The myopia susceptibility locus vasoactive intestinal peptide receptor 2 (VIPR2) contains variants with opposite effects.

Myopia is the commonest eye disorder in the world. High myopes are predisposed to ocular pathologies. The vasoactive intestinal peptide receptor 2 (VIPR2) gene was identified as a myopia susceptibility locus by our group and another group. We continued to fine-map this locus. A case-control study was performed in 4 sequential stages with a total of 941 highly myopic subjects and 846 control subjects, all unrelated Chinese. Stage 1 experimentally genotyped 64.4% of the entire cohort for 152 single-nucleotide polymorphisms (SNPs) and Stage 2 the remaining subjects for 21 SNPs. Stage 3 combined the genotypes for 21 SNPs for the entire cohort, and identified one group of high-risk haplotypes and one group of protective haplotypes significantly associated with high myopia. Stage 4 imputed genotypes for variants in the VIPR2 region and identified two independent groups of variants: one group with high-risk minor alleles and another with protective minor alleles. Variants within each group were generally in strong linkage disequilibrium among themselves while high-risk variants were in linkage equilibrium with protective variants. Therefore, the VIPR2 locus seems to contain variants with opposite effects. This is the first study that has examined the genetic architecture of a myopia susceptibility locus in detail.

Awareness of diabetic retinopathy and its association with attendance for systematic screening at the public primary care setting: a cross-sectional study in Hong Kong.

OBJECTIVE: To assess the association between awareness of diabetic retinopathy (DR) and actual attendance for DR screening. DESIGN: Cross-sectional study. SETTING: Two public general outpatient clinics. PARTICIPANTS: The subjects were people with diabetes mellitus (DM) who participated in a randomised controlled trial, set up in 2008, to test the impact of a copayment on attendance for DR screening. PRIMARY AND SECONDARY OUTCOME MEASURES: The subjects' awareness of DR was evaluated using a structured questionnaire conducted via a telephone interview. The attendance for screening was from the actual attendance data. Association between awareness and attendance for screening was determined using multivariate logistic regression model and was reported as ORs. RESULTS: A total of 2593 participants completed the questionnaire. A total of 42.9% (1113/2593) said they would worry if they had any vision loss and 79.6% (2063/2593) knew that DM could cause blindness. Only 17.5% (453/2593) knew that treatment was available for DR and 11.5% (297/2593) knew that early DR could be asymptomatic. The importance of having a regular eye examination was acknowledged by 75.7% (1964/2593), but 34% (881/2593) did not know how frequently their eyes should be examined. Worry about vision loss (OR=1.72, P<0.001), awareness of the importance of regular eye examination (OR=1.83, P=0.002) and awareness of the frequency of eye examinations ('every year' (OR=2.64, P<0.001) or 'every 6 months' (OR=3.27, P<0.001)) were the most significant factors associated with attendance. CONCLUSIONS: Deficits in knowledge of DR and screening were found among subjects with DM, and three awareness factors were associated with attendance for screening. These factors could be targeted for future interventions.

Genetic Association Study of KCNQ5 Polymorphisms with High Myopia.

Identification of genetic variations related to high myopia may advance our knowledge of the etiopathogenesis of refractive error. This study investigated the role of potassium channel gene (KCNQ5) polymorphisms in high myopia. We performed a case-control study of 1563 unrelated Han Chinese subjects (809 cases of high myopia and 754 emmetropic controls). Five tag single-nucleotide polymorphisms (SNPs) of KCNQ5 were genotyped, and association testing with high myopia was conducted using logistic regression analysis adjusted for sex and age to give Pasym values, and multiple comparisons were corrected by permutation test to give Pemp values. All five noncoding SNPs were associated with high myopia. The SNP rs7744813, previously shown to be associated with refractive error and myopia in two GWAS, showed an odds ratio of 0.75 (95% CI 0.63-0.90; Pemp = 0.0058) for the minor allele. The top SNP rs9342979 showed an odds ratio of 0.75 (95% CI 0.64-0.89; Pemp = 0.0045) for the minor allele. Both SNPs are located within enhancer histone marks and DNase-hypersensitive sites. Our data support the involvement of KCNQ5 gene polymorphisms in the genetic susceptibility to high myopia and further exploration of KCNQ5 as a risk factor for high myopia.

