Genes in the high-density lipoprotein metabolic pathway in age-related macular degeneration and polypoidal choroidal vasculopathy.

PURPOSE: To investigate the associations of genetic variants in the high-density lipoprotein (HDL) metabolism pathway with neovascular age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV). DESIGN: Cross-sectional, case-control association study. PARTICIPANTS: A Chinese case-control group of 200 neovascular AMD patients, 233 PCV patients, and 275 control subjects. METHODS: Eight single nucleotide polymorphisms (SNPs) from 6 genes of the HDL metabolism pathway and 2 known AMD-associated SNPs, rs800292 (from complement factor H [CFH]) and rs11200638 (from HtrA serine peptidase 1 [HTRA1]), were genotyped in all study subjects using the TaqMan genotyping technology (Applied Biosystems, Foster City, CA). MAIN OUTCOME MEASURES: Allele and genotypic frequencies of selected SNPs. RESULTS: The SNP rs3764261 in the cholesteryl ester transfer protein (CETP) gene was associated significantly with neovascular AMD (P = 1.82×10(-4); odds ratio [OR], 1.89) and PCV (P = 4.04×10(-4); OR, 1.80). The associations remained significant after adjusting for the CFH SNP rs800292 and the HTRA1 SNP rs11200638. A significant interaction between the CETP SNP rs3764261 and the CFH SNP rs800292 existed in both neovascular AMD and PCV, the rs800292 G allele conferring a significantly increased risk of the diseases only in individuals carrying the risk allele T of rs3764261. A borderline association was detected between the ATP-binding cassette, subfamily G, member 1 (ABCG1) gene SNP rs57137919 and PCV (P = 0.03). CONCLUSIONS: Our results showed that CETP is a susceptibility gene for neovascular AMD and PCV and that ABCG1 a putative gene for PCV. CETP exerts a modifying effect on CFH in the genetic risk. Our data suggest a link of the HDL metabolism pathway with neovascular AMD and PCV.

Associations of the C2-CFB-RDBP-SKIV2L locus with age-related macular degeneration and polypoidal choroidal vasculopathy.

PURPOSE: To investigate the associations of the C2-CFB-RDBP-SKIV2L region with neovascular age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV). DESIGN: Cross-sectional, case-control association study. PARTICIPANTS: A Chinese case-control group of 200 neovascular AMD patients, 233 PCV patients, and 275 control subjects. METHODS: An association analysis was performed of the C2-CFB-RDBP-SKIV2L locus with both neovascular AMD and PCV in a Chinese population using 19 haplotype-tagging single nucleotide polymorphisms (SNPs) and 6 previously reported SNPs across the C2-CFB-RDBP-SKIV2L region. All SNPs were genotyped using the TaqMan genotyping technology (TaqMan; Applied Biosystems [ABI], Foster City, CA). MAIN OUTCOME MEASURES: Allele and haplotype frequencies of the SNPs in the C2-CFB-RDBP-SKIV2L region. RESULTS: The SKIV2L SNPs rs429608 and rs453821 were significantly associated with neovascular AMD (P = 7.39 × 10(-5); odds ratio [OR], 0.22; 95% confidence interval [CI], 0.10-0.50; and P = 0.001; OR, 0.38; 95% CI, 0.21-0.70, respectively), whereas borderline associations were detected for C2 rs547154 (P = 0.002) and RDBP rs760070 (P = 0.003). Conditional haplotype analysis revealed that SKIV2L rs429608 could account fully for the global haplotype association identified in this region. The association of SKIV2L rs429608 with neovascular AMD remained significant after adjusting for CFH rs800292 and HTRA1 rs11200638. No individual SNP or haplotype was associated significantly with PCV. CONCLUSIONS: In this concurrent investigation of the associations of the entire C2-CFB-RDBP-SKIV2L region with neovascular AMD and PCV, the results suggested that SKIV2L is a likely causal gene for neovascular AMD, conferring a significant protective effect independent of CFH and HTRA1. These data do not support a significant role of this region in PCV, suggesting different molecular mechanisms between neovascular AMD and PCV.

SNP rs1533428 at 2p16.3 as a marker for late-onset primary open-angle glaucoma.

