Klaver, C.C.W.

Optimisation of an automated drusen-quantifying software for the analysis of drusen distribution in patients with age-related macular degeneration (Article)

2013-04-01T00:00:00Z

PurposeThe purpose of this study is to optimise the settings of the Retinal Image Analysis Laboratory (RIALAB), a semi-automatic drusen quantification software, in planning for high-throughput quantification of drusen in clinical studies of age-related macular degeneration (AMD).Patients and methodsA comparison of five different settings in RIALAB was made on 67 images from the Rotterdam eye study (population-based study) and 56 images from the fellow eye of patients with active neovascular AMD in King's College Hospital, London (hospital-based study).ResultsThe 'Few Outer' setting was the best setting, with it being most appropriate for 52 (77.6%) of the Rotterdam cohort and 47 (83.9%) for the London cohort. Pearson's χ 2 -test revealed both results to be statistically significant (P<0.0001).ConclusionsRIALAB is a viable algorithm and software package that can detect, quantify, and analyse drusen efficiently in both population-based and hospital-based studies. We have shown that the 'Few Outer' drusen setting can be employed as the default setting, with fine-tuning only needed in a minority of cases, thus helping to speed up workflow.

Identification of a candidate gene for astigmatism (Article)

2013-02-01T00:00:00Z

Purpose. Astigmatism is a common refractive error that reduces vision, where the curvature and refractive power of the cornea in one meridian are less than those of the perpendicular axis. It is a complex trait likely to be influenced by both genetic and environmental factors. Twin studies of astigmatism have found approximately 60% of phenotypic variance is explained by genetic factors. This study aimed to identify susceptibility loci for astigmatism. Methods. We performed a meta-analysis of seven genome-wide association studies that included 22,100 individuals of European descent, where astigmatism was defined as the number of diopters of cylinder prescription, using fixed effect inverse variance-weighted methods. Results. A susceptibility locus was identified with lead single nucleotide polymorphism rs3771395 on chromosome 2p13.3 (meta-analysis, P = 1.97 × 10-7) in the VAX2 gene. VAX2 plays an important role in the development of the dorsoventral axis of the eye. Animal studies have shown a gradient in astigmatism along the vertical plane, with corresponding changes in refraction, particularly in the ventral field. Conclusions. This finding advances the understanding of refractive error, and provides new potential pathways to be evaluated with regard to the development of astigmatism.

Comprehensive candidate gene study highlights UGT1A and BNC2 as new genes determining continuous skin color variation in Europeans (Article)

2012-10-11T00:00:00Z

Natural variation in human skin pigmentation is primarily due to genetic causes rooted in recent evolutionary history. Genetic variants associated with human skin pigmentation confer risk of skin cancer and may provide useful information in forensic investigations. Almost all previous gene-mapping studies of human skin pigmentation were based on categorical skin color information known to oversimplify the continuous nature of human skin coloration. We digitally quantified skin color into hue and saturation dimensions for 5,860 Dutch Europeans based on high-resolution skin photographs. We then tested an extensive list of 14,185 single nucleotide polymorphisms in 281 candidate genes potentially involved in human skin pigmentation for association with quantitative skin color phenotypes. Confirmatory association was revealed for several known skin color genes including HERC2, MC1R, IRF4, TYR, OCA2, and ASIP. We identified two new skin color genes: genetic variants in UGT1A were significantly associated with hue and variants in BNC2 were significantly associated with saturation. Overall, digital quantification of human skin color allowed detecting new skin color genes. The variants identified in this study may also contribute to the risk of skin cancer. Our findings are also important for predicting skin color in forensic investigations.

Clinical implications of old and new genes for open-angle glaucoma (Article)

2011-12-01T00:00:00Z

Objective: Genome-wide association studies have revealed new insights into the genetic determinants of open-angle glaucoma (OAG). This study was performed to determine to what extent variants within established genes (MYOC, OPTN, and WDR36) and newly identified common genetic variants (ATOH7, CDKN2B, and SIX1) contribute to the risk of OAG. Design: Population-based setting, family-based setting, and a case-control study. Participants: The Rotterdam Study I cohort (N = 5312; mean age±standard deviation [SD], 68.0±8.4 years). Findings were replicated in the Genetic Research in Isolated Populations combined with the Erasmus Rucphen Family study (N = 1750; mean age±SD, 48.3±15.2 years), and a cohort from Southampton (N = 702; mean age±SD, 72.5±10.7 years). Methods: After identifying common variants associated with OAG within the established genes, the risk of OAG was analyzed using logistic regression. Discriminative accuracy was assessed by comparing the area under the receiver operator characteristic curve (AUC) for models, including the number of risk alleles, intraocular pressure, age, and gender, with the AUC for the same model but without the risk alleles. Main Outcome Measures: Odds ratios and AUCs of individual and combined risk alleles. Results: No consistent significant associations for the established genes (MYOC, OPTN, and WDR36) with OAG were found. However, when comparing the load of risk variants between cases and controls, 2 of 3 studies showed a significant increased risk of OAG for participants carrying more risk alleles of the 3 established genes. When combining all 6 genes, participants carrying a high number of risk alleles (highest tertile) had a 2.29-fold to 3.19-fold increase in risk of OAG compared with those carrying only a few risk alleles. The addition of the newly identified genes to IOP, age, and gender resulted in a higher AUC compared with the AUC without the newly identified genes (P = 0.027). Conclusions: A significant contribution to the risk of OAG was found for the new common variants identified by recent genome-wide association studies, but not for variants within the established genes. Participants carrying a high number of risk alleles had an approximately 3-fold increase in the risk of OAG compared with those with a low number of risk alleles.

Sifting the wheat from the chaff: Prioritizing GWAS results by identifying consistency across analytical methods (Article)

2011-12-01T00:00:00Z

The curse of multiple testing has led to the adoption of a stringent Bonferroni threshold for declaring genome-wide statistical significance for any one SNP as standard practice. Although justified in avoiding false positives, this conservative approach has the potential to miss true associations as most studies are drastically underpowered. As an alternative to increasing sample size, we compare results from a typical SNP-by-SNP analysis with three other methods that incorporate regional information in order to boost or dampen an otherwise noisy signal: the haplotype method (Schaid et al. [2002] Am J Hum Genet 70:425-434), the gene-based method (Liu et al. [2010] Am J Hum Genet 87:139-145), and a new method (interaction count) that uses genome-wide screening of pairwise SNP interactions. Using a modestly sized case-control study, we conduct a genome-wide association studies (GWAS) of age-related macular degeneration, and find striking agreement across all methods in regions of known associated variants. We also find strong evidence of novel associated variants in two regions (Chromosome 2p25 and Chromosome 10p15) in which the individual SNP P-values are only suggestive, but where there are very high levels of agreement between all methods. We propose that consistency between different analysis methods may be an alternative to increasingly larger sample sizes in sifting true signals from noise in GWAS.

Evidence of association of APOE with age-related macular degeneration - a pooled analysis of 15 studies (Article)

2011-12-01T00:00:00Z

Age-related macular degeneration (AMD) is the most common cause of incurable visual impairment in high-income countries. Previous studies report inconsistent associations between AMD and apolipoprotein E (APOE), a lipid transport protein involved in low-density cholesterol modulation. Potential interaction between APOE and sex, and smoking status has been reported. We present a pooled analysis (n = 21,160) demonstrating associations between late AMD and APOε4 (odds ratio [OR] = 0.72 per haplotype; confidence interval [CI]: 0.65-0.74; P = 4.41×10-11) and APOε2 (OR = 1.83 for homozygote carriers; CI: 1.04-3.23; P = 0.04), following adjustment for age group and sex within each study and smoking status. No evidence of interaction between APOE and sex or smoking was found. Ever smokers had significant increased risk relative to never smokers for both neovascular (OR = 1.54; CI: 1.38-1.72; P = 2.8×10-15) and atrophic (OR = 1.38; CI: 1.18-1.61; P = 3.37×10-5) AMD but not early AMD (OR = 0.94; CI: 0.86-1.03; P = 0.16), implicating smoking as a major contributing factor to disease progression from early signs to the visually disabling late forms. Extended haplotype analysis incorporating rs405509 did not identify additional risks beyond ε2 and ε4 haplotypes. Our expanded analysis substantially improves our understanding of the association between the APOE locus and AMD. It further provides evidence supporting the role of cholesterol modulation, and low-density cholesterol specifically, in AMD disease etiology.