Pathway analysis of complex diseases for GWAS, extending to consider rare variants, multi-omics and interactions.

BACKGROUND: Genome-wide association studies (GWAS) is a major method for studying the genetics of complex diseases. Finding all sequence variants to explain fully the aetiology of a disease is difficult because of their small effect sizes. To better explain disease mechanisms, pathway analysis is used to consolidate the effects of multiple variants, and hence increase the power of the study. While pathway analysis has previously been performed within GWAS only, it can now be extended to examining rare variants, other "-omics" and interaction data. SCOPE OF REVIEW: 1. Factors to consider in the choice of software for GWAS pathway analysis. 2. Examples of how pathway analysis is used to analyse rare variants, other "-omics" and interaction data. MAJOR CONCLUSIONS: To choose appropriate software tools, factors for consideration include covariate compatibility, null hypothesis, one- or two-step analysis required, curation method of gene sets, size of pathways, and size of flanking regions to define gene boundaries. For rare variants, analysis performance depends on consistency between assumed and actual effect distribution of variants. Integration of other "-omics" data and interaction can better explain gene functions. GENERAL SIGNIFICANCE: Pathway analysis methods will be more readily used for integration of multiple sources of data, and enable more accurate prediction of phenotypes.

Identification of myopia-associated WNT7B polymorphisms provides insights into the mechanism underlying the development of myopia.

Myopia can cause severe visual impairment. Here, we report a two-stage genome-wide association study for three myopia-related traits in 9,804 Japanese individuals, which was extended with trans-ethnic replication in 2,674 Chinese and 2,690 Caucasian individuals. We identify WNT7B as a novel susceptibility gene for axial length (rs10453441, Pmeta=3.9 × 10(-13)) and corneal curvature (Pmeta=2.9 × 10(-40)) and confirm the previously reported association between GJD2 and myopia. WNT7B significantly associates with extreme myopia in a case-control study with 1,478 Asian patients and 4,689 controls (odds ratio (OR)meta=1.13, Pmeta=0.011). We also find in a mouse model of myopia downregulation of WNT7B expression in the cornea and upregulation in the retina, suggesting its possible role in the development of myopia.

Genome-wide association study for refractive astigmatism reveals genetic co-determination with spherical equivalent refractive error: the CREAM consortium.

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

Prevalence of myopia-related retinal changes among 12-18 year old Hong Kong Chinese high myopes.

PURPOSE: To determine the prevalence and risk factors of myopia-related retinal changes in Hong Kong Chinese adolescents with high myopia. METHODS: A cross-sectional study on Hong Kong Chinese teenage subjects with high myopia was conducted between January 2005 and June 2009. Subjects were recruited via newspaper advertisements, invitation letters to schools, leaflets and posters. Data collected included history related to myopia progression and retinal characteristics. RESULTS: In total, 120 subjects (61 boys and 59 girls) were recruited. The mean age was 14.8 ± 1.6 years (range: 12-18 years). The mean SER of the eyes was -8.41 ± 1.60 D. Ninety four of the 120 adolescents were found to have a retinal change of which 0.8% were sight-threatening, 2.5% were posterior pole changes, and 61.7% were peripheral retinal changes. The five most frequent retinal changes found were optic nerve crescents (52.5%), white-without-pressure (51.7%), lattice degeneration (5.8%), microcystoid degeneration (5%) and pigmentary degeneration (4.2%). After adjusting for myopia over -8 D, age, gender, duration of myopia, family retinal history and intraocular pressure (IOP), binary logistic regressions showed that an axial length longer than 26.5 mm was a significant risk factor for peripheral retinal changes, optic nerve crescents and white-without-pressure. CONCLUSIONS: Peripheral retinal degenerative changes and optic nerve crescent were found in a significant proportion of high myopic teenage subjects. There is increased risk of retinal changes in eyes with an axial length >26.5 mm in 12-18 year-olds.

Genetic susceptibility to refractive error: association of vasoactive intestinal peptide receptor 2 (VIPR2) with high myopia in Chinese.