PURPOSE: To investigate the associations between gene variants in cholesterol 24S-hydroxylase (CYP46A1), LIM homeobox transcription factor 1-beta (LMX1B), plexin domain containing 2 (PLXDC2), toll-like receptor 4 (TLR4), transmembrane and tetratricopeptide repeat containing 2 (TMTC2), zona pellucida glycoprotein 4 (ZP4), chromosome 2p16.3, and primary open-angle glaucoma (POAG). METHODS: We studied 462 POAG patients and 577 controls from three cohorts (Hong Kong, Shantou, and Beijing, China). Twelve single-nucleotide polymorphisms (SNPs) were genotyped in the Hong Kong cohort using TaqMan genotyping assay. Significant associations were validated in the Shantou and Beijing cohorts. RESULTS: Association of POAG with TLR4 rs7037117, in a recessive model, was identified in the Hong Kong and Shantou cohorts (both southern Chinese, p(rec)=0.0019) but not the Beijing cohort (northern Chinese). rs1533428 at chromosome 2p16.3 showed a consistent trend of age-specific association in all three cohorts. Genotypes TT + CT conferred a 2.16 fold of significantly increased risk to late-onset POAG (p(dom)=0.00025), but no significant risk to POAG of younger ages of onset in the combined cohort. A joint effect was found between rs7037117 and rs1533428, with carriers of both higher-risk genotypes having a 4.53 fold of increased disease risk (p=0.00028). CONCLUSIONS: Our study reveals discrepant association patterns of 12 candidate SNPs in 7 genes/loci with POAG in Chinese, provides positive replications for POAG markers rs1533428 at 2p16.3 and TLR4 rs7037117, and suggests that rs1533428 is a putative risk variant for late-onset POAG. The identification of an age-specific association between rs1533428 and late-onset POAG highlights a new genotype-phenotype association in POAG. Further studies are warranted to confirm the age-specific association.

Complement factor H and interleukin gene polymorphisms in patients with non-infectious intermediate and posterior uveitis.

OBJECTIVE: To investigate the associations of complement factor H (CFH), KIAA1109, and interleukin-27 (IL-27) gene polymorphisms in patients with non-infectious intermediate and posterior uveitis. METHODS: The study cohort consisted of a total of 95 Chinese non-infectious uveitis patients, including 38 patients with intermediate uveitis (IU), 38 patients with Vogt-Koyanagi-Harada disease (VKH), and 19 patients with Behçet's disease and 308 healthy controls. The genotypes of CFH-rs800292, KIAA1109-rs4505848, and IL27-rs4788084 were determined using TaqMan single nucleotide polymorphism genotyping assays. RESULTS: The frequency of carriers of G allele for CFH-rs800292 was significantly higher in patients with non-infectious intermediate and posterior uveitis than in controls (GG/AG versus AA; p=0.02). No significant association was found between uveitis and both KIAA1109-rs4505848 and IL27-rs4788084. In stratified analysis by gender, the frequency of carriers with G allele for KIAA1109-rs4505848 was significantly higher in male uveitis patients than in male controls (GG/AG versus AA; p=0.034). There was no significant difference in allelic and genotypic frequencies for CFH-rs800292 and IL27-rs4788084 in either male or female groups. In addition, higher frequency of KIAA1109-rs4505848 G allele was found in Behçet's disease patients compared with controls and IU patients (p=0.01 and p=0.003, respectively). CONCLUSIONS: Our results demonstrated that CFH-rs800292 and KIAA1109-rs4505848 are associated with non-infectious intermediate and posterior uveitis. Moreover, gender susceptibility for uveitis might be involved in the KIAA1109 gene and the KIAA1109-rs4505848 polymorphism might be associated with the development of Behçet's disease.

Evaluation of NTF4 as a causative gene for primary open-angle glaucoma.