Clinical course of cone dystrophy caused by mutations in the RPGR gene (Article)

2011-10-01T00:00:00Z

Background: Mutations in the RPGR gene predominantly cause rod photoreceptor disorders with a large variability in clinical course. In this report, we describe two families with mutations in this gene and cone involvement. Methods: We investigated an X-linked cone dystrophy family (1) with 25 affected males, 25 female carriers, and 21 non-carriers, as well as a small family (2) with one affected and one unaffected male. The RPGR gene was analyzed by direct sequencing. All medical records were evaluated, and all available data on visual acuity, color vision testing, ophthalmoscopy, fundus photography, fundus autofluorescence, Goldmann perimetry, SD-OCT, dark adaptation, and full-field electroretinography (ERG) were registered. Cumulative risks of visual loss were studied with Kaplan-Meier product-limit survival analysis. Results: Both families had a frameshift mutation in ORF15 of the RPGR gene; family 1 had p.Ser1107ValfsX4, and family 2 had p.His1100GlnfsX10. Mean follow up was 13 years (SD 10). Virtually all affected males showed reduced photopic and normal scotopic responses on ERG. Fifty percent of the patients had a visual acuity of <0.5 at age 35 years (SE 2.2), and 75% of the patients was legally blind at age 60 years (SE 2.3). Female carriers showed no signs of ocular involvement. Conclusions: This report describes the clinical course and visual prognosis in two families with cone dystrophy due to RPGR mutations in the 3' terminal region of ORF15. Remarkable features were the consistent, late-onset phenotype, the severe visual outcome, and the non-expression in female carriers. Expression of RPGR mutations in this particular region appears to be relatively homogeneous and predisposed to cones.

Common variants near FRK/COL10A1 and VEGFA are associated with advanced age-related macular degeneration (Article)

2011-09-01T00:00:00Z

Despite significant progress in the identification of genetic loci for age-related macular degeneration (AMD), not all of the heritability has been explained. To identify variants which contribute to the remaining genetic susceptibility, we performed the largest meta-analysis of genome-wide association studies to date for advanced AMD. We imputed 6 036 699 single-nucleotide polymorphisms with the 1000 Genomes Project reference genotypes on 2594 cases and 4134 controls with follow-up replication of top signals in 5640 cases and 52 174 controls. We identified two new common susceptibility alleles, rs1999930 on 6q21-q22.3 near FRK/COL10A1 [odds ratio (OR) 0.87; P 5 1.1 × 10-8] and rs4711751 on 6p12 near VEGFA (OR 1.15; P 5 8.7 × 10-9). In addition to the two novel loci, 10 previously reported loci in ARMS2/HTRA1 (rs10490924), CFH (rs1061170, and rs1410996), CFB (rs641153), C3 (rs2230199), C2 (rs9332739), CFI (rs10033900), LIPC (rs10468017), TIMP3 (rs9621532) and CETP (rs3764261) were confirmed with genomewide significant signals in this large study. Loci in the recently reported genes ABCA1 and COL8A1 were also detected with suggestive evidence of association with advanced AMD. The novel variants identified in this study suggest that angiogenesis (VEGFA) and extracellular collagen matrix (FRK/COL10A1) pathways contribute to the development of advanced AMD.

The Rotterdam Study: 2012 objectives and design update (Article)

2011-08-31T00:00:00Z

Abstract The Rotterdam Study is a prospective cohort study ongoing since 1990 in the city of Rotterdam in The Netherlands. The study targets cardiovascular, endocrine, hepatic, neurological, ophthalmic, psychiatric, dermatological, oncological, and respiratory diseases. As of 2008, 14,926 subjects aged 45 years or over comprise the Rotterdam Study cohort. The findings of the Rotterdam Study have been presented in over a 1,000 research articles and reports (see www.erasmus-epidemiology.nl/rotterdamstudy). This article gives the rationale of the study and its design. It also presents a summary of the major findings and an update of the objectives and methods.

Variations in apolipoprotein e frequency with age in a pooled analysis of a large group of older people (Article)

2011-06-15T00:00:00Z

Variation in the apolipoprotein E gene (APOE) has been reported to be associated with longevity in humans. The authors assessed the allelic distribution of APOE isoforms ε2, ε3, and ε4 among 10,623 participants from 15 case-control and cohort studies of age-related macular degeneration (AMD) in populations of European ancestry (study dates ranged from 1990 to 2009). The authors included only the 10,623 control subjects from these studies who were classified as having no evidence of AMD, since variation within the APOE gene has previously been associated with AMD. In an analysis stratified by study center, gender, and smoking status, there was a decreasing frequency of the APOE ε4 isoform with increasing age (χ2for trend = 14.9 (1 df); P = 0.0001), with a concomitant increase in the ε3 isoform (χ2for trend = 11.3 (1 df); P = 0.001). The association with age was strongest in ε4 homozygotes; the frequency of ε4 homozygosity decreased from 2.7% for participants aged 60 years or less to 0.8% for those over age 85 years, while the proportion of participants with the ε3/ε4 genotype decreased from 26.8% to 17.5% across the same age range. Gender had no significant effect on the isoform frequencies. This study provides strong support for an association of the APOE gene with human longevity.

Reducing the genetic risk of age-related macular degeneration with dietary antioxidants, zinc, and ω-3 fatty acids: The Rotterdam Study (Article)

2011-06-01T00:00:00Z

Objective: To investigate whether dietary nutrients can reduce the genetic risk of early age-related macular degeneration (AMD) conferred by the genetic variants CFH Y402H and LOC387715 A69S in a nested case-control study. Methods: For 2167 individuals (≥55 years) from the population-based Rotterdam Study at risk of AMD, dietary intake was assessed at baseline using a semiquantitative food frequency questionnaire and genetic variants were determined using TaqMan assay. Incident early AMD was determined on fundus photographs at 3 follow-up visits (median follow-up, 8.6 years). The synergy index was used to evaluate biological interaction between risk factors; hazard ratios were calculated to estimate risk of early AMD in strata of nutrient intake and genotypes. Results: Five hundred seventeen participants developed early AMD. Significant synergy indices supported the possibility of biological interaction between CFH Y402H and zinc, β-carotene, lutein/zeaxanthin, and eicosapentaenoic/ docosahexaenoic acid (EPA/DHA) and between LOC387715 A69S and zinc and EPA/DHA (all P < .05). Homozygotes of CFH Y402H with dietary intake of zinc in the highest tertile reduced their hazard ratio of early AMD from 2.25 to 1.27. For intakes of β-carotene, lutein/zeaxanthin, and EPA/DHA, these risk reductions were from 2.54 to 1.47, 2.63 to 1.72, and 1.97 to 1.30, respectively. Carriers of LOC387715 A69S with the highest intake of zinc and EPA/DHA reduced their risk from 1.70 to 1.17 and 1.59 to 0.95, respectively (all P trends <.05). Conclusions: High dietary intake of nutrients with antioxidant properties reduces the risk of early AMD in those at high genetic risk. Therefore, clinicians should provide dietary advice to young susceptible individuals to postpone or prevent the vision-disabling consequences of AMD.

Common genetic variants associated with open-angle glaucoma (Article)

2011-06-01T00:00:00Z

Open-angle glaucoma (glaucoma) is a major eye disorder characterized by optic disc pathology. Recent genome-wide association studies identified new loci associated with clinically relevant optic disc parameters, such as the optic disc area and vertical cup-disc ratio (VCDR). We examined to what extent these loci are involved in glaucoma. The loci studied include ATOH7, CDC7/TGFBR3 and SALL1 for optic disc area, and CDKN2B, SIX1, SCYL1/LTBP3, CHEK2, ATOH7 and DCLK1 for VCDR. We performed a metaanalysis using data from six independent studies including: the Rotterdam Study (n = 5736), Genetic Research in Isolated Populations combined with Erasmus Rucphen Family study (n = 1750), Amsterdam Glaucoma Study (n = 296) and cohorts from Erlangen and Tü bingen (n = 1363), Southampton (n = 702) and deCODE (n = 36 151) resulting in a total of 3161 glaucoma cases and 42 837 controls. Of the eight loci, we found significant evidence (P = 1.41 3 10-8) for the association of CDKN2B with glaucoma [odds ratio (OR) for those homozygous for the risk allele: 0.76; 95% confidence interval (CI): 0.70-0.84], for the role of ATOH7 (OR: 1.28; 95% CI: 1.12-1.47) and for SIX1 (OR: 1.20; 95% CI: 1.10-1.31) when adjusting for the number of tested loci. Furthermore, there was a borderline significant association of CDC7/TGFBR3.