Myopia is the most common ocular disease worldwide. We investigated the association of high myopia with the common single nucleotide polymorphisms (SNPs) of five candidate genes - early growth response 1 (EGR1), v-fos FBJ murine osteosarcoma viral oncogene homolog (FOS), jun oncogene (JUN), vasoactive intestinal peptide (VIP), and vasoactive intestinal peptide receptor 2 (VIPR2). We recruited 1200 unrelated Chinese subjects with 600 cases (spherical equivalent ≤-8.00 diopters) and 600 controls (spherical equivalent within ±1.00 diopter). A discovery sample set was formed from 300 cases and 300 controls, and a replication sample set from the remaining samples. Tag SNPs were genotyped for the discovery sample set, and the most significant haplotypes and their constituent SNPs were followed up with the replication sample set. The allele and haplotype frequencies in cases and controls were compared by logistic regression adjusted for sex and age to give P a values, and multiple comparisons were corrected by permutation test to give P aemp values. Odd ratios (OR) were calculated accordingly. In the discovery phase, EGR1, JUN and VIP did not show any significant association while FOS and VIPR2 demonstrated significant haplotype association with high myopia. In the replication phase, the haplotype association for VIPR2 was successfully replicated, but not FOS. In analysis combining both sample sets, the most significant association signals of VIPR2 were the single marker rs2071625 (P a = 0.0008, P aemp = 0.0046 and OR = 0.75) and the 4-SNP haplotype window rs2071623-rs2071625-rs2730220-rs885863 (omnibus test, P a = 9.10e-10 and P aemp = 0.0001) with one protective haplotype (GGGG: P aemp = 0.0002 and OR = 0.52) and one high-risk haplotype (GAGA: P aemp = 0.0027 and OR = 4.68). This 4-SNP haplotype window was the most significant in all sample sets examined. This is the first study to suggest a role of VIPR2 in the genetic susceptibility to high myopia. EGR1, JUN, FOS and VIP are unlikely to be important in predisposing humans to high myopia.

A novel missense mutation in the NYX gene associated with high myopia.

PURPOSE: Myopia is a complex eye disorder. The X-linked form of complete congenital stationary night blindness (CSNB1A) is usually associated with moderate to high myopia, and is caused by mutations in the NYX gene. We explored if NYX mutations could be associated with high myopia, but not CSNB1A. METHODS: The coding regions of the NYX gene were sequenced for 204 Chinese males with high myopia (-8.00 dioptres or worse for both eyes). The frequencies of any sequence variations identified were determined in 200 Chinese males without myopia. Electro-oculography, electroretinography and standard cone function tests were performed on a male high myope carrying a mutation. RESULTS: A missense mutation (c.529_530GC>AT or p.Ala177Met) was identified in one male subject with high myopia, but not in 200 male emmetropes. Neither was this variant found in any of the 529 male and 567 female subjects of various ethnic backgrounds whose genome sequences are documented in the 1000 Genomes Project database. The mutation was predicted to affect the protein function. From ocular electrophysiological tests, the proband was found to have normal rod function, but mildly abnormal cone function and inner retina function. He did not seem to suffer from CSNB1A. CONCLUSIONS: One novel missense NYX mutation was identified in an adult male presented with high myopia, but without the major electrophysiological features normally associated with CSNB1A. NYX gene mutations may be considered as one of the rare genetic risk factors for high myopia without key features of CSNB1A.

Corneal changes following near work in myopic anisometropia.

PURPOSE: To examine the symmetry of corneal changes following near work in the fellow eyes of non-amblyopic myopic anisometropes. METHODS: Thirty-four non-amblyopic, myopic anisometropes (minimum 1 D spherical equivalent anisometropia) had corneal topography measured before and after a controlled near work task. Subjects were positioned in a headrest to minimise head movements and read continuous text on a computer monitor for 10 min at an angle of 25 degrees downward gaze and an accommodation demand of 2.5 D. Measures of the morphology of the palpebral aperture during primary and downward gaze were also obtained. RESULTS: The more and less myopic eyes exhibited a high degree of interocular symmetry for measures of palpebral aperture morphology during both primary and downward gaze. Following the near work task, fellow eyes also displayed a symmetrical change in superior corneal topography (hyperopic defocus) which correlated with the position of the upper eyelid during downward gaze. Greater changes in the spherical corneal power vector (M) following reading were associated with a narrower palpebral aperture during downward gaze (p = 0.07 for more myopic and p = 0.03 for less myopic eyes). A significantly greater change in J0 (an increase in against the rule astigmatism) was observed in the more myopic eyes (-0.04 ± 0.04 D) compared to the less myopic eyes (-0.02 ± 0.06 D) over a 6 mm corneal diameter (p = 0.01). CONCLUSIONS: Changes in corneal topography following near work are highly symmetrical between the fellow eyes of myopic anisometropes due to the interocular symmetry of the palpebral aperture. However, the more myopic eye exhibits changes in corneal astigmatism of greater magnitude compared to the less myopic eye.