PURPOSE: The neurotrophin-4 (NTF4) gene has been recently implicated in primary open-angle glaucoma (POAG). In this study, we investigated the implication of NTF4 in POAG among three Chinese cohorts. METHODS: The coding regions and exon-intron boundaries of NTF4 was sequenced in 950 unrelated Chinese subjects, including a Hong Kong cohort of 390 patients and 230 controls, a Shantou cohort of 130 patients, and a Beijing cohort of 200 patients. Constructs carrying the detected variants were generated using site-directed mutagenesis and transfected into HeLa cells, followed by solubility and migration analyses. RESULTS: Three variants were identified. p.Pro151Pro was detected in three POAG patients and one control subject. Two novel missense variants, p.Gly157Ala and p.Ala182Val, were identified each in one POAG patient from the Hong Kong cohort, but not in controls. Functional assays showed that the p.Gly157Ala mutant protein was less soluble in Triton X-100, and that migration of HeLa cells transfected with either mutant construct was less than cells transfected with the wildtype. CONCLUSIONS: The NTF4 variants p.Gly157Ala and p.Ala182Val have been shown to be functional mutations, occurring in 2 of a total of 720 Chinese POAG patients. NTF4 is functionally related to POAG pathogenesis but its mutation frequency is low. Therefore, NTF4 does not have a major contribution in the molecular genetics of POAG.

CFH 184G as a genetic risk marker for anterior uveitis in Chinese females.

OBJECTIVE: To investigate the association of three single nucleotide polymorphisms (SNPs) in the complement factor H (CFH), KIAA1109, and interleukin-27 (IL-27) genes in patients with anterior uveitis (AU). METHODS: A case-control study was performed in 98 Chinese AU patients and 308 healthy controls. Three SNPs including CFH-rs800292, KIAA1109-rs4505848, and IL27-rs4788084 were detected using TaqMan SNP Genotyping Assays. Analyses were also stratified according to gender, clinical features and human leukocyte antigen (HLA)-B27 status of the patients. RESULTS: No significant association was found between all three SNPs and AU. However, when stratified by gender, there were significant increases in the frequency of the CFH-rs800292 184G allele and GG homozygosity in female patients compared with control subjects (p=0.003 and p=0.009, respectively). Similar association was not detected in males. No significant association was found between AU and KIAA1109-rs4505848 or IL27-rs4788084 even stratified by gender. There was no significant difference in genotypes of AU patients stratified by various clinical features. Subgroup analyses showed that all three SNPs (rs800292, rs4505848, and rs4788084) were not associated with AU in HLA-B27-positive patients, neither in HLA-B27-negative patients. CONCLUSIONS: Our results showed an association between AU and CFH polymorphism in Chinese female patients but not in males, indicating gender-specific genetic differences in CFH. Gender should be considered in genetic studies of anterior uveitis even extending to other immunologic diseases.

Differential pattern of RP1 mutations in retinitis pigmentosa.

PURPOSE: Retinitis pigmentosa 1 (RP1) is a major gene responsible for both autosomal dominant and autosomal recessive retinitis pigmentosa (RP). We have previously identified three disease-causing mutations out of 174 RP patients. In this study, we investigated a new cohort of Chinese RP patients to further evaluate the contribution of RP1 mutations to cause RP. METHODS: A group of 55 nonsyndromic RP patients, the majority of them isolated cases or without information on family history, were screened for mutations in the entire coding sequences of RP1, using direct DNA sequencing. All detected variants were genotyped in 190 controls, while the three putative mutations were additionally genotyped in 362 controls subjects. Web-based programs, including PolyPhen, Sorting Intolerant from Tolerant (SIFT), Prediction of Pathological Mutations (PMUT), Single Amino Acid Polymorphism Disease-Association Predictor (SAP), ScanProsite, and ClustalW2, were used to predict the potential functional and structural impacts of the missense variants on RP1. RESULTS: A total of 14 sequence changes were identified. Among them, five were novel and found only in the RP patients. Two missense variants (p.K1370E and p.R1652L), which are conserved in primates, were predicted to have functional and structural impacts on the RP1 protein. The other three variants (c.787+34T>C, p.I408L and p.L2015L) were considered benign. CONCLUSIONS: If these two novel missense variants are in fact pathogenic, then RP1 mutations account for approximately 2.18% (5/229) of RP cases in our Chinese cohort; this is similar to other ethnic groups. However, a relatively higher frequency of missense mutations found in the Chinese patients may suggest an ethnic diversity in the RP1 mutation patterns.

Evaluation of SPARC as a candidate gene of juvenile-onset primary open-angle glaucoma by mutation and copy number analyses.