Genome-wide association studies in Asians confirm the involvement of ATOH7 and TGFBR3, and further identify CARD10 as a novel locus influencing optic discarea (Article)

2011-05-01T00:00:00Z

Damage to the optic nerve (e.g. from glaucoma) has an adverse and often irreversible impact on vision. Earlier studies have suggested that the size of the optic nerve head could be governed by hereditary factors. We conducted a genome-wide association study (GWAS) on 4445 Singaporean individuals (n 5 2132 of Indian and n 5 2313 of Malay ancestry, respectively), with replication in Rotterdam, the Netherlands (n 5 9326 individuals of Caucasian ancestry) using the most widely reported parameter for optic disc traits, the optic disc area. We identified a novel locus on chromosome 22q13.1, CARD10, which strongly associates with optic disc area in both Singaporean cohorts as well as in the Rotterdam Study (RS; rs9607469, perallele change in optic disc area 5 0.051 mm2; Pmeta5 2.73310-12) and confirmed the association between CDC7/TGFBR3 (lead single nucleotide polymorphism (SNP) rs1192415, Pmeta5 7.57310-17) and ATOH7 (lead SNP rs7916697, Pmeta5 2.00 3 10-15) and optic disc area in Asians. This is the first Asian-based GWAS on optic disc area, identifying a novel locus for the optic disc area, but also confirming the results found in Caucasian persons suggesting that there are general genetic determinants applicable to the size of the optic disc across different ethnicities.

Genetic architecture of open angle glaucoma and related determinants (Article)

2011-01-01T00:00:00Z

Background: Although the vertical cup-disc ratio (VCDR) and intraocular pressure (IOP) are important determinants of open angle glaucoma (OAG), it is unclear to what extent the genetic origin of these traits overlap with those of OAG. We evaluated whether the same genes that determine VCDR and IOP also predict OAG. Methods: Genetic risk scores were constructed from single nucleotide polymorphisms (SNPs) using genome wide association data of 9326 participants from the Rotterdam Study cohorts (mean±SD age: 64.6±9.1 years). These risk scores were used to calculate the explained variance of VCDR and IOP in an independent cohort (Erasmus Rucphen Family study) consisting of 1646 participants (mean±SD age: 46.8614.1 years) and the OAG risk in a subset of the Rotterdam Study cohorts. To evaluate false positive findings, we generated two new variables containing randomly sampled values to serve as a negative control. Results: The explained variance of VCDR increased when increasing the number of SNPs included in the risk score, suggesting a polygenic model. We found no clear evidence for a similar model for IOP, suggesting that a small number of SNPs determine the susceptibility to IOP. The SNPs related to IOP in terms of p values contributed little to VCDR. The risk scores associated with VCDR were also associated significantly with OAG. This suggests a common polygenic background for VCDR and OAG Conclusions: We found evidence for a polygenic model underlying one of the major traits of OAG, VCDR, and OAG itself. The IOP did not show any evidence for such a model.

Simultaneous mutation detection in 90 retinal disease genes in multiple patients using a custom-designed 300-kb retinal resequencing chip (Article)

2011-01-01T00:00:00Z

Purpose: To develop a high-throughput, cost-effective diagnostic strategy for the identification of known and new mutations in 90 retinal disease genes. Design: Evidence-based study. Participants: Sixty patients with a variety of retinal disorders, including Leber's congenital amaurosis, ocular albinism, pseudoxanthoma elasticum, retinitis pigmentosa, and Stargardt's disease. Methods: We designed a custom 300-kb resequencing chip. Polymerase chain reaction (PCR) amplification, DNA fragmentation, and chip hybridization were performed according to Affymetrix recommendations. Hybridization signals were analyzed using Sequence pilot module seq-C mutation detection software (2009). This resequencing approach was validated by Sanger sequence technology. Main Outcome Measures: Disease-causing sequence changes. Results: We developed a retinal resequencing chip that covers all exons of 90 retinal disease genes. We developed and tested multiplex primer sets for 1445 amplicons representing the genes included on the chip. We validated our approach by screening 87 exons from 25 retinal disease genes containing 87 known sequence changes previously identified in our patient group using Sanger sequencing. Call rates for successfully hybridized amplicons were 98% to 100%. Of the known single nucleotide changes, 99% could be detected on the chip. As expected, deletions could not be detected reliably. Conclusions: We designed a custom resequencing chip that can detect known and new sequence changes in 90 retinal disease genes using a new high-throughput strategy with a high sensitivity and specificity for one tenth of the cost of conventional direct sequencing. The developed amplification strategy allows for the pooling of multiple patients with non-overlapping phenotypes, enabling many patients to be analyzed simultaneously in a fast and cost-effective manner. Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Genetic testing and clinical characterization of patients with cone-rod dystrophy (Article)

2010-12-01T00:00:00Z

The ERCC6 Gene and Age-Related Macular Degeneration (Article)

2010-11-19T00:00:00Z

Background: Age-related macular degeneration (AMD) is the leading cause of irreversible visual loss in the developed countries and is caused by both environmental and genetic factors. A recent study (Tuo et al., PNAS) reported an association between AMD and a single nucleotide polymorphism (SNP) (rs3793784) in the ERCC6 (NM_000124) gene. The risk allele also increased ERCC6 expression. ERCC6 is involved in DNA repair and mutations in ERCC6 cause Cockayne syndrome (CS). Amongst others, photosensitivity and pigmentary retinopathy are hallmarks of CS. Methodology/Principal Findings: Separate and combined data from three large AMD case-control studies and a prospective population-based study (The Rotterdam Study) were used to analyse the genetic association between ERCC6 and AMD (2682 AMD cases and 3152 controls). We also measured ERCC6 mRNA levels in retinal pigment epithelium (RPE) cells of healthy and early AMD affected human donor eyes. Rs3793784 conferred a small increase in risk for late AMD in the Dutch population (The Rotterdam and AMRO-NL study), but this was not replicated in two non-European studies (AREDS, Columbia University). In addition, the AMRO-NL study revealed no significant association for 9 other variants spanning ERCC6. Finally, we determined that ERCC6 expression in the human RPE did not depend on rs3793784 genotype, but, interestingly, on AMD status: Early AMD-affected donor eyes had a 50% lower ERCC6 expression than healthy donor eyes (P = 0.018). Conclusions/Significance: Our meta analysis of four Caucasian cohorts does not replicate the reported association between SNPs in ERCC6 and AMD. Nevertheless, our findings on ERCC6 expression in the RPE suggest that ERCC6 may be functionally involved in AMD. Combining our data with those of the literature, we hypothesize that the AMD-related reduced transcriptional activity of ERCC6 may be caused by diverse, small and heterogeneous genetic and/or environmental determinants.

Genetic architecture of open angle glaucoma and related determinants (Article)

2010-11-07T00:00:00Z

Background: Although the vertical cup-disc ratio (VCDR) and intraocular pressure (IOP) are important determinants of open angle glaucoma (OAG), it is unclear to what extent the genetic origin of these traits overlap with those of OAG. We evaluated whether the same genes that determine VCDR and IOP also predict OAG. Methods: Genetic risk scores were constructed from single nucleotide polymorphisms (SNPs) using genome wide association data of 9326 participants from the Rotterdam Study cohorts (mean±SD age: 64.6±9.1 years). These risk scores were used to calculate the explained variance of VCDR and IOP in an independent cohort (Erasmus Rucphen Family study) consisting of 1646 participants (mean±SD age: 46.8±14.1 years) and the OAG risk in a subset of the Rotterdam Study cohorts. To evaluate false positive findings, we generated two new variables containing randomly sampled values to serve as a negative control. Results: The explained variance of VCDR increased when increasing the number of SNPs included in the risk score, suggesting a polygenic model. We found no clear evidence for a similar model for IOP, suggesting that a small number of SNPs determine the susceptibility to IOP. The SNPs related to IOP in terms of p values contributed little to VCDR. The risk scores associated with VCDR were also associated significantly with OAG. This suggests a common polygenic background for VCDR and OAG Conclusions: We found evidence for a polygenic model underlying one of the major traits of OAG, VCDR, and OAG itself. The IOP did not show any evidence for such a model.

Progressive loss of cones in achromatopsia: An imaging study using spectral-domain optical coherence tomography (Article)

2010-11-01T00:00:00Z

PURPOSE. Achromatopsia (ACHM) is a congenital autosomal recessive cone disorder with a presumed stationary nature and only a few causative genes. Animal studies suggest that ACHM may be a good candidate for corrective gene therapy. Future implementation of this therapy in humans requires the presence of viable cone cells in the retina. In this study the presence of cone cells in ACHM was determined, as a function of age. METHODS. The appearance and thickness of all retinal layers were evaluated by spectral-domain optical coherence tomography (SD-OCT) in 40 ACHM patients (age range, 4-70 years) with known mutations in the CNGB3, CNGA3, and PDE6C genes. A comparison was made with 55 healthy age-matched control subjects. RESULTS. The initial feature of cone cell decay was loss of inner and outer segments with disruption of the ciliary layer on OCT, which was observed as early as 8 years of age. Cone cell loss further progressed with age and occurred in 8 (42%) of 19 patients below 30 years and in 20 (95%) of 21 of those aged 30+ years. Retinal thickness was significantly thinner in the fovea of all patients (126 μm in ACHM vs. 225 μm in the control; P < 0.001) and correlated with age (β = 0.065; P = 0.011). Foveal hypoplasia was present in 24 (80%) of 30 patients and in 1 of 55 control subjects. CONCLUSIONS. ACHM is not a stationary disease. The first signs of cone cell loss occur in early childhood. If intervention becomes available in the future, the present results imply that it should be applied in the first decade.

