LncRNA VEAL2 regulates PRKCB2 to modulate endothelial permeability in diabetic retinopathy.

Long non-coding RNAs (lncRNAs) are emerging as key regulators of endothelial cell function. Here, we investigated the role of a novel vascular endothelial-associated lncRNA (VEAL2) in regulating endothelial permeability. Precise editing of veal2 loci in zebrafish (veal2gib005Δ8/+ ) induced cranial hemorrhage. In vitro and in vivo studies revealed that veal2 competes with diacylglycerol for interaction with protein kinase C beta-b (Prkcbb) and regulates its kinase activity. Using PRKCB2 as bait, we identified functional ortholog of veal2 in humans from HUVECs and named it as VEAL2. Overexpression and knockdown of VEAL2 affected tubulogenesis and permeability in HUVECs. VEAL2 was differentially expressed in choroid tissue in eye and blood from patients with diabetic retinopathy, a disease where PRKCB2 is known to be hyperactivated. Further, VEAL2 could rescue the effects of PRKCB2-mediated turnover of endothelial junctional proteins thus reducing hyperpermeability in hyperglycemic HUVEC model of diabetic retinopathy. Based on evidence from zebrafish and hyperglycemic HUVEC models and diabetic retinopathy patients, we report a hitherto unknown VEAL2 lncRNA-mediated regulation of PRKCB2, for modulating junctional dynamics and maintenance of endothelial permeability.

Binding of Gtf2i-ß/δ transcription factors to the ARMS2 gene leads to increased circulating HTRA1 in AMD patients and in vitro.

The disease-initiating molecular events for age-related macular degeneration (AMD), a multifactorial retinal disease affecting many millions of elderly individuals worldwide, are still unknown. Of the over 30 risk and protective loci so far associated with AMD through whole genome-wide association studies (GWAS), the Age-Related Maculopathy Susceptibility 2 (ARMS2) gene locus represents one of the most highly associated risk regions for AMD. A unique insertion/deletion (in/del) sequence located immediately upstream of the High Temperature Requirement A1 (HTRA1) gene in this region confers high risk for AMD. Using electrophoretic mobility shift assay (EMSA), we identified that two Gtf2i-ß/δ transcription factor isoforms bind to the cis-element 5'- ATTAATAACC-3' contained in this in/del sequence. The binding of these transcription factors leads to enhanced upregulation of transcription of the secretory serine protease HTRA1 in transfected cells and AMD patient-derived induced pluripotent stem cells (iPSCs). Overexpression of Htra1 in mice using a CAG-promoter demonstrated increased blood concentration of Htra1 protein, caused upregulation of vascular endothelial growth factor (VEGF), and produced a choroidal neovascularization (CNV)-like phenotype. Finally, a comparison of 478 AMD patients to 481 healthy, age-matched controls from Japan, India, Australia, and the USA showed a statistically increased level of secreted HTRA1 blood concentration in AMD patients compared with age-matched controls. Taken together, these results suggest a common mechanism across ethnicities whereby increased systemic blood circulation of secreted serine protease HTRA1 leads to subsequent degradation of Bruch's membrane and eventual CNV in AMD.

Angiopoietin receptor TEK interacts with CYP1B1 in primary congenital glaucoma.