PURPOSE: To investigate the involvement of SPARC (secreted protein acidic and rich in cysteine) mutations and copy number variation in juvenile-onset primary open-angle glaucoma (JPOAG). METHODS: This study involved the 27 family members from the GLC1M (glaucoma 1, open angle, M)-linked Philippine pedigree with JPOAG, 46 unrelated Chinese patients with JPOAG and 95 controls. Mutation screening of the SPARC sequence, covering the promoter, 5'-untranslated region (UTR), entire coding regions, exon-intron boundaries, and part of the 3'-UTR, was performed using polymerase chain reaction and direct DNA sequencing. Copy number of the gene was analyzed by three TaqMan copy number assays. RESULTS: No putative SPARC mutation was detected in the Philippine family. In the Chinese participants, 11 sequence variants were detected. Two were novel: IVS2+8G>T and IVS2+32C>T. For the 9 known SNPs, one was synonymous (rs2304052, p.Glu22Glu) and the others were located in noncoding regions. No individual SNP was associated with JPOAG. Five of the most common SNPs, i.e., rs2116780, rs1978707, rs7719521, rs729853, and rs1053411, were contained in a LD (linkage disequilibrium) block. Haplotype-based analysis showed that no haplotype was associated with the disorder. Copy number analysis revealed that all study subjects had two copies of the gene, suggesting no correlation between the copy number of SPARC and JPOAG. CONCLUSIONS: We have excluded SPARC as the causal gene at the GLC1M locus in the Philippine pedigree and, for the first time, revealed that the coding sequences, splice sites and copy number of SPARC do not contribute to JPOAG. Further investigations are warranted to unravel the involvement of SPARC in the pathogenesis of other forms of glaucoma.

Novel and homozygous BEST1 mutations in Chinese patients with Best vitelliform macular dystrophy.

PURPOSE: The purpose of this study was to investigate the BEST1 gene mutations in Chinese patients with Best vitelliform macular dystrophy (BVMD). METHODS: Twenty-six subjects from 7 Chinese families with BVMD and 100 unrelated healthy Chinese subjects without a family history of BVMD were screened for mutations in the BEST1 gene by direct sequencing. The subjects underwent complete ophthalmologic examination and BEST1 gene screening. RESULTS: Six novel missense mutations (Thr2Asn, Leu75Phe, Ser144Asn, Arg255Trp, Pro297Thr, and Asp301Gly) and 1 previously reported mutation (Arg218Cys) were identified. Each family was found to have a unique BEST1 mutation that segregated with the disease. Two of the six novel mutations are located within the four previously reported common mutation clusters within the BEST1 gene. One family with patients having homozygous Leu75Phe mutations did not have the more severe BVMD phenotype. None of the patients with mutations was identified among the 100 healthy control subjects. CONCLUSION: A large number of unique novel missense mutations was found in Chinese patients with BVMD, suggesting considerable interethnic differences between the mutation sites in the BEST1 gene in different populations. The few truncating BEST1 mutations and the lack of a more severe phenotype in homozygous patients suggest that the missense BEST1 mutation may produce a dominant negative effect on wild-type BEST1 gene.

Tyrosinase gene (TYR) mutations in Chinese patients with oculocutaneous albinism type 1.

BACKGROUND: To identify the sequence variants of the tyrosinase (TYR) gene in Chinese families with oculocutaneous albinism. METHODS: Three families with oculocutaneous albinism type 1 and 95 unrelated healthy Chinese individuals with normal pigmentation were screened for mutations in the TYR gene by direct sequencing. Computational algorithms were used to characterize the biological significance of the mutants. RESULTS: Four previously reported mutations (R299C, R299H, W400L and frame-shift c.930insC) and one novel mutation (F214del) were identified, and probands had homozygous or compound heterozygous TYR mutant alleles. None of the mutants were identified among the 95 normal control subjects. Computational analysis predicted that the R299C mutant inactivates the tyrosinase enzyme by misfolding of protein tertiary structure and/or retention of the misfolded tyrosinase within the endoplasmic reticulum, and F214del causes dysfunction of tyrosine enzyme by affecting the copper binding sites and altering substrate orientation and electronic transfers during catalytic reactions for melanosynthesis. CONCLUSION: We have identified five different TYR mutations, including one novel mutation, which caused oculocutaneous albinism type 1 in Chinese. Further analysis of the patients will be useful to determine the effects of these mutations on the tyrosinase activities.

