Zinc supplementation inhibits complement activation in age-related macular degeneration.
ABSTRACT: Age-related macular degeneration (AMD) is the leading cause of blindness in the Western world. AMD is a multifactorial disorder but complement-mediated inflammation at the level of the retina plays a pivotal role. Oral zinc supplementation can reduce the progression of AMD but the precise mechanism of this protective effect is as yet unclear. We investigated whether zinc supplementation directly affects the degree of complement activation in AMD and whether there is a relation between serum complement catabolism during zinc administration and the complement factor H (CFH) gene or the Age-Related Maculopathy susceptibility 2 (ARMS2) genotype. In this open-label clinical study, 72 randomly selected AMD patients in various stages of AMD received a daily supplement of 50 mg zinc sulphate and 1 mg cupric sulphate for three months. Serum complement catabolism-defined as the C3d/C3 ratio-was measured at baseline, throughout the three months of supplementation and after discontinuation of zinc administration. Additionally, downstream inhibition of complement catabolism was evaluated by measurement of anaphylatoxin C5a. Furthermore, we investigated the effect of zinc on complement activation in vitro. AMD patients with high levels of complement catabolism at baseline exhibited a steeper decline in serum complement activation (p<0.001) during the three month zinc supplementation period compared to patients with low complement levels. There was no significant association of change in complement catabolism and CFH and ARMS2 genotype. In vitro zinc sulphate directly inhibits complement catabolism in hemolytic assays and membrane attack complex (MAC) deposition on RPE cells. This study provides evidence that daily administration of 50 mg zinc sulphate can inhibit complement catabolism in AMD patients with increased complement activation. This could explain part of the mechanism by which zinc slows AMD progression.The Netherlands National Trial Register NTR2605.
Project description:BACKGROUND/AIMS:The Age-Related Eye Disease Study (AREDS) reported the beneficial impact of antioxidant and zinc supplements on the risk of progression to advanced stages of age-related macular degeneration (AMD). We evaluated the role of genetic variants in modifying the relationship between supplementation and progression to advanced AMD. METHODS:Among 4124 eyes (2317 subjects with a genetic specimen), 882 progressed from no AMD, early or intermediate AMD to overall advanced disease, including geographic atrophy (GA) and neovascular disease (NV) over the course of the clinical trial. Survival analysis using individual eyes as the unit of analysis was used to assess the effect of supplementation on AMD outcomes, with adjustment for demographic, environmental, ocular and genetic covariates. Interaction effects between supplement groups and individual complement factor H (CFH) Y402H and age-related maculopathy susceptibility 2 (ARMS2) genotypes, and composite genetic risk groups combining the number of risk alleles for both loci, were evaluated for their association with progression. RESULTS:Among antioxidant and zinc supplement users compared with the placebo group, subjects with a non-risk genotype for CFH (TT) had a lower risk of progression to advanced AMD (HR: 0.55, 95% CI 0.32 to 0.95, p=0.033). No significant treatment effect was apparent among subjects who were homozygous for the CFH risk allele (CC). A protective effect was observed among high-risk ARMS2 (TT) carriers (HR: 0.52, 95% CI 0.33 to 0.82, p=0.005). Similar results were seen for the NV subtype but not GA. CONCLUSIONS:The effectiveness of antioxidant and zinc supplementation appears to differ by genotype. Further study is needed to determine the biological basis for this interaction. TRIAL REGISTRATION NUMBER:NCT00594672, pre-results.