A genome-wide association study for myopia and refractive error identifies a susceptibility locus at 15q25 (Article)

2010-10-01T00:00:00Z

Myopia and hyperopia are at opposite ends of the continuum of refraction, the measure of the eyeĝ€2s ability to focus light, which is an important cause of visual impairment (when aberrant) and is a highly heritable trait. We conducted a genome-wide association study for refractive error in 4,270 individuals from the TwinsUK cohort. We identified SNPs on 15q25 associated with refractive error (rs8027411, P = 7.91 × 10-8). We replicated this association in six adult cohorts of European ancestry with a combined 13,414 individuals (combined P = 2.07 × 10-9). This locus overlaps the transcription initiation site of RASGRF1, which is highly expressed in neurons and retina and has previously been implicated in retinal function and memory consolidation. Rasgrf1-/-mice show a heavier average crystalline lens (P = 0.001). The identification of a susceptibility locus for refractive error on 15q25 will be important in characterizing the molecular mechanism responsible for the most common cause of visual impairment.

A genome-wide association study identifies a susceptibility locus for refractive errors and myopia at 15q14 (Letter To Editor)

2010-09-01T00:00:00Z

Refractive errors are the most common ocular disorders worldwide and may lead to blindness. Although this trait is highly heritable, identification of susceptibility genes has been challenging. We conducted a genome-wide association study for refractive error in 5,328 individuals from a Dutch population-based study with replication in four independent cohorts (combined 10,280 individuals in the replication stage). We identified a significant association at chromosome 15q14 (rs634990, P = 2.21 × 10−14). The odds ratio of myopia compared to hyperopia for the minor allele (minor allele frequency = 0.47) was 1.41 (95% CI 1.16–1.70) for individuals heterozygous for the allele and 1.83 (95% CI 1.42–2.36) for individuals homozygous for the allele. The associated locus is near two genes that are expressed in the retina, GJD2 and ACTC1, and appears to harbor regulatory elements which may influence transcription of these genes. Our data suggest that common variants at 15q14 influence susceptibility for refractive errors in the general population.

A genome-wide association study identifies a susceptibility locus for refractive errors and myopia at 15q14 (Letter To Editor)

2010-09-01T00:00:00Z

Mutations in IMPG2, Encoding interphotoreceptor matrix proteoglycan 2, cause autosomal-recessive retinitis pigmentosa (Article)

2010-08-01T00:00:00Z

Retinitis pigmentosa (RP) is a heterogeneous group of inherited retinal diseases caused by progressive degeneration of the photoreceptor cells. Using autozygosity mapping, we identified two families, each with three affected siblings sharing large overlapping homozygous regions that harbored the IMPG2 gene on chromosome 3. Sequence analysis of IMPG2 in the two index cases revealed homozygous mutations cosegregating with the disease in the respective families: three affected siblings of Iraqi Jewish ancestry displayed a nonsense mutation, and a Dutch family displayed a 1.8 kb genomic deletion that removes exon 9 and results in the absence of seven amino acids in a conserved SEA domain of the IMPG2 protein. Transient transfection of COS-1 cells showed that a construct expressing the wild-type SEA domain is properly targeted to the plasma membrane, whereas the mutant lacking the seven amino acids appears to be retained in the endoplasmic reticulum. Mutation analysis in ten additional index cases that were of Dutch, Israeli, Italian, and Pakistani origin and had homozygous regions encompassing IMPG2 revealed five additional mutations; four nonsense mutations and one missense mutation affecting a highly conserved phenylalanine residue. Most patients with IMPG2 mutations showed an early-onset form of RP with progressive visual-field loss and deterioration of visual acuity. The patient with the missense mutation, however, was diagnosed with maculopathy. The IMPG2 gene encodes the interphotoreceptor matrix proteoglycan IMPG2, which is a constituent of the interphotoreceptor matrix. Our data therefore show that mutations in a structural component of the interphotoreceptor matrix can cause arRP.

Course of visual decline in relation to the best1 genotype in vitelliform macular dystrophy (Article)

2010-07-01T00:00:00Z

Purpose: To describe the disease course in patients with vitelliform macular dystrophy (VMD) with a Best1 mutation and to determine the association between Best1 genotype and visual prognosis. Design: Consecutive case series. Participants: Fifty-three patients with VMD with Best1 mutations from 27 Dutch families, aged 11 to 87 years. Methods: Best-corrected visual acuity (VA), fundus appearance, and Arden ratio on the electro-oculogram (EOG) during clinical follow-up were assessed from medical records. Mutation analysis of the Best1 gene was performed on DNA samples using denaturing high-pressure liquid chromatography and direct sequencing. Main Outcome Measures: Cumulative lifetime risk of visual decline below 0.5, 0.3, and 0.1 for the entire group and stratified for genotype. Results: Median age of onset of visual symptoms was 33 years (range: 278). The cumulative risk of VA below 0.5 (20/40) was 50% at 55 years and 75% at 66 years. The cumulative risk of decline less than 0.3 (20/63) was 50% by age 66 years and 75% by age 74 years. Two patients progressed to VA less than 0.1 (20/200). Fourteen different mutations were found. Most patients (96%) had missense mutations; the Thr6Pro, Ala10Val, and Tyr227Asn mutations were most common. Visual decline was significantly faster in patients with an Ala10Val mutation than either the Thr6Pro or the Tyr227Asn mutation (P=0.001). Conclusions: Age of onset of visual symptoms varies greatly among patients with VMD. All patients show a gradual decrease in VA, and most progress to visual impairment at a relatively late age. Our data suggest a phenotypegenotype correlation, because the Ala10Val mutation has a more rapid disease progression than other common mutations. © 2010 by the American Academy of Ophthalmology.

A novel nonsense mutation in CEP290 induces exon skipping and leads to a relatively mild retinal phenotype (Article)

2010-07-01T00:00:00Z

Purpose. To identify the genetic defect in a family with variable retinal phenotypes. The proband had a diagnosis of Leber congenital amaurosis (LCA), whereas her two cousins had an early-onset severe retinal dystrophy (EOSRD) with useful vision. A distant family member had retinitis pigmentosa (RP). Methods. DNA samples of the affected family members were genotyped with 250 K genome-wide SNP microarrays. Genetic defects were localized by linkage analysis and homozygosity mapping, and candidate genes were analyzed by sequencing. Patients underwent a full ophthalmic examination. Results. Compound heterozygous mutations in CEP290 were identified in the proband and her two cousins: the frequent c.2991 + 1655A>G founder mutation and a novel nonsense mutation in exon 7 (c.451C>T, p.Arg151X). The proband had nystagmus, hyperopia, a flat electroretinogram (ERG), and decreased visual acuity (20/250) from birth. The two cousins had minimal scotopic ERG responses at the age of 2. In one of these patients, visual acuity had reached a level of 20/32 at age 5, which is high for patients with CEP290 mutations. Analysis of the CEP290 mRNA in affected individuals revealed altered splice forms in which either exon 7 or exons 7 and 8 were skipped. In both mutant cDNA products, the open reading frame was not disrupted. Furthermore, homozygosity mapping and mutation analysis in the distant family member affected by RP revealed a homozygous mutation in MERTK, but no CEP290 mutations. This MERTK mutation was heterozygously present in the most severely affected (LCA) patient, but was absent in the two more mildly affected cousins. Conclusions. A novel nonsense mutation in CEP290 results in nonsense-associated altered splicing. That the remaining open reading frame is intact may explain the less severe phenotype observed in the two affected cousins. The additional heterozygous mutation in MERTK may clarify the more severe pheno-type in the proband. This study extends the phenotypic spectrum of CEP290-associated diseases at the mild end.