Investigating the relationship between UMODL1 gene polymorphisms and high myopia: a case-control study in Chinese.

BACKGROUND: The UMODL1 gene was found to be associated with high myopia in Japanese. This study aimed to investigate this gene for association with high myopia in Chinese. METHODS: Two groups of unrelated Han Chinese from Hong Kong were recruited using the same criteria: Sample Set 1 comprising 356 controls (spherical equivalent, SE, within ±1 diopter or D) and 356 cases (SE ≤ -8D), and Sample Set 2 comprising 394 controls and 526 cases. Fifty-nine tag single nucleotide polymorphisms (SNPs) were selected and genotyped for Sample Set 1. Four SNPs were followed up with Sample Set 2. Both single-marker and haplotype analyses were performed with cases defined by different SE thresholds. Secondary phenotypes were also analyzed for association with genotypes. RESULTS: Data filtering left 57 SNPs for analysis. Single-marker analysis did not reveal any significant differences between cases and controls in the initial study. However, haplotype GCT for markers rs220168-rs220170-rs11911271 showed marginal significance (empirical P = 0.076; SE ≤ -12D for cases), but could not be replicated in the follow-up study. In contrast, non-synonymous SNP rs3819142 was associated with high myopia (SE ≤ -10D) in the follow-up study, but could not be confirmed using Sample Set 1. The SNP rs2839471, positive in the original Japanese study, gave negative results in all our analyses. Exploratory analysis of secondary phenotypes indicated that allele C of rs220120 was associated with anterior chamber depth (adjusted P = 0.0460). CONCLUSIONS: Common UMODL1 polymorphisms were unlikely to be important in the genetic susceptibility to high myopia in Han Chinese.

Association of high myopia with crystallin beta A4 (CRYBA4) gene polymorphisms in the linkage-identified MYP6 locus.

BACKGROUND: Myopia is the most common ocular disorder worldwide and imposes tremendous burden on the society. It is a complex disease. The MYP6 locus at 22 q12 is of particular interest because many studies have detected linkage signals at this interval. The MYP6 locus is likely to contain susceptibility gene(s) for myopia, but none has yet been identified. METHODOLOGY/PRINCIPAL FINDINGS: Two independent subject groups of southern Chinese in Hong Kong participated in the study an initial study using a discovery sample set of 342 cases and 342 controls, and a follow-up study using a replication sample set of 316 cases and 313 controls. Cases with high myopia were defined by spherical equivalent ≤ -8 dioptres and emmetropic controls by spherical equivalent within ±1.00 dioptre for both eyes. Manual candidate gene selection from the MYP6 locus was supported by objective in silico prioritization. DNA samples of discovery sample set were genotyped for 178 tagging single nucleotide polymorphisms (SNPs) from 26 genes. For replication, 25 SNPs (tagging or located at predicted transcription factor or microRNA binding sites) from 4 genes were subsequently examined using the replication sample set. Fisher P value was calculated for all SNPs and overall association results were summarized by meta-analysis. Based on initial and replication studies, rs2009066 located in the crystallin beta A4 (CRYBA4) gene was identified to be the most significantly associated with high myopia (initial study: P = 0.02; replication study: P = 1.88e-4; meta-analysis: P = 1.54e-5) among all the SNPs tested. The association result survived correction for multiple comparisons. Under the allelic genetic model for the combined sample set, the odds ratio of the minor allele G was 1.41 (95% confidence intervals, 1.21-1.64). CONCLUSIONS/SIGNIFICANCE: A novel susceptibility gene (CRYBA4) was discovered for high myopia. Our study also signified the potential importance of appropriate gene prioritization in candidate selection.

Association of IGF1 gene haplotypes with high myopia in Chinese adults.