Project description:OBJECTIVE:To determine whether genotypes at 2 major loci associated with late age-related macular degeneration (AMD), complement factor H (CFH) and age-related maculopathy susceptibility 2 (ARMS2), influence the relative benefits of Age-Related Eye Disease Study (AREDS) supplements. DESIGN:Unplanned retrospective evaluation of a prospective, randomized, placebo-controlled clinical trial of vitamins and minerals for the treatment of AMD. SUBJECTS:AREDS participants (mean age, 69 years) who were at risk of developing late AMD and who were randomized to the 4 arms of AREDS supplement treatment. METHODS:Analyses were performed using the Cox proportional hazards model to predict progression to late AMD (neovascular or central geographic atrophy). Statistical models, adjusted for age, gender, smoking status, and baseline AMD severity, were used to examine the influence of genotypes on the response to therapy with 4 randomly assigned arms of AREDS supplement components: placebo, antioxidants (vitamin C, vitamin E, ?-carotene), zinc, or a combination. MAIN OUTCOME MEASURES:The influence of the genotype on the relative treatment response to the randomized components of the AREDS supplement, measured as progression to late AMD. RESULTS:Of the 1237 genotyped AREDS participants of white ethnicity, late AMD developed in 385 (31.1%) during the mean follow-up of 6.6 years. As previously demonstrated, CFH genotype (P = 0.005), ARMS2 (P< 0.0001), and supplement were associated individually with progression to late AMD. An interaction analysis found no evidence that the relative benefits of AREDS supplementation varied by genotype. Analysis of (1) CFH rs1061170 and rs1410996 combined with ARMS2 rs10490924 with the 4 randomly assigned arms of AREDS supplement and (2) analysis of the combination of CFH rs412852 and rs3766405 with ARMS2 c.372_815del443ins54 with the AREDS components resulted in no interaction (P = 0.06 and P = 0.45, respectively, before multiplicity adjustment). CONCLUSIONS:The AREDS supplements reduced the rate of AMD progression across all genotype groups. Furthermore, the genotypes at the CFH and ARMS2 loci did not statistically significantly alter the benefits of AREDS supplements. Genetic testing remains a valuable research tool, but these analyses suggest it provides no benefits in managing nutritional supplementation for patients at risk of late AMD.
Project description:BACKGROUND: To determine whether the functional effects of oral supplementation with Saffron, a natural compound that proved to be neuroprotective in early age-related macular degeneration, are influenced by complement factor H (CFH) and age-related maculopathy susceptibility 2 (ARMS2) risk genotypes. METHODS: Thirty-three early AMD patients, screened for CFH (rs1061170) and ARMS2 (rs10490924) polymorphisms and receiving Saffron oral supplementation (20 mg/day) over an average period of treatment of 11 months (range, 6-12), were longitudinally evaluated by clinical examination and focal electroretinogram (fERG)-derived macular (18°) flicker sensitivity estimate. fERG amplitude and macular sensitivity, the reciprocal value of the estimated fERG amplitude threshold, were the main outcome measures. RESULTS: After three months of supplementation, mean fERG amplitude and fERG sensitivity improved significantly when compared to baseline values (p < 0.01). These changes were stable throughout the follow-up period. No significant differences in clinical and fERG improvements were observed across different CFH or ARMS2 genotypes. CONCLUSIONS: The present results indicate that the functional effect of Saffron supplementation in individual AMD patients is not related to the major risk genotypes of disease.
Project description:We evaluated the influence of an antioxidant and zinc nutritional supplement [the Age-Related Eye Disease Study (AREDS) formulation] on delaying or preventing progression to neovascular AMD (NV) in persons with age-related macular degeneration (AMD). AREDS subjects (n = 802) with category 3 or 4 AMD at baseline who had been treated with placebo or the AREDS formulation were evaluated for differences in the risk of progression to NV as a function of complement factor H (CFH) and age-related maculopathy susceptibility 2 (ARMS2) genotype groups. We used published genetic grouping: a two-SNP haplotype risk-calling algorithm to assess CFH, and either the single SNP rs10490924 or 372_815del443ins54 to mark ARMS2 risk. Progression risk was determined using the Cox proportional hazard model. Genetics-treatment interaction on NV risk was assessed using a multiiterative bootstrap validation analysis. We identified strong interaction of genetics with AREDS formulation treatment on the development of NV. Individuals with high CFH and no ARMS2 risk alleles and taking the AREDS formulation had increased progression to NV compared with placebo. Those with low CFH risk and high ARMS2 risk had decreased progression risk. Analysis of CFH and ARMS2 genotype groups from a validation dataset reinforces this conclusion. Bootstrapping analysis confirms the presence of a genetics-treatment interaction and suggests that individual treatment response to the AREDS formulation is largely determined by genetics. The AREDS formulation modifies the risk of progression to NV based on individual genetics. Its use should be based on patient-specific genotype.