A genome-wide association study of optic disc parameters (Article)

2010-06-01T00:00:00Z

The optic nerve head is involved in many ophthalmic disorders, including common diseases such as myopia and open-angle glaucoma. Two of the most important parameters are the size of the optic disc area and the vertical cup-disc ratio (VCDR). Both are highly heritable but genetically largely undetermined. We performed a meta-analysis of genome-wide association (GWA) data to identify genetic variants associated with optic disc area and VCDR. The gene discovery included 7,360 unrelated individuals from the population-based Rotterdam Study I and Rotterdam Study II cohorts. These cohorts revealed two genome-wide significant loci for optic disc area, rs1192415 on chromosome 1p22 (p =6.72*10-19) within 117 kb of the CDC7 gene and rs1900004 on chromosome 10q21.3-q22.1 (p=2.67*10-33) within 10 kb of the ATOH7 gene. They revealed two genome-wide significant loci for VCDR, rs1063192 on chromosome 9p21 (p =6.15*10-11) in the CDKN2B gene and rs10483727 on chromosome 14q22.3-q23 (p=2.93*10-10) within 40 kbp of the SIX1 gene. Findings were replicated in two independent Dutch cohorts (Rotterdam Study III and Erasmus Rucphen Family study; N=3,612), and the TwinsUK cohort (N=843). Meta-analysis with the replication cohorts confirmed the four loci and revealed a third locus at 16q12.1 associated with optic disc area, and four other loci at 11q13, 13q13, 17q23 (borderline significant), and 22q12.1 for VCDR. ATOH7 was also associated with VCDR independent of optic disc area. Three of the loci were marginally associated with open-angle glaucoma. The protein pathways in which the loci of optic disc area are involved overlap with those identified for VCDR, suggesting a common genetic origin. © 2010 Ramdas et al.

Comprehensive Analysis of the Achromatopsia Genes CNGA3 and CNGB3 in Progressive Cone Dystrophy (Article)

2010-04-01T00:00:00Z

Objective: To investigate whether the major achromatopsia genes (CNGA3 and CNGB3) play a role in the cause of progressive cone dystrophy (CD). Design: Prospective multicenter study. Participants: Probands (N = 60) with autosomal recessive (ar) CD from various ophthalmogenetic clinics in The Netherlands. Methods: All available ophthalmologic data from the arCD probands were registered from medical charts and updated by an additional ophthalmologic examination. Mutations in the CNGA3 and CNGB3 genes were analyzed by direct sequencing. Main Outcome Measures: CNGA3 and CNGB3 mutations and clinical course in arCD probands. Results: In 3 arCD probands (3/60; 5%) we found 2 mutations in the CNGB3 gene. Two of these probands had compound heterozygous mutations (p.R296YfsX9/p.R274VfsX12 and p.R296YfsX9/c.991-3T>g). The third proband revealed homozygous missense mutations (p.R403Q) with 2 additional variants in the CNGA3 gene (p.E228K and p.V266M). These probands did not have a congenital nystagmus, but had a progressive deterioration of visual acuity, color vision, and photopic electroretinogram, with onset in the second decade. In 6 other unrelated probands, we found 6 different heterozygous amino acid changes in the CNGA3 (N = 4) and CNGB3 (N = 2) gene. Conclusions: The CNGB3 gene accounts for a small fraction of the later onset progressive form of cone photoreceptor disorders, and CNGA3 may have an additive causative effect. Our data indicate that these genes are involved in a broader spectrum of cone dysfunction, and it remains intriguing why initial cone function can be spared despite similar gene defects. Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

The Complement Component 5 Gene and Age-Related Macular Degeneration (Article)

2010-03-01T00:00:00Z

Objective: To investigate the association between variants in the complement component 5 (C5) gene and age-related macular degeneration (AMD). Design: Separate and combined data from 3 large AMD case-control studies and a prospective population-based study (The Rotterdam Study). Participants: A total of 2599 AMD cases and 3458 ethnically matched controls. Methods: Fifteen single nucleotide polymorphisms (SNPs) spanning the C5 gene were initially genotyped in 375 cases and 199 controls from The Netherlands (The Amsterdam/Rotterdam-Netherlands [AMRO-NL] study population). Replication testing of selected SNPs was performed in the Rotterdam Study (NL) and study populations from Southampton, United Kingdom (UK), and New York, United States (US). Main Outcome Measures: Early and late stages of prevalent and incident AMD, graded according to (a modification of) the international grading and classification system of AMD. Results: Significant allelic or genotypic associations between 8 C5 SNPs and AMD were found in the AMRO-NL study and this risk seemed to be independent of CFH Y402H, LOC387715 A69S, age, and gender. None of these findings could be confirmed consistently in 3 replication populations. Conclusions: Although the complement pathway, including C5, plays a crucial role in AMD, and the C5 protein is present in drusen, no consistent significant associations between C5 SNPs and AMD were found in any of these studies. The implications for genetic screening of AMD are discussed. Financial Disclosure(s): The authors have no proprietary or commercial interest in any of the materials discussed in this article.

Association of cognitive functioning with retinal nerve fiber layer thickness (Article)

2009-12-01T00:00:00Z

PURPOSE. The brain areas that are responsible for cognitive functioning have the same embryonic origin as the retina. The association between cognitive functioning and retinal nerve fiber layer (RNFL) thickness was assessed in a large, population-based sample. METHODS. Neuropsychological and ophthalmic examinations were performed in 1485 healthy individuals (mean age, 46 years; range, 18 - 85) from the Erasmus Rucphen Family (ERF) study, a study in a genetic isolate from the Netherlands. Different domains of cognitive functioning were assessed with the Dutch Adult Reading Test, the Rey Auditory Verbal Memory Test, semantic fluency, the Trail-Making Test, the Stroop Color-Word Test, and Block Design. RNFL thickness was measured with scanning laser polarimetry. The association between cognitive test scores and peripapillary RNFL thickness was studied with linear regression analyses, adjusting for age, sex, level of inbreeding, and refractive error. RESULTS. After adjustment for confounders, a better cognitive performance was significantly associated with a thicker RNFL in all tests (P < 0.03) except for the Stroop Color-Word Test (P = 0.15). RNFL thickness explained up to 2.8% (R2= 0.028) of the total variance in cognitive test scores. The association diminished in age groups beyond 40 years. CONCLUSIONS. The present study shows that cognitive functioning is associated with RNFL thickness in healthy young individuals. The lack of association in older individuals suggests that loss of neurons in the cerebrum and retina is not concomitant and may have different origins.

Heterozygous NTF4 Mutations Impairing Neurotrophin-4 Signaling in Patients with Primary Open-Angle Glaucoma (Article)

2009-10-09T00:00:00Z

Glaucoma, a main cause of blindness in the developed world, is characterized by progressive degeneration of retinal ganglion cells (RGCs), resulting in irreversible loss of vision. Although members of the neurotrophin gene family in various species are known to support the survival of numerous neuronal populations, including RGCs, it is less clear whether they are also required for survival and maintenance of adult neurons in humans. Here, we report seven different heterozygous mutations in the Neurotrophin-4 (NTF4) gene accounting for about 1.7% of primary open-angle glaucoma patients of European origin. Molecular modeling predicted a decreased affinity of neurotrophin 4 protein (NT-4) mutants with its specific tyrosine kinase receptor B (TrkB). Expression of recombinant NT-4 carrying the most frequent mutation was demonstrated to lead to decreased activation of TrkB. These findings suggest a pathway in the pathophysiology of glaucoma through loss of neurotrophic function and may eventually open the possibility of using ligands activating TrkB to prevent the progression of the disease.

Genetic Etiology and Clinical Consequences of Complete and Incomplete Achromatopsia (Article)

2009-10-01T00:00:00Z

Objective: To investigate the genetic causes of complete and incomplete achromatopsia (ACHM) and assess the association between disease-causing mutations, phenotype at diagnosis, and visual prognosis. Design: Clinic-based, longitudinal, multicenter study. Participants: Probands with complete ACHM (n = 35), incomplete ACHM (n = 26), or nonspecific ACHM (n = 2) and their affected relatives (n = 18) from various ophthalmogenetic clinics in The Netherlands. Methods: Ophthalmologic clinical data were assessed over a life time and were registered from medical charts and updated by ophthalmologic examination. Mutations in the CNGB3, CNGA3, and GNAT2 genes were analyzed by direct sequencing. Main Outcome Measures: Genetic mutations and clinical course of ACHM. Results: CNGB3 mutations were identified in 55 of 63 (87%) of probands and all caused premature truncation of the protein. The most common mutation was p.T383IfsX13 (80%); among the 4 other mutations was the novel frameshift mutation p.G548VfsX35. CNGA3 mutations were detected in 3 of 63 (5%) probands; all caused an amino acid change of the protein. No mutations were found in the GNAT2 gene. The ACHM subtype, visual acuity, color vision, and macular appearance were equally distributed among the CNGB3 genotypes, but were more severely affected among CNGA3 genotypes. Visual acuity deteriorated from infancy to adulthood in 12% of patients, leading to 0.10 in 61%, and even lower than 0.10 in 20% of patients. Conclusions: In this well-defined cohort of ACHM patients, the disease seemed much more genetically homogeneous than previously described. The CNGB3 gene was by far the most important causal gene, and T383IfsX13 the most frequent mutation. The ACHM subtype did not associate with a distinct genetic etiology, nor were any other genotype-phenotype correlations apparent. The distinction between complete and incomplete subtypes of ACHM has no clinical value, and the assumption of a stationary nature is misleading. Financial Disclosure(s): The authors have no proprietary or commercial interest in any of the materials discussed in this article.