OBJECTIVE: To investigate the association of high myopia with common single-nucleotide polymorphisms (SNPs) in the IGF1, IGFBP3, and IGFBP4 genes in a Chinese population. METHODS: For our case-control study, we recruited 600 unrelated participants: 300 case participants with high myopia (-8.00 diopters or less) and 300 emmetropic controls (within ±1.00 diopter). Twenty-one tag SNPs were selected from these candidate genes and were genotyped. Genotype data were analyzed by logistic regression. Multiple comparisons were corrected by running 15 000 permutations of case-control status. RESULTS: Although we did not find any significant association for IGF1 SNPs using single-marker analysis, we identified many windows with haplotype frequencies significantly different between case participants and control participants using a variable-sized sliding-window strategy. In particular, the most significant association was shown by a 3-SNP window consisting of rs12423791, rs7956547, and rs5742632 (omnibus test: asymptotic P(asym) = 3.70 × 10(-9) and empirical P(emp) = 6.67 × 10(-5)). There were 3 high-risk haplotypes: CAC (P(asym) = 2.35 × 10(-6); odds ratio [OR], 5.06), GAT (P(asym) = 3.56 × 10(-4); OR, 3.18), and GGC (P(asym) = 2.25 × 10(-3); OR, 25.10). There was 1 protective haplotype: GAC (P(asym) = 8.43 × 10(-4); OR, 0.63). On the other hand, our single-marker and haplotype analyses did not show any significant association for IGFBP3 and IGFBP4. CONCLUSIONS: IGF1 haplotypes are associated with genetic susceptibility to high myopia in Chinese adults, whereas IGFBP3 and IGFBP4 are unlikely to be important in the genetic predisposition to high myopia. CLINICAL RELEVANCE: IGF1 is associated with high myopia, and identifying the causal variants is important for understanding the underlying mechanisms.

UPDG: utilities package for data analysis of pooled DNA GWAS.

BACKGROUND: Despite being a well-established strategy for cost reduction in disease gene mapping, pooled DNA association study is much less popular than the individual DNA approach. This situation is especially true for pooled DNA genomewide association study (GWAS), for which very few computer resources have been developed for its data analysis. This motivates the development of UPDG (Utilities package for data analysis of Pooled DNA GWAS). RESULTS: UPDG represents a generalized framework for data analysis of pooled DNA GWAS with the integration of Unix/Linux shell operations, Perl programs and R scripts. With the input of raw intensity data from GWAS, UPDG performs the following tasks in a stepwise manner: raw data manipulation, correction for allelic preferential amplification, normalization, nested analysis of variance for genetic association testing, and summarization of analysis results. Detailed instructions, procedures and commands are provided in the comprehensive user manual describing the whole process from preliminary preparation of software installation to final outcome acquisition. An example dataset (input files and sample output files) is also included in the package so that users can easily familiarize themselves with the data file formats, working procedures and expected output. Therefore, UPDG is especially useful for users with some computer knowledge, but without a sophisticated programming background. CONCLUSIONS: UPDG provides a free, simple and platform-independent one-stop service to scientists working on pooled DNA GWAS data analysis, but with less advanced programming knowledge. It is our vision and mission to reduce the hindrance for performing data analysis of pooled DNA GWAS through our contribution of UPDG. More importantly, we hope to promote the popularity of pooled DNA GWAS, which is a very useful research strategy.

Interocular symmetry in myopic anisometropia.

PURPOSE: To investigate the interocular symmetry of optical, biometric, and biomechanical characteristics between the fellow eyes of myopic anisometropes. METHODS: Thirty-four young, healthy myopic anisometropic adults (≥ 1 D spherical equivalent difference between eyes) without amblyopia or strabismus were recruited. A range of biometric and optical parameters were measured in both eyes of each subject including axial length, ocular aberrations, intraocular pressure, corneal topography, and biomechanics. Ocular sighting dominance was also measured. RESULTS: Mean absolute spherical equivalent anisometropia was 1.70 ± 0.74 D, and there was a strong correlation between the degree of anisometropia and the interocular difference in axial length (r = 0.81, p < 0.001). The more and less myopic eyes displayed a high degree of interocular symmetry for the majority of biometric, biomechanical, and optical parameters measured. When the level of anisometropia exceeded 1.75 D, the more myopic eye was more likely to be the dominant sighting eye than for lower levels of anisometropia (p = 0.002). Subjects with greater levels of anisometropia (>1.75 D) also showed high levels of correlation between the dominant and non-dominant eyes in their biometric, biomechanical, and optical characteristics. CONCLUSIONS: Although significantly different in axial length, anisometropic eyes display a high degree of interocular symmetry for a range of anterior eye biometrics and optical parameters. For higher levels of anisometropia, the more myopic eye tends to be the dominant sighting eye.