Primary congenital glaucoma (PCG) is a severe autosomal recessive ocular disorder associated with considerable clinical and genetic heterogeneity. Recently, rare heterozygous alleles in the angiopoietin receptor-encoding gene TEK were implicated in PCG. We undertook this study to ascertain the second mutant allele in a large cohort (n = 337) of autosomal recessive PCG cases that carried heterozygous TEK mutations. Our investigations revealed 12 rare heterozygous missense mutations in TEK by targeted sequencing. Interestingly, four of these TEK mutations (p.E103D, p.I148T, p.Q214P, and p.G743A) co-occurred with three heterozygous mutations in another major PCG gene CYP1B1 (p.A115P, p.E229K, and p.R368H) in five families. The parents of these probands harbored either of the heterozygous TEK or CYP1B1 alleles and were asymptomatic, indicating a potential digenic mode of inheritance. Furthermore, we ascertained the interactions of TEK and CYP1B1 by co-transfection and pull-down assays in HEK293 cells. Ligand responsiveness of the wild-type and mutant TEK proteins was assessed in HUVECs using immunofluorescence analysis. We observed that recombinant TEK and CYP1B1 proteins interact with each other, while the disease-associated allelic combinations of TEK (p.E103D)::CYP1B1 (p.A115P), TEK (p.Q214P)::CYP1B1 (p.E229K), and TEK (p.I148T)::CYP1B1 (p.R368H) exhibit perturbed interaction. The mutations also diminished the ability of TEK to respond to ligand stimulation, indicating perturbed TEK signaling. Overall, our data suggest that interaction of TEK and CYP1B1 contributes to PCG pathogenesis and argue that TEK-CYP1B1 may perform overlapping as well as distinct functions in manifesting the disease etiology.

Mutation spectrum of the Norrie disease pseudoglioma (NDP) gene in Indian patients with FEVR.

PURPOSE: Mutations in the Norrie disease pseudoglioma (NDP; Xp11.3) gene have been involved in retinal blood vessel formation and neural differentiation and are implicated in familial exudative vitreoretinopathy (FEVR) cases. However, the role of the gene has not been explored in the Indian context. Thus, this study was designed to understand the involvement of NDP among Indian patients with FEVR. METHODS: The study cohort comprised 225 subjects, including unrelated patients with FEVR (n = 110) and ethnically matched healthy subjects (n = 115) recruited from a tertiary eye care center in India. The entire coding regions, intron-exon boundaries, along with the 5' and 3' untranslated regions of NDP were screened with resequencing following standard protocols. The spectrum of the observed variants was analyzed in conjunction with data available from other populations. RESULTS: Eight potentially pathogenic mutations (p.His4ArgfsX21, p.Asp23GlufsX9, p.Ile48ValfsX55, p.His50Asp, p.Ser57*, p.Gly113Asp, p.Arg121Gln, and p.Cys126Arg, including five novel ones), were observed in the coding region of the NDP gene in ten unrelated FEVR probands (9%). The novel changes were not observed in the control subjects and were unavailable in the dbSNP, ESP5400, NIEHS95, and ExAC databases. All probands with NDP mutations exhibited classical features of the disease as observed among patients with FEVR worldwide. CONCLUSIONS: This is perhaps the first study to demonstrate the involvement of NDP among patients with Indian FEVR that further expands its mutation spectrum. The data generated could have broad implications in genetic counseling, disease management, and early intervention for a better prognosis in FEVR.

Mutation spectrum of the FZD-4, TSPAN12 AND ZNF408 genes in Indian FEVR patients.

BACKGROUND: Mutations in candidate genes that encode for a ligand (NDP) and receptor complex (FZD4, LRP5 and TSPAN12) in the Norrin ß-catenin signaling pathway are involved in the pathogenesis of familial exudative vitreoretinopathy (FEVR, MIM # 133780). Recently, a transcription factor (ZNF408) has also been implicated in FEVR. We had earlier characterized the variations in NDP among FEVR patients from India. The present study aimed at understanding the involvement of the remaining genes (FZD4, TSPAN12 and ZNF408) in the same cohort. METHODS: The DNA of 110 unrelated FEVR patients and 115 unaffected controls were screened for variations in the entire coding and untranslated regions of these 3 genes by resequencing. Segregation of the disease-associated variants was assessed in the family members of the probands. The effect of the observed missense changes were further analyzed by SIFT and PolyPhen-2 scores. RESULTS: The screening of FZD4, TSPAN12 and ZNF408 genes identified 11 different mutations in 15/110 FEVR probands. Of the 11 identified mutations, 6 mutations were novel. The detected missense mutations were mainly located in the domains which are functionally crucial for the formation of ligand-receptor complex and as they replaced evolutionarily highly conserved amino acids with a SIFT score < 0.005, they are predicted to be pathogenic. Additionally 2 novel and 16 reported single nucleotide polymorphisms (SNP) were also detected. CONCLUSIONS: Our genetic screening revealed varying mutation frequencies in the FZD4 (8.0 %), TSPAN12 (5.4 %) and ZNF408 (2.7 %) genes among the FEVR patients, indicating their potential role in the disease pathogenesis. The observed mutations segregated with the disease phenotype and exhibited variable expressivity. The mutations in FZD4 and TSPAN12 were involved in autosomal dominant and autosomal recessive families and further validates the involvement of these gene in FEVR development.