Association of CTLA-4 and IL-13 gene polymorphisms with Graves' disease and ophthalmopathy in Chinese children.

PURPOSE: The frequency of childhood Graves' disease (GD) in Hong Kong Chinese is among the highest in the world but childhood Graves' ophthalmopathy (GO) appears to have milder clinical severity than does the adult disease. This study was conducted to investigate CTLA-4 and IL-13 polymorphisms in Chinese pediatric patients with GD and GO. METHODS: Recruited for the study were 177 childhood patients with GD (age range, 2-16 years) and 151 unrelated control subjects (age range, 4-16 years) for genotype analysis of IL-13 single-nucleotide polymorphisms (SNPs) (-1112C/T and 2044G/A), CTLA-4 SNPs (-318C/T, 49A/G, and CT60A/G), and the repeat length of (AT)n in the 3' untranslated region (UTR) of CTLA-4. RESULTS: The patients with GD revealed higher frequencies of CTLA-4 49 GG genotype and G alleles than did the control subjects (P = 0.005 and P = 0.03, respectively). The CT60 GG genotype and G alleles were more prevalent in GD (P = 0.07 and P = 0.02, respectively). The CTLA-4 SNPs (-318C/T, 49A/G, and CT60A/G) were in the same haplotype block, and the CGG haplotype was associated with GD (P = 0.0071) but not GO. The shortest allele of (AT)n was protective against GD (P = 8.4 x 10(-6)). The IL-13 SNPs did not affect GD or GO risk. IL-13 -1112C/T was associated with IgE elevation (P = 0.044) and 2044G/A with proptosis (P = 0.02), but these associations became insignificant after Bonferroni correction (P = 0.22 and 0.10, respectively). CONCLUSIONS: Three SNPs and the AT repeat length in CTLA-4 conferred susceptibility to childhood GD, whereas IL-13 polymorphisms did not. No association was found between CTLA-4 and IL-13 with GO.

HTRA1 variants in exudative age-related macular degeneration and interactions with smoking and CFH.

PURPOSE: Mapping the genes for age-related macular degeneration (AMD) had not been successful until recent genome-wide association studies revealed Tyr402His in CFH and rs11200638 in HTRA1 as AMD-related genetic variants. This study was conducted to identify other critical factors in HTRA1 that are associated with exudative AMD. METHODS: The promoter, splice regions, and coding exons of HTRA1 were sequenced in 163 patients with exudative AMD and 183 sex- and age-matched control subjects. Also documented were the CFH genotype and smoking status. RESULTS: Four significant SNPs were found in the promoter and the first exon of HTRA1: rs11200638 (-625G>A), rs2672598 (-487T>C), rs1049331 (102C>T, Ala34Ala), and rs2293870 (108G>T, Gly36Gly) with respective P = 1.7 x 10(-14), 3.0 x 10(-10), 3.7 x 10(-12), and 3.7 x 10(-12). Among them, rs11200638 is the most significant associated SNP with a high odds ratio (OR) of 7.6 (95% CI: 3.94-14.51). One risk haplotype block across the promoter and exon 1, ACCTT, significantly predisposes to AMD (P = 6.68 x 10(-14)). In both models, significant independent additive effects were identified with smoking and rs800292 (184G>A, Val62Ile) of CFH. Smoking and rs11200638 (HTRA1) combined caused a 15.7-fold increased risk, whereas combined rs800292 and rs11200638 caused a 23.3-fold increased risk. An extremely high population attributable risk (PAR) of 78% was also found. CONCLUSIONS: A high impact of the additive effect of CFH and HTRA1 in the development of exudative AMD was shown. The HTRA1-smoking additive effect found in this study further suggests the importance of this environmental risk factor in AMD.

Evaluation of LOXL1 polymorphisms in primary open-angle glaucoma in southern and northern Chinese.