Project description:PURPOSE:Several genes encoding complement system components and fragments are associated with age-related macular degeneration (AMD). This study was conducted to determine whether alterations in circulating levels of these markers of complement activation and regulation are also independently associated with advanced AMD and whether they are related to AMD genotypes. METHODS:Plasma and DNA samples were selected from individuals in our AMD registry who had progressed to or developed the advanced stages of AMD, including 58 with geographic atrophy and 62 with neovascular disease. Subjects of similar age and sex, but without AMD, and who did not progress were included as controls (n = 60). Plasma complement components (C3, CFB, CFI, CFH, and factor D) and activation fragments (Bb, C3a, C5a, iC3b, and SC5b-9) were analyzed. DNA samples were genotyped for seven single-nucleotide polymorphisms in six genes previously shown to be associated with AMD: CFB, CFH, C2, C3, and CFI and the LOC387715/ARMS2 gene region. The association between AMD and each complement biomarker was assessed by using logistic regression, controlling for age, sex, and proinflammatory risk factors: smoking and body mass index (BMI). Functional genomic analyses were performed to assess the relationship between the complement markers and genotypes. Concordance, or C, statistics were calculated to assess the effect of complement components and activation fragments in an AMD gene-environment prediction model. RESULTS:The highest quartiles of Bb and C5a were significantly associated with advanced AMD, when compared with the lowest quartiles. In multivariate models without genetic variants, the odds ratio (OR) for Bb was 3.3 (95% confidence interval [CI] = 1.3-8.6), and the OR for C5a was 3.6 (95% CI = 1.2-10.3). With adjustment for genetic variants, these ORs were substantially higher. The alternative pathway regulator CFH was inversely associated with AMD in the model without genotypes (OR = 0.3; P = 0.01). Positive associations were found between BMI and plasma C3, CFB, CFH, iC3b, and C3a. There were also significant associations between C5a fragment and LOC387715/ARMS2 and C3 genotypes (P for trend = 0.02, 0.04), respectively. C statistics for models with behavioral and genetic factors increased to 0.94 +/- 0.20 with the addition of C3a, Bb, and C5a. CONCLUSIONS:Increased levels of activation fragments Bb and C5a are independently associated with AMD. Higher BMI is related to increased levels of complement components. C5a is associated with AMD genotypes. C statistics are stronger with the addition of C3a, Bb, and C5a in predictive models. Results implicate ongoing activation of the alternative complement pathway in AMD pathogenesis.
Project description:<h4>Purpose</h4>To assess whether genotypes at 2 major loci associated with age-related macular degeneration (AMD), complement factor H (CFH), or age-related maculopathy susceptibility 2 (ARMS2), modify the response to oral nutrients for the treatment of AMD in the Age-Related Eye Disease Study 2 (AREDS2).<h4>Design</h4>Post hoc analysis of a randomized trial.<h4>Participants</h4>White AREDS2 participants.<h4>Methods</h4>AREDS2 participants (n = 4203) with bilateral large drusen or late AMD in 1 eye were assigned randomly to lutein and zeaxanthin, omega-3 fatty acids, both, or placebo, and most also received the AREDS supplements. A secondary randomization assessed modified AREDS supplements in 4 treatment arms: lower zinc dosage, omission of β-carotene, both, or no modification. To evaluate the progression to late AMD, fundus photographs were obtained at baseline and annual study visits, and history of treatment for late AMD was obtained at study visits and 6-month interim telephone calls. Participants were genotyped for the single-nucleotide polymorphisms rs1061170 in CFH and rs10490924 in ARMS2. Bivariate frailty models using both eyes were conducted, including a gene-supplement interaction term and adjusting for age, gender, level of education, and smoking status. The main treatment effects, as well as the direct comparison between lutein plus zeaxanthin and β-carotene, were assessed for genotype interaction.<h4>Main outcome measures</h4>The interaction between genotype and the response to AREDS2 supplements regarding progression to late AMD, any geographic atrophy (GA), and neovascular AMD.<h4>Results</h4>Complete data were available for 2775 eyes without baseline late AMD (1684 participants). The participants (mean age ± standard deviation, 72.1±7.7 years; 58.5% female) were followed up for a median of 5 years. The ARMS2 risk allele was associated significantly with progression to late AMD and neovascular AMD (P = 2.40 × 10<sup>-5</sup> and P = 0.002, respectively), but not any GA (P = 0.097). The CFH risk allele was not associated with AMD progression. Genotype did not modify significantly the response to any of the AREDS2 supplements.<h4>Conclusions</h4>CFH and ARMS2 risk alleles do not modify the response to the AREDS2 nutrient supplements with respect to the progression to late AMD (GA and neovascular AMD).