Longitudinal Andhra Pradesh Eye Disease Study: rationale, study design and research methodology.

BACKGROUND: The rationale, objectives, study design and procedures for the longitudinal Andhra Pradesh Eye Disease Study are described. DESIGN: A longitudinal cohort study was carried out. PARTICIPANTS: Participants include surviving cohort from the rural component of Andhra Pradesh Eye Disease Study. METHODS: During 1996-2000, Andhra Pradesh Eye Disease Survey was conducted in three rural (n = 7771) and one urban (n = 2522) areas (now called Andhra Pradesh Eye Disease Study 1). In 2009-2010, a feasibility exercise (Andhra Pradesh Eye Disease Study 2) for a longitudinal study (Andhra Pradesh Eye Disease Study 3) was undertaken in the rural clusters only, as urban clusters no longer existed. In Andhra Pradesh Eye Disease Study 3, a detailed interview will be carried out to collect data on sociodemographic factors, ocular and systemic history, risk factors, visual function, knowledge of eye diseases and barriers to accessing services. All participants will also undergo a comprehensive eye examination including photography of lens, optic disc and retina, Optic Coherence Tomography of the posterior segment, anthropometry, blood pressure and frailty measures. MAIN OUTCOME MEASURES: Measures include estimates of the incidence of visual impairment and age-related eye disease (lens opacities, glaucoma and age-related macular degeneration) and the progression of eye disease (lens opacities and myopia) and associated risk factors. RESULTS: Of the 7771 respondents examined in rural areas in Andhra Pradesh Eye Disease Study 1, 5447 (70.1%) participants were traced in Andhra Pradesh Eye Disease Study 2. These participants will be re-examined. CONCLUSIONS: Andhra Pradesh Eye Disease Study 3 will provide data on the incidence and progression of visual impairment and major eye diseases and their associated risk factors in India. The study will provide further evidence to aid planning eye care services.

Exome sequencing reveals the likely involvement of SOX10 in uveal melanoma.

PURPOSE: To identify the spectrum of somatic mutations in an Asian Indian patient with uveal melanoma (UM) without metastasis using exome sequencing. CASE REPORT: A 49-year-old man from India was diagnosed as having cilio-choroidal (uveal) melanoma (UM), without metastasis, in his right eye with the help of magnetic resonance imaging. This was later confirmed by histopathological evaluation. Two individuals from India with non-neoplastic blind eyes were recruited as controls. The affected eyes from the UM patient and the two control individuals were enucleated, and uveal tissues were collected. DNA was extracted from uveal tissue, and the matched blood sample from each of the three individuals was followed by exome sequencing. Statistical and bioinformatic analyses were done to identify somatic mutations and their putative associations with UM. Thirty-one somatic mutations (25 amino acid altering) in protein-coding (exonic) regions were detected in the UM patient. Of the amino acid-altering somatic mutations, 16 mutations were predicted to be candidate mutations relevant to UM. Somatic mutations, putatively causal for UM, were identified in GNAQ, SF3B1, and SOX10. CONCLUSIONS: Somatic mutations in GNAQ and SF3B1 genes were probable drivers of UM in the Indian patient; these were also reported earlier in some White patients. In addition, a frameshift deletion of 20 base pairs has been identified in SOX10 in the UM patient. Somatic mutations in SOX10, a transcription factor, which acts upstream of microphthalmia-associated transcription factor and synergizes with microphthalmia-associated transcription factor, was identified in some melanoma cell lines. The transcription factor SOX10 was found to have an essential role in melanocyte development and pigmentation. Our finding of the frameshift deletion (p.H387fs) in exon 4 of SOX10 in UM provides an important insight and complements earlier findings of mutations in GNAQ and SF3B1 on the genomic basis of UM.