PURPOSE: The lysyl oxidase-like protein 1 (LOXL1) gene is strongly associated with exfoliation glaucoma, which is very rare in the Chinese population. The implicated LOXL1 polymorphisms have not been associated with primary open-angle glaucoma (POAG). In this study, we investigated three of the LOXL1 polymorphisms in POAG in a southern Chinese population of Hong Kong and northern Chinese from Beijing. METHODS: The Hong Kong group included 293 POAG patients and 250 controls, and the Beijing group included 169 POAG patients and 197 controls. LOXL1 single nucleotide polymorphisms (SNPs), rs1048661, rs3825942, and rs2165241, were genotyped by direct DNA sequencing. Individual association was analyzed using the chi(2) test, and haplotype-based association analysis was performed in WHAP. RESULTS: Each of the candidate SNPs was not statistically associated with POAG in either group (p>0.017, Bonferroni correction). Haplotype-based association analysis had identified a significant omnibus association (Omnibus chi(2)=18.16, p=0.00115) between these SNPs and POAG in the Hong Kong group. A minor haplotype (T-G-T) showed significant statistical association with POAG. It presented in 2.1% of cases and 0.4% of controls, conferring a 5.24 fold of increased risk to the disease (95% CI: 1.17-23.54, P(perm)=0.00108). However, this haplotype was absent in the Beijing group. CONCLUSIONS: Individual LOXL1 SNPs, rs1048661, rs3825942, and rs2165241, were not associated with POAG in the Chinese population. However, a minor haplotype T-G-T was found to be associated with the disorder in the southern Chinese. The low frequencies of the at-risk alleles at rs1048661 and rs2165241 may be one of the factors that led to the low prevalence of exfoliation syndrome in the general populations of the Chinese.

Ethnic specific association of the CAV1/CAV2 locus with primary open-angle glaucoma.

A single-nucleotide polymorphism (SNP) rs4236601 at the CAV1/CAV2 locus is associated with primary open-angle glaucoma (POAG). Rs4236601 is common in Caucasians but rare in East Asians. Here we conducted a haplotype-tagging SNP analysis followed by replication in a total of 848 POAG cases and 1574 controls drawn from 3 cities in China and 1 city in Japan. Two SNPs, rs4236601 (odds ratio [OR] = 6.25; P = 0.0086) and a tagging-SNP rs3801994 (OR = 1.32; P = 0.042), were associated with POAG in the Hong Kong Chinese cohort after age and gender adjustments. Rs4236601 was associated with POAG also in Shantou (OR = 6.09; P = 0.0037) and Beijing (OR = 3.92; P = 0.030) cohorts after age and gender adjustment, with a pooled-OR of 5.26 (P = 9.0 × 10(-6)) in Chinese; but it is non-polymorphic in the Osaka cohort. SNP rs3801994 showed a similar trend of effect in the Shantou and Beijing cohorts, with a pooled-OR of 1.23 (P = 0.022) and 1.20 (P = 0.063) in Chinese, prior to and after age and gender adjustment, respectively; but it showed a reverse effect in the Osaka cohort (OR = 0.58; P = 0.033) after the adjustments. We have thus confirmed the association of rs4236601 with POAG in different Chinese cohorts. Also, we found a common SNP rs3801994 of diverse associations with POAG between Chinese and Japanese.

The toxic and stress responses of cultured human retinal pigment epithelium (ARPE19) and human glial cells (SVG) in the presence of triamcinolone.

PURPOSE: To compare the cytotoxic effect of TA on human retinal pigment epithelium (ARPE19) and human glial (SVG) cells over a range of concentrations and durations of exposure. METHODS: TA (0.01-1 mg/mL) or vehicle (benzyl alcohol, 0.025%) was added to the ARPE19 and SVG cultures on day 0 and then subsequently for 1, 3, or 5 days. The amount of cell proliferations with or without TA treatment was performed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. All samples were read in triplicate (n = 4 in all cases). c-Fos, c-jun, caspase-3, c-myc, and p53 expression was determined after TA treatments after 0, 10, 20, 30, 40, 50, 60, and 90 minutes. All results were analyzed with ANOVA. RESULTS: TA (0.01-1 mg/mL) caused a significant reduction in ARPE19 cells that had been exposed to it for more than 1 day. Significant reductions in the number of SVG cells were observed as early as day 1 at 0.1 and 1 mg/mL TA. In general, the level of remaining SVG cells was less than that of the APRE19 cells over the 5 days. SVG cells appeared more susceptible to TA. Caspase-3 was elevated in both ARPE19 and SVG cells after TA treatment. c-Fos and c-jun expression was also increased in ARPE19 cells but not in SVG. The vehicle of TA had no effect, and there was no change in p53 or c-myc expression. CONCLUSIONS: TA was cytotoxic to both SVG and ARPE19 cells, with higher efficacy on SVG. TA caused the activation of the caspase-3 pathway more readily than the cell-protective c-fos and c-jun pathways in SVG cells, making those cells more vulnerable than the ARPE19 cells. The results suggest that TA toxicity in one cell type may not reliably indicate its toxicity in other cells. Different cells within the retina may react to TA differently, or TA may cause changes in the gene expressions differentially with different concentrations of the same stimulus.