The SERPING1 gene and age-related macular degeneration (Article)

2009-09-11T00:00:00Z

Homozygosity Mapping Reveals PDE6C Mutations in Patients with Early-Onset Cone Photoreceptor Disorders (Article)

2009-08-14T00:00:00Z

Cone photoreceptor disorders form a clinical spectrum of diseases that include progressive cone dystrophy (CD) and complete and incomplete achromatopsia (ACHM). The underlying disease mechanisms of autosomal recessive (ar)CD are largely unknown. Our aim was to identify causative genes for these disorders by genome-wide homozygosity mapping. We investigated 75 ACHM, 97 arCD, and 20 early-onset arCD probands and excluded the involvement of known genes for ACHM and arCD. Subsequently, we performed high-resolution SNP analysis and identified large homozygous regions spanning the PDE6C gene in one sibling pair with early-onset arCD and one sibling pair with incomplete ACHM. The PDE6C gene encodes the cone α subunit of cyclic guanosine monophosphate (cGMP) phosphodiesterase, which converts cGMP to 5′-GMP, and thereby plays an essential role in cone phototransduction. Sequence analysis of the coding region of PDE6C revealed homozygous missense mutations (p.R29W, p.Y323N) in both sibling pairs. Sequence analysis of 104 probands with arCD and 10 probands with ACHM revealed compound heterozygous PDE6C mutations in three complete ACHM patients from two families. One patient had a frameshift mutation and a splice defect; the other two had a splice defect and a missense variant (p.M455V). Cross-sectional retinal imaging via optical coherence tomography revealed a more pronounced absence of cone photoreceptors in patients with ACHM compared to patients with early-onset arCD. Our findings identify PDE6C as a gene for cone photoreceptor disorders and show that arCD and ACHM constitute genetically and clinically overlapping phenotypes.

The spectrum of phenotypes caused by variants in the CFH gene (Article)

2009-05-01T00:00:00Z

Complement factor H (CFH) is a complement inhibitor, which is present as a soluble protein and attached to cell surfaces throughout the human body. As such, CFH is a key player in complement homeostasis, inhibiting excessive activation of the complement cascade, with an emphasis on the alternative pathway. The significance of CFH is demonstrated by the broad range of phenotypes associated with specific CFH gene variants. This phenotypic spectrum includes renal phenotypes, such as membranoproliferative glomerulonephritis and atypical hemolytic uremic syndrome, as well as ocular phenotypes, such as basal laminar drusen and age-related macular degeneration. In addition, several overlapping phenotypes have been described in association with CFH gene variants. The phenotypic outcome of these CFH variants depends on their differential impact on plasma- and surface-bound CFH function. Consequently, distinct genotype-phenotype correlations may be observed.

Complement Component C3 and Risk of Age-Related Macular Degeneration (Article)

2009-01-26T00:00:00Z

Objective: To explore the association between polymorphisms in the complement component 3 (C3) gene and age-related macular degeneration (AMD), and to investigate the modifying effect of complement factor H(CFH) Y402H, LOC387715 A69S and smoking. Design: Pooled data from the prospective, population-based Rotterdam Study (enrolment between 1990 and 1993, and 3 follow-up examinations between September 1, 1993, and December 31, 2004) and an independent case-control study from the Netherlands. Participants: The Rotterdam Study comprised a total of 6418 persons aged ≥55 years who had gradable fundus photographs. The case-control study consisted of 357 unrelated AMD patients and 173 control individuals aged ≥55 years. Methods: The variants R102G and P314L of the C3 gene, CFH Y402H and LOC387715 A69S, were genotyped in all study participants. Information on cigarette smoking was obtained by interview at baseline. Main Outcome Measures: Early and late stages of prevalent and incident AMD, graded according to the international classification and grading system for AMD. Results: We found a population frequency of 0.217 for R102G and 0.211 for P314L in the Rotterdam Study. Both alleles significantly increased the risk of early AMD and all subtypes of late AMD, and this risk seemed to be independent of CFH Y402H, LOC387715 A69S, and smoking. Detailed analysis showed that the haplotype carrying both alleles had the highest frequency difference between cases and controls (P = 0.006). We estimated a total population-attributable risk of 14.6%. A meta-analysis of all currently available data yielded a pooled odds ratio (OR) of 1.61 (95% confidence interval [CI], 1.46-1.78) for the R102G allele, and an OR of 1.50 (95% CI, 1.31-1.71) for the P314L allele. Conclusions: Our study showed a significant association between variants in the C3 gene and AMD and further highlights the crucial role of the complement pathway in the etiology of AMD. Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this article.

Three Genome-wide Association Studies and a Linkage Analysis Identify HERC2 as a Human Iris Color Gene (Article)

2008-02-08T00:00:00Z

Human iris color was one of the first traits for which Mendelian segregation was established. To date, the genetics of iris color is still not fully understood and is of interest, particularly in view of forensic applications. In three independent genome-wide association (GWA) studies of a total of 1406 persons and a genome-wide linkage study of 1292 relatives, all from the Netherlands, we found that the 15q13.1 region is the predominant region involved in human iris color. There were no other regions showing consistent genome-wide evidence for association and linkage to iris color. Single nucleotide polymorphisms (SNPs) in the HERC2 gene and, to a lesser extent, in the neighboring OCA2 gene were independently associated to iris color variation. OCA2 has been implicated in iris color previously. A replication study within two populations confirmed that the HERC2 gene is a new and significant determinant of human iris color variation, in addition to OCA2. Furthermore, HERC2 rs916977 showed a clinal allele distribution across 23 European populations, which was significantly correlated to iris color variation. We suggest that genetic variants regulating expression of the OCA2 gene exist in the HERC2 gene or, alternatively, within the 11.7 kb of sequence between OCA2 and HERC2, and that most iris color variation in Europeans is explained by those two genes. Testing markers in the HERC2-OCA2 region may be useful in forensic applications to predict eye color phenotypes of unknown persons of European genetic origin.

Comprehensive analysis of the candidate genes CCL2, CCR2, and TLR4 in age-related macular degeneration (Article)

2008-01-01T00:00:00Z

PURPOSE. To determine whether variants in the candidate genes TLR4, CCL2, and CCR2 are associated with age-related macular degeneration (AMD). METHODS. This study was performed in two independent Caucasian populations that included 357 cases and 173 controls from the Netherlands and 368 cases and 368 controls from the United States. Exon 4 of the TLR4 gene and the promoter, all exons, and flanking intronic regions of the CCL2 and CCR2 genes were analyzed in the Dutch study and common variants were validated in the U.S. study. Quantitative (q)PCR reactions were performed to evaluate expression of these genes in laser-dissected retinal pigment epithelium from 13 donor AMD and 13 control eyes. RESULTS. Analysis of single nucleotide polymorphisms (SNPs) in the TLR4 gene did not show a significant association between D299G or T399I and AMD, nor did haplotypes containing these variants. Univariate analyses of the SNPs in CCL2 and CCR2 did not demonstrate an association with AMD. For CCR2, haplotype frequencies were not significantly different between cases and controls. For CCL2, one haplotype containing the minor allele of C35C was significantly associated with AMD (P = 0.03), but this did not sustain after adjustment for multiple testing (q = 0.30). Expression analysis did not demonstrate altered RNA expression of CCL2 and CCR2 in the retinal pigment epithelium from AMD eyes (for CCL2 P = 0.62; for CCR2 P = 0.97). CONCLUSIONS. No evidence was found of an association between TLR4, CCR2, and CCL2 and AMD, which implies that the common genetic variation in these genes does not play a significant role in the etiology of AMD. Copyright

Macrophages feel their age in macular degeneration (Article)

2007-11-01T00:00:00Z

Macular degeneration, during which the posterior part of the eye known as the macula suffers from thinning, atrophy, and bleeding caused by abnormal angiogenesis (blood vessel formation), predominantly affects elderly adults and results in the loss of central vision. In this issue of the JCI, Kelly et al. investigate the regulation of innate immune cells, specifically macrophages, in ocular neovascularization following eye injury in mice (see the related article beginning on page 3421). They found that, as the mice aged, increased expression of IL-10 by senescent macrophages and changes in their expression of other cytokines altered the ability of these cells to restrain trauma-induced angiogenesis in the eye. These data provide insight into the effect of senescence on macrophage function and angiogenesis and have important implications for age-related diseases such as macular degeneration.