Genotyping performance assessment of whole genome amplified DNA with respect to multiplexing level of assay and its period of storage.

Whole genome amplification can faithfully amplify genomic DNA (gDNA) with minimal bias and substantial genome coverage. Whole genome amplified DNA (wgaDNA) has been tested to be workable for high-throughput genotyping arrays. However, issues about whether wgaDNA would decrease genotyping performance at increasing multiplexing levels and whether the storage period of wgaDNA would reduce genotyping performance have not been examined. Using the Sequenom MassARRAY iPLEX Gold assays, we investigated 174 single nucleotide polymorphisms for 3 groups of matched samples: group 1 of 20 gDNA samples, group 2 of 20 freshly prepared wgaDNA samples, and group 3 of 20 stored wgaDNA samples that had been kept frozen at -70°C for 18 months. MassARRAY is a medium-throughput genotyping platform with reaction chemistry different from those of high-throughput genotyping arrays. The results showed that genotyping performance (efficiency and accuracy) of freshly prepared wgaDNA was similar to that of gDNA at various multiplexing levels (17-plex, 21-plex, 28-plex and 36-plex) of the MassARRAY assays. However, compared with gDNA or freshly prepared wgaDNA, stored wgaDNA was found to give diminished genotyping performance (efficiency and accuracy) due to potentially inferior quality. Consequently, no matter whether gDNA or wgaDNA was used, better genotyping efficiency would tend to have better genotyping accuracy.

Evaluation of proteoglycan gene polymorphisms as risk factors in the genetic susceptibility to high myopia.

PURPOSE. To investigate the relationship between high myopia and single nucleotide polymorphisms (SNPs) in six proteoglycan genes: aggrecan (ACAN), fibromodulin (FMOD), decorin (DCN), lumican (LUM), keratocan (KERA), and epiphycan (EPYC). These genes were selected for study because they are involved in induced myopia in animals and/or are within the human MYP3 locus identified by linkage analysis of families with high myopia. METHODS. Two groups of Chinese subjects were studied: group 1 (300 cases and 300 controls) and group 2 (356 cases and 354 controls). Cases were high myopes with spherical equivalent (SE) ≤ -8.00 D, and controls had SE between +1.0 and -1.0 D. From these candidate genes, 60 tagging SNPs were selected. First, 12 DNA pools were each constructed from 50 samples of the same phenotype from group 1 subjects and were tested for association with the SNPs. Second, putatively positive SNPs were confirmed by individual genotyping of group 1 subjects. Finally, positive results were replicated in group 2 subjects. RESULTS. Of the 58 SNPs successfully screened by DNA pooling, 8 ACAN SNPs passed the threshold of P ≤ 0.10 (nested ANOVA) and were then genotyped in the individual samples. Haplotypes rs3784757 and rs1516794 showed significant association with high myopia. However, the positive result could not be replicated in the second subject group. CONCLUSIONS. These six proteoglycan genes were not associated with high myopia in these Chinese subjects and hence are unlikely to be important in the genetic predisposition to high myopia.

Incidence of diabetic retinopathy in a Hong Kong Chinese population.

BACKGROUND: The aim was to examine the progression and regression of diabetic retinopathy within a four-year period in a Chinese population with type 2 diabetes mellitus in a community optometry clinic in Hong Kong. METHODS: During the period May 2005 to November 2009, 5,160 patients with type 2 diabetes mellitus who had attended at least two diabetic retinopathy screening sessions at a community optometry clinic were included as subjects in this study. All had retinal photographs taken of both eyes, which were of sufficiently good quality for grading. For the purpose of this study, diabetic retinopathy grading was based on the results of the worst eye. The main outcomes were the within four-year incidence of diabetic retinopathy and the incidence of progression and regression of diabetic retinopathy. RESULTS: Of the 5,160 subjects in this study, 3,647 had no diabetic retinopathy, while 1,513 had diabetic retinopathy at the baseline visit. Of those 3,647 subjects with no diabetic retinopathy, the within four-year cumulative incidence of any diabetic retinopathy, mild or moderate non-proliferative diabetic retinopathy and sight-threatening diabetic retinopathy was 15.16 per cent, 14.45 per cent, 0.69 per cent and 0.03 per cent, respectively. Of those 1,513 subjects with diabetic retinopathy at baseline, the within four-year progression incidence of diabetic retinopathy was 6.61 per cent and the regression incidence of diabetic retinopathy was 45.54 per cent. CONCLUSION: The high regression incidence of diabetic retinopathy suggests that it might not be necessary for all patients with diabetes to be screened annually. Other methods to determine the screening frequency for an individual patient should be explored.

