusen formation.

Is There Vitamin D Deficiency in AMD Patients?

Is There Vitamin D Deficiency in AMD Patients?

Warfarin use among wet AMD patients

PurposeTo evaluate the prevalence of warfarin use among wet AMD patients, and impact on clinical outcome.MethodsA retrospective analysis of patient records was conducted in Kuopio University Hospital, Finland. We obtained best possible medication history of 414 wet AMD patients and 247 controls, aged 60 years or over, and already excluding other causes of macular oedema by ophthalmologist. Controls were patients with glaucoma or age‐related cataract, with no wet AMD. Due missing medication history, 139 wet AMD patients and 71 controls (p = 0.216) were kept out from the study. When possible, we obtained real‐life 2‐year outcome data of wet AMD treatment for the patient's first eye, solely treated with anti‐VEGF pro re nata.ResultsThe study included 276 wet AMD patients (mean age: 79.5 years) and 175 controls (mean age: 75.3 years). A positive warfarin medication history was detected in 57 (20.7%) wet AMD patients, compared to 16 (9.1%) in control group. Crude OR was 2.59 (95% CI 1.43–4.67, p = 0.001). Age‐adjusted OR was 1.92 (95% CI 1.02–3.60, p = 0.018). Treatment outcome data was obtained from 11 patients (mean age: 77.3 years) using warfarin and 54 patients (mean age: 79.1 years) without warfarin medication. Snellen visual acuity didn't differ between these wet AMD groups at the baseline (0.3 vs. 0.2, p = 0.319). After 2‐year follow‐up, no difference was seen in the change of visual acuity (+0.3 vs. +0.3, p = 0.696), or in number of anti‐VEGF injections (7.8 vs. 9.0, p = 0.435) with respect to the use of warfarin.ConclusionsOur study suggest that warfarin use may be more common among wet AMD patients, but it does not have any effect on the outcome of wet AMD treatment. As the study sample is small and wet AMD‐related cardiovascular factors are susceptible to interfere, these initial findings should be verified in a larger prospective study.

Spontaneous anatomical improvement on OCT findings in patients with neovascular age‐related macular degeneration without anti‐VEGF treatment

PurposeTo present some patients with wet AMD can be improved spontaneously on OCT findings without anti‐VEGF treatment.MethodsWe reviewed the medical records of wet AMD patients visiting Gangneung Asan Hospital from December 2013 to May 2015. There were group of patients who refused to receive anti‐VEGF treatment due to various reasons, even though there were remaining subretinal fluid and/or pigment epithelial detachment on OCT. Among those patients, we investigated spontaneous anatomical improvement cases without anti‐VEGF injection.ResultsThere were nine cases showing spontaneous anatomical improvement on OCT findings. Average age was 70.0 ± 9.4, male was seven, and female was two. Number of anti‐VEGF injection was 4.4 ± 1.7, and injection‐free interval was 12 ± 5.7 months before showing spontaneous anatomical improvement on OCT. Seven cases showed resolution of subretinal fluid, and two cases showed decrease of pigment epithelial detachment size. Initial visual acuity was 0.64 ± 0.36 in LogMAR scale, and final visual acuity improved to 0.40 ± 0.28 in LogMAR scale. Six patients were treated with bevacizumab, two patients received aflibercept, and one patient was injected with ranibizumab. Total follow up periods was 21.0 ± 5.3 months. During the periods, three cases worsened and re‐treatment was performed with anti‐VEGF agents. Six patients have not shown recurrence for more than 6 months. Those nine patients showed relatively small number of anti‐VEGF injection and improvement of visual acuity.ConclusionsThese findings might imply that some patients showing small amount of remaining SRF and/or PED might not need frequent anti‐VEGF injection, especially in patients showing good response to anti‐VEGF therapy with relatively good initial visual acuity. And closed follow up will be essential and needed for those patients.