Predictive value of multiple genetic testing for age-related macular degeneration (Article)

2007-09-01T00:00:00Z

Usefulness of Combining Complement Factor H and C-Reactive Protein Genetic Profiles for Predicting Myocardial Infarction (from the Rotterdam Study) (Article)

2007-08-15T00:00:00Z

Complement factor H (CFH) is an important regulator of the complement cascade. Binding of C-reactive protein (CRP) to CFH augments the ability of CFH to downregulate the effect of complement in atherosclerotic lesions. The CFH Tyr402His polymorphism has been suggested to influence the ability of CFH to bind CRP. We hypothesized that the combined presence of unfavorable CRP and CFH genetic profiles is associated with risk of myocardial infarction (MI). The Rotterdam Study is a population-based cohort study in 7,983 men and women aged ≥55 years. The CFH Tyr402His (rs1061170) polymorphism was determined (His402allele 37%), and using 3 tagging polymorphisms (rs1130864, rs1205, and rs3093068), CRP haplotypes were inferred (1 = CTC, 2 = TCC, 3 = CCC, 4 = CCG; frequencies of 33%, 32%, 30%, and 6%, respectively). Participants were grouped by CFH genotype (TyrTyr [reference], TyrHis, and HisHis) and CRP haplotype (haplotype 1 homozygotes [reference], haplotype 2 carriers, haplotype 3 carriers, and haplotype 4 carriers), which resulted in a total of 12 groups. CFH His402homozygotes who were also CRP haplotype 3 carriers had an age- and gender-adjusted hazard ratio of 5.9 (95% confidence interval 2.1 to 16.5) to develop MI compared with the reference group. In conclusion, this population-based study suggests that the combined presence of unfavorable CFH and CRP genetic profiles is associated with risk of MI.

Genetic contributions to glaucoma: Heritability of intraocular pressure, retinal nerve fiber layer thickness, and optic disc morphology (Article)

2007-08-01T00:00:00Z

PURPOSE. The genetic etiology of primary open-angle glaucoma (POAG) is still largely unknown, because of its complexity and disparities in its classification. This study was undertaken to determine the genetic contribution to various early, continuous markers of POAG by assessing the heritability of intraocular pressure (IOP), retinal nerve fiber layer (RNFL) thickness, and neuroretinal rim and optic disc parameters in a genetically isolated population. METHODS. A total of 2620 subjects (mean age, 48 years; range 18-86) from extended pedigrees living in a small town in The Netherlands underwent an extensive ophthalmic examination. Their IOP was measured by Goldmann applanation tonometry, their RNFL thickness by scanning laser polarimetry (GDx VCC), and their optic disc parameters by confocal scanning laser ophthalmoscopy (HRT II). Risk associations were explored by linear regression analyses and heritability estimates by variance component methods. RESULTS. Inbreeding was present in 2042 (81%) participants, and was significantly associated with a higher IOP (P < 0.001). The heritability estimate for IOP was 0.35 (95% confidence interval [CI], 0.27-0.43); for RNFL thickness, 0.48 (95% CI, 0.35-0.60); and for neuroretinal rim area, 0.39 (95% CI, 0.20-0.58). Nongenetic factors accounted for only a small proportion (≤0.13) of the variance in all three traits. CONCLUSIONS. Early, continuous markers of POAG are strongly determined by additive genetic effects. The results support a quantitative trait linkage strategy to discover new genes for POAG. Copyright

Dietary intake of antioxidants and risk of age-related macular degeneration. (Article)

2005-12-28T00:00:00Z

CONTEXT: Age-related macular degeneration (AMD) is the most prevalent cause of irreversible blindness in developed countries. Recently, high-dose supplementation with beta carotene, vitamins C and E, and zinc was shown to slow the progression of AMD. OBJECTIVE: To investigate whether regular dietary intake of antioxidants is associated with a lower risk of incident AMD. DESIGN: Dietary intake was assessed at baseline in the Rotterdam Study (1990-1993) using a semiquantitative food frequency questionnaire. Incident AMD until final follow-up in 2004 was determined by grading fundus color transparencies in a masked way according to the International Classification and Grading System. SETTING: Population-based cohort of all inhabitants aged 55 years or older in a middle-class suburb of Rotterdam, the Netherlands. PARTICIPANTS: Of 5836 persons at risk of AMD at baseline, 4765 had reliable dietary data and 4170 participated in the follow-up. MAIN OUTCOME MEASURE: Incident AMD, defined as soft distinct drusen with pigment alterations, indistinct or reticular drusen, geographic atrophy, or choroidal neovascularization. RESULTS: Incident AMD occurred in 560 participants after a mean follow-up of 8.0 years (range, 0.3-13.9 years). Dietary intake of both vitamin E and zinc was inversely associated with incident AMD. The hazard ratio (HR) per standard deviation increase of intake for vitamin E was 0.92 (95% confidence interval [CI], 0.84-1.00) and for zinc was 0.91 (95% CI, 0.83-0.98). An above-median intake of all 4 nutrients, beta carotene, vitamin C, vitamin E, and zinc, was associated with a 35% reduced risk (HR, 0.65; 95% CI, 0.46-0.92) of AMD. Exclusion of supplement users did not affect the results. CONCLUSION: In this study, a high dietary intake of beta carotene, vitamins C and E, and zinc was associated with a substantially reduced risk of AMD in elderly persons.

Concurrent loss of chromosome arm 1p and chromosome 3 predicts a decreased disease-free survival in uveal melanoma patients. (Article)

2005-12-01T00:00:00Z

PURPOSE: Uveal melanoma is a highly malignant disease with a mortality rate of 50% at 10 to 15 years. Previous studies have shown that chromosomal changes are associated with decreased survival of the patient. However, in these studies the small number of tumors analyzed did not allow robust statistical analysis. In the present study, the independent numerical changes in chromosomes 1, 3, 6, and 8 on disease-free survival (DFS) was assessed in a large series of patients with uveal melanoma. METHODS: One hundred twenty tumors from patients with uveal melanoma were analyzed for numerical changes in chromosomes 1, 3, 6, and 8, with cytogenetic analysis, fluorescent in situ hybridization, and/or comparative genomic hybridization. Data were correlated with disease outcome in univariate and multivariate analyses, by Kaplan-Meier and Cox regression analyses. RESULTS: At a mean follow-up time of 45 months, 42 patients had died or had metastatic disease. In the univariate analysis, loss of chromosome 3, gain of 8q, largest tumor diameter, or the presence of epithelioid cells was associated with a decreased DFS. In the multivariate analysis, the effect of monosomy 3 on survival was largely modified by changes in 1p36. Regarding all chromosomal changes, only the concurrent loss of the short arm of chromosome 1 and all of chromosome 3 was an independent prognostic parameter for disease-free survival (P < 0.001). CONCLUSIONS: In uveal melanoma, concurrent loss of the short arm of chromosome 1 and all of chromosome 3 is an independent predictor of decreased DFS.

Cholesterol and age-related macular degeneration: is there a link? (Article)

2004-04-01T00:00:00Z

PURPOSE: To examine the relation among serum cholesterol, apolipoprotein E genotype (APOE), and the risk of early and late age-related macular degeneration (AMD). DESIGN: The Rotterdam Study, a population based prospective cohort study. METHODS: Serum levels of total and high-density lipoprotein (HDL) cholesterol as well as APOE genotype were determined at baseline. Of 3,944 subjects, 400 were diagnosed with incident early and late AMD after a mean follow-up of 5.2 years. RESULTS: Serum HDL, but not total, cholesterol was associated with an increased risk of AMD (odds ratio/SD, 1.20; 95% confidence interval; 1.06-1.35). The association remained unchanged after adjustment for APOE genotype. When stratifying for APOE genotype, the association was strongest in persons with the e 4 allele; an inverse association seemed to be present for e 2 carriers. CONCLUSION: Elevated HDL but not total cholesterol is associated with an increased risk of AMD. Apolipoprotein E genotype does not explain this association but may be an effect modifier.