How genetic studies have advanced our understanding of age-related macular degeneration and their impact on patient care: a review.

The last 10 years have seen an unprecedented explosion in our knowledge regarding the genomic basis of age-related macular degeneration. This has come about through major advances in computing power, microfabrication of large numbers of molecular markers on chips and improved statistical algorithms for analysis. In tandem, it has become clear that age-related macular degeneration appears to be a multifactorial disease with influences from genetic and structural variants, as well as epigenetic involvement. The combination of these factors with known environmental determinants indicates the highly complex nature of this disease, but at the same time also offers insights into risk prediction and disease stratification through genotype profiling.

Changing trends in barriers to accessing eye care services in rural South India: results from the longitudinal Andhra Pradesh Eye Disease Study III (APEDS III) cohort.

PURPOSE: To assess the changing trends in barriers towards accessing eye care in a rural population cohort from Southern India. METHODS: This is a population-based longitudinal cohort of participants (the Andhra Pradesh Eye Disease study [APEDS]) from three rural regions of Telangana and Andhra Pradesh who were evaluated at baseline (APEDS I; 1996-2000), along with follow-ups at 10 years (APEDS II; 2009-10) and 15 years (APEDS III; 2012-2016). At follow-up, all participants 30 years and above were administered a structured questionnaire on barriers to uptake of eye care services. RESULTS: Of 3810 participants, 1449 had visual impairment (VI). Among them, 1302 noticed a reduction in vision over last five years and 722 sought treatment, a significant improvement from baseline (P < 0.001). Participants were more likely to seek treatment if they were educated (OR = 1.43, 95%CI: 1.07-1.89), had hypertension (OR = 1.36, 95%CI: 1.04-1.77), had VI from causes other than cataract and refractive error (OR = 2.49, 95%CI: 1.56-3.99) and were residents of Adilabad (OR = 2.21; 95%CI: 1.58-3.08) and Mahbubnagar (OR = 3.55; 95%CI: 2.48-5.08) districts. Those with moderate or worse VI were less likely to seek treatment (moderate VI: OR = 0.56; 95%CI: 0.42-0.75, severe VI: OR = 0.34; 95%CI: 0.19-0.57, blindness: OR = 0.38; 95%CI: 0.2-0.73). The most important barriers to uptake of services were, not perceiving loss of vision as a serious problem (25.9%), accepting it an aging process (21.4%) or due to economic reasons (16.0%). CONCLUSION: Personal and economic elements accounted for considerable amounts of barriers for utilization of eye care services. The uptake of services could be improved by addressing these specific barriers and risk factors for non-compliance.

Association of the del443ins54 at the ARMS2 locus in Indian and Australian cohorts with age-related macular degeneration.

PURPOSE: The ARMS2/HTRA1 genes at the 10q26 locus have been associated with risk of age-related macular degeneration (AMD), with the most significantly associated variants being A69S (rs10490924), del443ins54 (EU427539) and rs11200638. We wished to explore the association of the del443ins54 in two ethnically different populations from India and Australia. METHODS: The del443ins54 was screened in a large cohort of ~1500 subjects from these two populations by a combination of PCR-based agarose gel electrophoresis and validated by resequencing. Statistical analysis comprised the calculations of allele, genotype and haplotype frequencies along with their p values and corresponding odds ratios (OR), and 95% confidence intervals (95% CI) and measures of linkage disequilibrium (LD). RESULTS: The del443ins54 was significantly associated with AMD in both the Indian (p=1.74 × 10(-13); OR = 2.80, 95%CI, 2.12-3.70) and Australian cohorts (p = 2.78 × 10(-30); OR = 3.15, 95%CI, 2.58-3.86). These associations were similar to those previously identified for the A69S and the rs11200638 variant in these populations that also exhibited high degrees of LD (D' of 0.87-0.99). A major risk haplotype of "T-indel-A" (p = 5.7 × 10(-16); OR = 3.16, 95%CI, 2.34-4.19 and p=6.33 × 10(-30); OR = 3.15, 95%CI, 2.57-3.85) and a protective haplotype of "G-wild type-G" (p=2.35 × 10(-11); OR = 0.39, 95%CI, 0.29-0.52 and p=1.02 × 10(-30); OR = 0.31, 95%CI, 0.25-0.38) were identified in the Indian and Australian cohorts, respectively. CONCLUSIONS: These data provide an independent replication of the association of del443ins54 variant in two different ethnicities, despite differences in allele and haplotype frequencies between them. High levels of LD in both populations limit further genetic dissection of this region in AMD.