PAX6 haplotypes are associated with high myopia in Han chinese.

BACKGROUND: The paired box 6 (PAX6) gene is considered as a master gene for eye development. Linkage of myopia to the PAX6 region on chromosome 11p13 was shown in several studies, but the results for association between myopia and PAX6 were inconsistent so far. METHODOLOGY/PRINCIPAL FINDINGS: We genotyped 16 single nucleotide polymorphisms (SNPs) in the PAX6 gene and its regulatory regions in an initial study for 300 high myopia cases and 300 controls (Group 1), and successfully replicated the positive results with another independent group of 299 high myopia cases and 299 controls (Group 2). Five SNPs were genotyped in the replication study. The spherical equivalent of subjects with high myopia was ≤-8.0 dioptres. The PLINK package was used for genetic data analysis. No association was found between each of the SNPs and high myopia. However, exhaustive sliding-window haplotype analysis highlighted an important role for rs12421026 because haplotypes containing this SNP were found to be associated with high myopia. The most significant results were given by the 4-SNP haplotype window consisting of rs2071754, rs3026393, rs1506 and rs12421026 (P = 3.54×10(-10), 4.06×10(-11) and 1.56×10(-18) for Group 1, Group 2 and Combined Group, respectively) and the 3-SNP haplotype window composed of rs3026393, rs1506 and rs12421026 (P = 5.48×10(-10), 7.93×10(-12) and 6.28×10(-23) for the three respective groups). The results remained significant after correction for multiple comparisons by permutations. The associated haplotyes found in a previous study were also successfully replicated in this study. CONCLUSIONS/SIGNIFICANCE: PAX6 haplotypes are associated with susceptibility to the development of high myopia in Chinese. The PAX6 locus plays a role in high myopia.

A DNA pooling-based case-control study of myopia candidate genes COL11A1, COL18A1, FBN1, and PLOD1 in a Chinese population.

PURPOSE: We examined the relationship between high myopia and common polymorphisms in four candidate genes: collagen, type XI, alpha 1 (COL11A1); collagen, type XVIII, alpha 1 (COL18A1); fibrillin 1 (FBN1); and procollagen-lysine 1,2-oxoglutarate 5-dioxygenase 1 (PLOD1). These genes were selected because rare pathogenic mutations in these genes cause disease syndromes that have myopia, usually high myopia, as one of the common presenting features. METHODS: This study recruited 600 unrelated Han Chinese subjects including 300 cases with high myopia (spherical equivalent or SE≤-8.00 diopters) and 300 controls (SE within ±1.00 diopter). A total of 66 tag single nucleotide polymorphisms (SNPs) were selected for study from these four candidate genes. The study adopted a DNA pooling strategy with an initial screen of DNA pools to identify putatively positive SNPs and then confirmed the "positive" SNPs by genotyping individual samples forming the original DNA pools. DNA pools were each constructed by mixing equal amounts of DNA from 50 individuals with the same phenotype status. Six case pools were prepared from 300 cases and six control pools from 300 controls. Allele frequencies of DNA pools were estimated by analyzing the primer-extended products with denaturing high performance liquid chromatography and compared between case pools and control pools with nested ANOVA. RESULTS: In the first stage, 60 SNPs from the 4 candidate genes were successfully screened using the DNA pooling approach. Of these, 6 SNPs showed a statistical significant difference in estimated allele frequencies between case pools and controls at p<0.10. In the second stage, these "positive" SNPs were followed up by individual genotyping, but failed to be confirmed via standard single-marker and haplotype analyses. CONCLUSIONS: Common polymorphisms in these four candidate genes (COL11A1, COL18A1, FBN1 and PLOD1) were unlikely to play important roles in the genetic susceptibility to high myopia.