Project description:PURPOSE: To evaluate the contribution of genetic variants of complement factor H (CFH), complement component 2 and 3 (C2 and C3), complement factor B (CFB), and age-related maculopathy susceptibility 2 (ARMS2) to age-related macular degeneration (AMD) risk in the Mexican Mestizo population. METHODS: Analysis included 282 unrelated Mexican patients with advanced AMD, 205 healthy controls, and 280 population controls. Stereoscopic fundus images were graded on the Clinical Age-Related Maculopathy System (CARMS). We designed a resequencing strategy using primers with M13 adaptor for the 23 exons of the CFH gene in a subgroup of 96 individuals clinically evaluated: 48 AMD cases and 48 age- and sex-matched healthy controls. Single nucleotide polymorphisms (SNPs) in C3 (Arg80Gly and Pro292Leu), C2 (rs547154), CFB (Leu9His), and ARMS2 (Ala69Ser) were genotyped in all patients, healthy and population controls using TaqMan assay. RESULTS: All evaluated individuals were Mexican Mestizos, and their genetic ancestry was validated using 224 ancestry informative markers and calculating F(st) values. The CFH resequencing revealed 19 SNPs and a common variant in the intron 2 splice acceptor site; three CFH haplotypes inferred from individual genotypes, showed significant differences between cases and controls. The risk alleles in C3 (rs1047286, odds ratio [OR]=2.48, 95% confidence interval [CI]=1.64-3.75, p=1.59E-05; rs2230199, OR=2.15, 95% CI=1.48-3.13, p=6.28E-05) and in ARMS2 (rs10490924, OR=3.09, 95% CI=2.48-3.86, p=5.42E-23) were strongly associated with risk of AMD. The protective effect of alleles in C2 (rs547154) and CFB (rs4151667) showed a trend but was not significantly associated after correction for multiple testing. CONCLUSIONS: Our results show that ARMS2 and C3 are major contributors to advanced AMD in Mexican patients, while the contributions of CFH, C2, and CFB are minor to those of other populations, reveling significant ethnic differences in minor allele frequencies. We provide evidence that two specific common haplotypes in the CFH gene predispose individuals to AMD, while another may confer reduced risk of disease in this admixed population.
Project description:PURPOSE:To examine the association between sequence variants in genetic risk factors for age-related macular degeneration (AMD) and delayed rod-mediated dark adaptation (RMDA), the first functional biomarker for incident AMD, in older adults with normal macular health and early AMD. DESIGN:Cross-sectional. PARTICIPANTS:Adults 60 years of age or older showing normal macular health (defined as both eyes at step 1 on the Age-Related Eye Disease Study 9-step AMD classification system) and those with AMD in one or both eyes (defined as steps 2-9). METHODS:Single nucleotide polymorphisms were genotyped in the complement factor H (CFH) and ARMS2 genes using a Taqman assay. Rod-mediated dark adaptation was assessed in 1 eye after photobleach with targets centered at 5° on the inferior vertical meridian. Rate of dark adaptation was defined by rod intercept time (RIT), duration (in minutes) required for sensitivity to reach a criterion sensitivity level in the latter half of the second component of rod recovery. Associations between CFH and ARMS2 polymorphisms and RMDA were adjusted for age and smoking. MAIN OUTCOME MEASURE:Rod intercept time. RESULTS:The sample consisted of 543 participants having both genotype and RIT determination; 408 showed normal macular health and 135 demonstrated AMD, most having early AMD (124 of 135). For the combined sample, higher RIT (slower RMDA) was observed for both the A69S variant in ARMS2 and the Y402H variant in CFH (adjusted P = 0.0001 and P = 0.0023, respectively). For healthy participants, the A69S variant in ARMS2 was associated with higher RIT (adjusted P = 0.0011), whereas the Y402H variant in CFH was not (adjusted P = 0.2175). For AMD patients, the A69S variant of ARMS2 and the Y402H variant of CFH were associated with higher RIT (adjusted P = 0.0182 and P = 0.0222, respectively). Those with a larger number of high-risk ARMS2 and CFH alleles showed higher RIT, in both healthy and AMD groups (adjusted P = 0.0002 and P < 0.0001, respectively). CONCLUSIONS:We report a novel association wherein older adults with high-risk ARMS2 and CFH genotypes are more likely to demonstrate delayed RMDA, the first functional biomarker for incident early AMD. Before the AMD clinical phenotype is present, those showing normal macular health with the ARMS2 A69S allele demonstrate delayed RMDA. Understanding ARMS2 function is a research priority.