Barriers to uptake of referral eye care services among the elderly in residential care: the Hyderabad Ocular Morbidity in Elderly Study (HOMES).

BACKGROUND: To report on the barriers to uptake of eye care services after referral in the elderly in 'homes for the aged' in Hyderabad, India. METHODS: Individuals aged ≥60 years were recruited from 41 'homes for the aged' and were examined in the 'make-shift' clinics in homes. All participants who had vision impairment or needed further eye examination other than spectacles were referred to the higher centres for 'free services'. Three months after the referral, the participants were interviewed and asked about the uptake of services, and their reasons for not attending. RESULTS: In all, 731/1182 (61.8%) participants were referred of which 375 (49.9%) attended. In multiple logistic regression, participants aged ≥80 years were less likely to utilise the services (OR 0.60; 95% CI 0.39 to 0.03). Similarly, the participants living in free homes (OR 3.53; 95% CI 2.15 to 5.79) and subsidised homes (OR 2.24: 95% CI 1.55 to 3.23) and those independently mobile had higher odds for uptake of services (OR 5.74; 95% CI 3.31 to 10.51). The major reasons for not availing the referral services were 'lack of felt need' reported by 136 (45.4%) participants followed by other health issues in 100 (33.4%) participants and non-consenting family members in 49 (16.4%) participants. In all, 14 (4.7%) participants gave other reasons. CONCLUSIONS: The uptake of eye care services in the elderly in residential care remains poor despite the provision of services for free. Lack of felt need for services is the main reason for non-compliance to the referral for care. Counselling on the benefit of interventions could potentially improve referral compliance in this population.

Visual impairment and blindness due to retinitis pigmentosa in India: 15-year follow-up of the Andhra Pradesh Eye Disease Study cohort.

Purpose: To assess the incidence, visual impairment, and blindness due to retinitis pigmentosa (RP) in a rural southern Indian cohort. Methods: This is a population-based longitudinal cohort study of participants with RP from the Andhra Pradesh Eye Disease Study (APEDS) cohorts I and III, respectively. The study included participants with RP of APEDS I who were followed until APEDS III. Their demographic data along with ocular features, fundus photographs, and visual fields (Humphrey) were collected. Descriptive statistics using mean ± standard deviation with interquartile range (IQR) were calculated. The main outcome measures were RP incidence, visual impairment, and blindness as per the World Health Organization (WHO) definitions. Results: At baseline (APEDS I), 7771 participants residing in three rural areas were examined. There were nine participants with RP with a mean age at baseline of 47.33 ± 10.89 years (IQR: 39-55). There was a male preponderance (6:3), and the mean best-corrected visual acuity (BCVA) of 18 eyes from nine participants with RP was 1.2 ± 0.72 logarithm of minimum angle of resolution (logMAR; IQR: 0.7-1.6). Over a mean follow-up duration of 15 years, 5395/7771 (69.4%) were re-examined, which included seven RP participants from APEDS 1. Additionally, two new participants with RP were identified; so, the overall incidence was 370/ million in 15 years (24.7/million per year). The mean BCVA of 14 eyes of seven participants with RP who were re-examined in APEDS III was 2.17 ± 0.56 logMAR (IQR: 1.8-2.6), and five of these seven participants with RP developed incident blindness during the follow-up period. Conclusion: RP is a prevalent disease in southern India that warrants appropriate strategies to prevent this condition.

Regional variation in the incidence of pseudo-exfoliation in the Andhra Pradesh Eye Disease Study (APEDS).