A pooled case-control study of the apolipoprotein E (APOE) gene in age-related maculopathy (Article)

2002-12-01T00:00:00Z

Age-related maculopathy (ARM) is a multifactorial disorder known to have a substantial genetic component. The epsilon4 allele of the apolipoprotein E gene (APOE-4) has previously been reported to have a protective effect on ARM risk, while the APOE-2 allele may increase disease risk. This study combined four independent data sets (three US and one European) of Caucasian ARM patients and controls in order to obtain better statistical power to examine the role of APOE in ARM. APOE genotype and allele frequencies were compared for 617 ARM cases and 1260 controls, adjusting for age and sex differences between the two groups via multiple logistic regression. The protective effect of the APOE-4 allele on ARM risk was confirmed (age- and sex-adjusted odds ratio (OR) for APOE-4 carriers 0.54, 95% confidence interval (CI) 0.41-0.70, p < 0.0001). The effect of APOE-4 did not differ significantly between males and females and was observed consistently for both atrophic and neovascular ARM. Evidence for an increased risk of ARM due to the APOE-2 allele was found for men, but not for women (OR for men 1.54, 95% CI 0.97-2.45; OR for women 0.74, 95% CI 0.52-1.06, p = 0.01 for interaction of sex and APOE-2 carrier status). These data confirm that the APOE-4 allele, or an allele in linkage disequilibrium with it, reduces the risk of ARM. They also suggest that the effect of the APOE-2 allele may vary by gender, and that APOE-2 may confer an increased risk only to males.

Incidence and progression rates of age-related maculopathy: the Rotterdam Study (Article)

2001-01-01T00:00:00Z

PURPOSE: To describe the incidence rate of age-related macular degeneration (AMD) and the progression rates of early stages of age-related maculopathy (ARM), and to study the hierarchy of fundus features that determine progression. METHODS: A group of 4953 subjects aged 55 years and older living in Rotterdam, The Netherlands, was studied at baseline and at 2-year follow-up to determine the incidence of neovascular and atrophic AMD. A subgroup of 1244 subjects was studied for progression of early stages of ARM. Fundus transparencies were graded for features of ARM using the International Classification System. ARM was stratified in four exclusive stages, according to type of drusen and presence of pigmentary irregularities. RESULTS: The overall 2-year cumulative incidence of AMD was 0.2%, increasing to 1.8% in subjects of 85 years and older. Of those in the early stages, one fourth showed progression to a more severe stage. The most important predictors for progression were more than 10% of macular area covered by drusen (odds ratio [OR] 5.7, 95% confidence interval [CI] 2.9-11.3), presence of depigmentation (OR 4.0, 95% CI 2.5-6.4), and hyperpigmentation (OR 3.4, 95% CI 2.1-5.4). CONCLUSIONS: The incidence of AMD appears to be lower in The Netherlands than in the United States. Progression of early ARM stages occurs in a distinct pattern at a stable rate, with a large area of drusen and presence of pigmentary changes as the most important predictors.

Genetic epidemiologic studies on age-related maculopathy: a population-based approach (Doctoral Thesis)

2000-01-05T00:00:00Z

The western world is aging rapidly. In the Netherlands, the current mean life expectancy for men and women is 74.6 and 80.4 years, respectively, and those over 65 years of age comprise 13.6% of the total population. This proportion of elderly is expected to increase considerably within the coming years, and this will lead to higher frequencies of diseases. Age-related maculopathy (ARM) is one of those frequent geriatric diseases. It is an eye disease ultimately leading to blindness. The prevalence of the clinical end stages of this disorder range from 1 % in those aged 60 years of age to 10% in those aged 85 years and older. At least 60000 Dutch subjects are severely affected by these end stages, also called age-related macular degeneration (AMD). AMD has a great impact on visual function and the performance of daily tasks, in particular because there are still no means for long term restoration of vision. During the last decade there has been steadily increasing research activity investigating the disease etiology. It became better known that the pathogenesis was complex with a variety of risk factors involved. Family reports and twin studies pointed to a genetic background, and epidemiologic studies suggested environmental influences from vascular and dietmy factors, sunlight and smoking. However, findings were not unequivocal, and the evidence on most of these relations was insufficient and inconclusive. This called for more extensive research into the causes of ARM. This thesis aimed to answer the following questions: Pal'l I: What is the current genetic epidemiologic knowledge on ARM? Pari Jl: What is the incidence of AMD, what is the natural course of the disease, and what is the relation with visual impairment?

Genetic Risk of age-related maculopathy (Article)

1998-01-01T00:00:00Z

OBJECTIVE: To investigate to what extent age-related maculopathy (ARM) is genetically determined. DESIGN AND SETTING: Familial aggregation study based on probands derived from the population-based Rotterdam Study. PARTICIPANTS: First-degree relatives of 87 patients with late ARM, i.e., atrophic or neovascular macular degeneration, were compared with first-degree relatives of 135 control subjects without ARM. MAIN OUTCOME MEASURES: Presence and stage of ARM as diagnosed on fundus transparencies, odds ratio, lifetime risk, risk ratio, and population-attributable risk. RESULTS: Independent of other risk factors, the prevalence of early (odds ratio = 4.8, 95% confidence interval [CI] = 1.8-12.2) and late (odds ratio = 19.8, 95% CI = 3.1-126.0) ARM was significantly higher in relatives of patients with late ARM. The lifetime risk estimate of late ARM was 50% (95% CI = 26%-73%) for relatives of patients vs 12% (95% CI = 2%-16%) for relatives of controls (P < .001), yielding a risk ratio of 4.2 (95% CI = 2.6-6.8). Relatives of patients expressed the various features of ARM at a younger age. The population-attributable risk related to genetic factors was 23%. CONCLUSIONS: First-degree relatives of patients with late ARM developed ARM at an increased rate at a relatively young age. Our findings indicate that approximately one fourth of all late ARM is genetically determined and suggest that genetic susceptibility may play an important role in determining the onset of disease.

Genetic Risk of primary open-angle glaucoma. A population-based familial aggregation study (Article)

1998-01-01T00:00:00Z

OBJECTIVES: To study familial aggregation of primary open-angle glaucoma in a general population and to determine the absolute and relative risks for first-degree relatives. METHODS: First-degree relatives of patients with glaucoma (n = 48) and control subjects (n = 155) from the population-based Rotterdam Study underwent a standardized examination, including perimetry. MAIN OUTCOME MEASURES: Intraocular pressure, vertical cup-disc ratio; and the presence of glaucoma, defined as a visual field defect with a cup-disc ratio of 0.7 or higher or asymmetry of 0.3 or higher between both eyes. RESULTS: The prevalence of glaucoma was 10.4% in siblings of patients, 1.1% in offspring of patients, 0.7% in siblings of controls, and 0% in offspring of controls. Life-time risk of elevated intraocular pressure in relatives of patients vs relatives of controls was 42.5% vs 6.7%, of enlarged cup-disc ratio was 62.2% vs 16.6%, and of glaucoma was 22.0% vs 2.3%, yielding a risk ratio for glaucoma of 9.2 (95% confidence interval = 1.2-73.9). The population-attributable risk of glaucoma was 16.4%. CONCLUSIONS: In a general population, relatives of patients with glaucoma have a strongly increased risk of glaucoma. Enlarged cup-disc ratio, not intraocular pressure, was the earliest and most prominent feature of familial aggregation. Further studies are needed to disentangle the genetic components of the increased familial risk.

Genetic association of apolipoprotein E with age-related macular degeneration (Article)

1998-01-01T00:00:00Z

Age-related macular degeneration (AMD) is the most common geriatric eye disorder leading to blindness and is characterized by degeneration of the neuroepithelium in the macular area of the eye. Apolipoprotein E (apoE), the major apolipoprotein of the CNS and an important regulator of cholesterol and lipid transport, appears to be associated with neurodegeneration. The apoE gene (APOE) polymorphism is a strong risk factor for various neurodegenerative diseases, and the apoE protein has been demonstrated in disease-associated lesions of these disorders. Hypothesizing that variants of APOE act as a potential risk factor for AMD, we performed a genetic-association study among 88 AMD cases and 901 controls derived from the population-based Rotterdam Study in the Netherlands. The APOE polymorphism showed a significant association with the risk for AMD; the APOE epsilon4 allele was associated with a decreased risk (odds ratio 0.43 [95% confidence interval 0.21-0. 88]), and the epsilon2 allele was associated with a slightly increased risk of AMD (odds ratio 1.5 [95% confidence interval 0.8-2. 82]). To investigate whether apoE is directly involved in the pathogenesis of AMD, we studied apoE immunoreactivity in 15 AMD and 10 control maculae and found that apoE staining was consistently present in the disease-associated deposits in AMD-maculae-that is, drusen and basal laminar deposit. Our results suggest that APOE is a susceptibility gene for AMD.