Project description:PURPOSE: Single nucleotide polymorphisms (SNPs) of age-related maculopathy susceptibility protein 2/HtrA serine peptidase 1 (ARMS2/HTRA1) and complement factor H (CFH) have been reported to be associated with age-related macular degeneration (AMD). The purpose of this study was to investigate the association of ARMS2/HTRA1 and CFH SNPs with early age-related maculopathy (ARM) in a Han Chinese cohort. METHODS: The cohort consisted of 315 unrelated subjects, including 158 patients with early ARM and 157 recruited controls. Early ARM was diagnosed and graded according to the Age-Related Eye Disease Study criteria. Four SNPs in ARMS2/HTRA1 and six SNPs in CFH previously reported to be associated with AMD were genotyped using TaqMan genotyping assays. Logistic regression implemented with the R statistical language was used for association analysis. RESULTS: None of the ARMS2/HTRA1 and CFH SNPs showed any significant association with early ARM (all p>0.453), with the odds ratios ranging from 0.88 to 1.17. None of the SNPs were associated with unilateral or bilateral early ARM or any grade of early ARM (all p>0.249). CONCLUSIONS: The association of ARMS2/HTRA1 and CFH SNPs in early ARM was not detected in our cohort. The findings in the current study indicated that the effects of ARMS2/HTRA1 and CFH in early ARM could be much lower compared to those in AMD.
Project description:Importance:Previous studies investigating the association of single-nucleotide polymorphisms (SNPs) that confer increased risk of age-related macular degeneration (AMD) with pseudodrusen have yielded conflicting results and have not evaluated other AMD SNPs or pseudodrusen subtypes. Objective:To determine the association of SNPs in the complement factor H (CFH), age-related maculopathy susceptibility 2 (ARMS2), HtrA serine peptidase 1 (HTRA1), complement C2 (C2), complement C3 (C3), lipase C (LIPC), and complement factor B (CFB) genes with the presence of pseudodrusen and pseudodrusen subtypes (ie, dot, reticular, and confluent). Design, Setting, and Participants:In this post hoc analysis of cross-sectional data from US participants in the Comparison of AMD Treatments Trials, genotyping was performed in 835 participants with TaqMan assays for the SNPs rs1061170 (Y402H variant in CFH), rs800292 (I62V variant in CFH), rs10490924 (A69S variant in ARMS2), rs11200638 (HTRA1), rs547154 (C2), rs2230199 (R102G variant in C3), rs10468017 (LIPC), and rs4151667 (L9H variant in CFB). Main Outcomes and Measures:Presence and subtype of baseline pseudodrusen in either eye determined using color fundus photography, red-free images, and fluorescein angiograms. Results:Among 835 participants enrolled for genotyping, 755 (90.4%) were evaluated for pseudodrusen. Of these, 471 (62.4%) were female and 750 (99.3%) were white, and the mean (SD) age was 78.3 (7.5) years. A total of 213 of 755 participants (28.2%) had pseudodrusen (107 [14.2%] had dot pseudodrusen, 180 [23.8%] had reticular pseudodrusen, and 102 [13.5%] had confluent pseudodrusen). After adjusting for age, sex, and smoking status, the ARMS2 risk allele T was associated with higher risk of pseudodrusen (odds ratio [OR], 1.93; 95% CI, 1.19-3.12) for TT vs GG (P?=?.04). A similar association was found for HTRA1 (OR, 2.04; 95% CI, 1.26-3.31) for AA vs GG (P?=?.03). The CFH Y402H risk allele C was associated with lower risk of pseudodrusen (OR, 0.61; 95% CI, 0.38-0.97) for CC vs TT but was not statistically significant after correcting for multiple comparison (P?=?.20). CFH Y402H, ARMS2, HTRA1, and C3 were significantly associated with reticular pseudodrusen. Conclusions and Relevance:Among patients with neovascular AMD, the AMD risk alleles ARMS2 and HTRA1 were associated with an increased risk of pseudodrusen and the risk allele CFH Y402H was associated with lower risk of pseudodrusen, supporting findings from previous studies. Understanding the role of these SNPs in the development of pseudodrusen might improve our understanding of the pathogenesis of AMD and help develop future therapies.