Gender specific association of a complement component 3 polymorphism with polypoidal choroidal vasculopathy.

Neovascular age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV) are leading causes of irreversible blindness in developed countries. In this study, we investigated the associations of haplotype-tagging single nucleotide polymorphisms (SNPs) in the complement component 3 (C3) gene with both neovascular AMD and PCV, and potential epistatic effects on C3. Eight tagging SNPs in C3 were genotyped in 708 unrelated study subjects: 200 neovascular AMD patients, 233 PCV patients and 275 controls. Among the eight C3 SNPs, rs17030 was associated with PCV after adjusted for gender and SNP-gender interaction (P = 0.008, OR = 2.94; 95% CI: 1.32-6.52). Moreover, an interaction between rs17030 and gender was identified in PCV (P = 0.02). After stratification by gender, the rs17030 G allele was found to confer an increased risk for PCV in male (P = 0.010, OR = 1.56) but not in female. The haplotype AG defined by the major alleles of rs17030 and rs344555 was also associated with PCV in male (P = 0.010, OR = 0.64). In contrast to PCV, none of the eight SNPs was significantly associated with neovascular AMD. This study shows an association of C3 rs17030 with PCV in male, indicating that C3 may have an epistatic effect with gender in the pathogenesis of PCV.

Association of CFH and SERPING1 polymorphisms with anterior uveitis.

AIMS: To investigate the association of the complement factor H (CFH) and complement component 1 inhibitor (SERPING1) genes with anterior uveitis (AU). METHODS: A total of 406 subjects (98 patients with AU and 308 controls) were recruited for this study. Two CFH polymorphisms (rs3753394 and rs1065489) and two SERPING1 polymorphisms (rs1005511 and rs3824988) were genotyped using TaqMan genotyping assays. Analyses were stratified for gender and human leukocyte antigen (HLA)-B27 status. Correlations of the genotypes with multiple clinical features were also evaluated. RESULTS: No significant association was found between any of the four polymorphisms and AU after multiple testing corrections. However, stratified analyses showed that there were significant increases in the frequencies of T allele and TT homozygosity for CFH-rs1065489 in female patients compared with that of controls (pcorr=0.004 and pcorr=0.012 respectively). In addition, CFH-rs1065489 was also associated with AU in patients who were HLA-B27 positive. No significant association with AU was found for the other three single nucleotide polymorphisms (SNPs), even stratified by gender or HLA-B27 status. Genotype-phenotype analyses found that CFH-rs1065489 TT genotype was associated with higher uveitis recurrence frequency. No correlation was found between CFH-rs1065489 and other clinical features. CONCLUSIONS: This study revealed an association of CFH-rs1065489 with AU as well as uveitis recurrence frequency. The influence on AU could be gender specific and dependent on HLA-B27 status. Our results also suggested that SERPING1 does not play a significant role in the development of AU.

Association of C2 and CFB polymorphisms with anterior uveitis.

PURPOSE: Association of rs800292 (I62V) in the complement factor H (CFH) gene with anterior uveitis (AU) was identified in our previous study. We proceeded to investigate whether polymorphisms of two tightly linked genes in the complement pathway, complement component 2 (C2) and complement factor B (CFB), are associated with AU. METHODS: Five single-nucleotide polymorphisms (SNPs), rs1048709, rs537160, rs4151657, rs2072633 in CFB, and rs3020644 in C2, were examined using genotyping assays in 98 Chinese AU patients and 291 unrelated controls. Adjustments and stratifications were given for sex, clinical manifestations, and HLA-B27 status. RESULTS: There were significant increases in the frequency of A allele and AA homozygosity for CFB-rs1048709 in AU patients compared with that of controls (P value after Bonferroni correction [P(corr)] = 2.67 × 10⁻4, P(corr) = 0.001, respectively). No association was found between AU and the other four SNPs after adjustment for multiple testing. Logistic regression analysis showed none of the 5 SNPs had significant interaction with sex. Stratified analyses showed that only rs1048709 was significantly associated with AU in HLA-B27-positive patients but not in HLA-B27-negative patients. No association was found in the 5 tested SNPs with clinical manifestations. A haplotype block across CFB (AATA) was significantly predisposed to AU with increased risk of 1.97 (P(corr) = 0.0005). Additive effect of CFB-rs1048709 and CFH-rs800292 was identified with an odds ratio of 7.48. CONCLUSIONS: Our results revealed an association between AU and CFB-rs1048709. The influence on AU might differ depending on HLA-B27 status. The joint effect in CFB and CFH strengthens the concept that the complement system plays an important role in the pathogenesis of AU.

Differentiation of exudative age-related macular degeneration and polypoidal choroidal vasculopathy in the ARMS2/HTRA1 locus.

PURPOSE: differentiate the associations of exudative age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV) with the ARMS2/HTRA1 locus. METHODS: The entire ARMS2 sequence was sequenced and HTRA1 rs11200638 genotyped in 568 unrelated Chinese individuals: 156 exudative AMD patients, 164 PCV patients, and 248 controls. A meta-analysis was performed to examine the effects of rs10490924 and rs11200638 at the ARMS2/HTRA1 locus in PCV. RESULTS: In total, 31 polymorphisms in ARMS2 were identified. Significant associations with both exudative AMD and PCV were observed in 11 of them and HTRA1 rs11200638, with different genotypic distributions between exudative AMD and PCV (P < 0.001). After adjusting for rs11200638, ARMS2 rs10490924 remained significantly associated with exudative AMD (P = 0.011), but not with PCV (P = 0.077). Meta-analysis showed consistent allelic associations of rs10490924 and rs11200638 with PCV in different study populations. CONCLUSIONS: There is a strong and consistent association of the ARMS2/HTRA1 locus with both exudative AMD and PCV, suggesting the two disorders share, at least partially, similar molecular mechanisms. Different effect sizes indicate the existence of additional genetic and environmental factors affecting them to different extents.

Association of NR2E3 but not NRL mutations with retinitis pigmentosa in the Chinese population.

Purpose. Mutations in the NR2E3 and NRL genes have been implicated in both autosomal dominant and autosomal recessive retinitis pigmentosa (RP). In this study, the mutation profiles of these two genes were investigated in Chinese RP patients. Methods. In 172 RP patients and 360 normal control subjects (180 from Hong Kong and 180 from Beijing), the coding exons and the exon-intron boundaries of NR2E3 and NRL were screened by direct DNA sequencing after PCR. Association analysis was performed for common single-nucleotide polymorphisms (SNPs), whereas in silico programs were used for analysis of rare missense variants. Results. In NR2E3, 14 novel sequence changes have been identified. Two missense variants, p.G56R and p.V118M, were exclusively found in RP patients with frequencies at 1.2% (2/172) and 1.7% (3/172), respectively. All five patients were found to be heterozygous for these two mutations. Computational analysis suggested functional defects on the NR2E3 protein, indicating disease-causing roles. The p.E121K variant of NR2E3, which reportedly caused enhanced S-cone syndrome (ESCS) in Caucasians, was found concurrently in RP patients (13.4%) and control subjects from Hong Kong (10.5%) and Beijing (12.8%). In NRL, six novel sequence changes were identified, none of them associated with RP. Conclusions. In this study, NR2E3 mutations (p.G56R, p.V118M) were found to be responsible for approximately 2.9% of overall RP in Chinese patients, comparable to the contributions of RHO and RP1 mutations. The p.E121K in NR2E3 is a common SNP in the Chinese, suggesting another genetic or environmental factor is involved in its causative role in ESCS in Caucasians.