BACKGROUND: To report the 15-year incidence rate of pseudo-exfoliation (PXF),  PXF glaucoma and regional variation among rural participants in the Andhra Pradesh Eye Disease Study (APEDS) III. METHODS: This population-based longitudinal study was carried out at three rural study sites. Individuals of all ages who participated at baseline with a mean 15-year follow-up visit were included. Detailed Comprehensive ophthalmic examination was performed on all participants. The main outcome measure was development of PXF during the follow-up period in participants who were phakic in one or both eyes without PXF at baseline. RESULTS: Among 5395 participants, 5108 (94.6%) met the inclusion criteria. There were 93 (1.82%; 95% confidence interval (CI), 1.47-2.22) cases of incident PXF. Their median baseline age (1st, 3rd quartiles) was 51 (44, 59) years and the male: female ratio was 1.3:1. There was no case of incident PXF in participants aged <30 years at baseline. The incidence rate per 100 person years (95% CI) among all ages and those aged ≥30 years at baseline was 1.73 (1.64-1.82) and 3.73 (3.53-3.93), respectively. PXF material was located on iris as well as anterior surface of lens and it was often bilateral. Participants living in two study sites and increasing age were associated with the incidence of PXF. The 15-year incidence of PXF glaucoma (95% CI) in participants ≥30 years of age at baseline was 0.33% (0.14-0.66). CONCLUSION: There is significant regional variation in incidence of PXF in south India which warrants further investigation.

Incidence of primary open angle glaucoma in the Andhra Pradesh Eye Disease Study (APEDS).

BACKGROUND: To report 15-year incidence rate of primary open angle glaucoma (POAG) in the Andhra Pradesh Eye Disease Study (APEDS). METHODS: A population-based longitudinal study was carried out at three rural study sites. Phakic participants aged ≥40 years who participated at baseline (APEDS I) and the mean 15-year follow-up visit (APEDS III) were included. A comprehensive ophthalmic examination was performed on all participants. Mean intraocular pressure (IOP) was average of IOPs of right and left eyes. The definition of glaucoma was based on the International Society of Geographical and Epidemiological Ophthalmology (ISGEO) classification. The main outcome measure was incidence of POAG during the follow-up period in participants without glaucoma or suspicion of glaucoma at baseline. RESULTS: Data from the available and eligible participants from the original cohort (1241/2790; 44.4%) were analysed. The mean age (standard deviation) of participants at baseline was 50.2 (8.1) years; 580 (46.7%) were men. Thirty-six participants developed POAG [bilateral in 17 (47.2%)] over 15 years. The incidence rate of POAG per 100-person years (95% confidence interval) was 2.83 (2.6, 3.08). Compared to baseline, the reduction in mean IOP [median (range) mm Hg] was -0.75 (-7.5, 9) in participants with incident POAG and -2.5 (-14.5, 14.5) in those without. The inter-visit difference in mean IOP was a significant risk factor on logistic regression analysis. CONCLUSION: We report the long-term incidence of POAG in rural India. A longitudinal change in IOP, specifically a less pronounced reduction in IOP with increasing age, was a novel risk factor.

The association between complement component 2/complement factor B polymorphisms and age-related macular degeneration: a HuGE review and meta-analysis.

The authors performed a systematic review of the association of complement component 2(C2)/complement factor B (CFB) gene polymorphisms with age-related macular degeneration (AMD). In total, data from 19 studies published between 2006 and 2011 were pooled for 4 polymorphisms: rs9332739 and rs547154 in the C2 gene and rs4151667 and rs641153 in the CFB gene. Data extraction and assessments for risk of bias were independently performed by 2 reviewers. Allele frequencies and allele and genotypic effects were pooled. Heterogeneity and publication bias were explored. Pooled minor allele frequencies for all 4 SNPs were between 4.7% and 9.6% for all polymorphisms, except for an Indian population in which the C allele at rs9332739 was the major allele. For the C2 polymorphisms, the minor C allele at rs9332739 and the minor T allele at rs547154 carried estimated relative risks (odds ratios) of 0.55 (95% confidence interval (CI): 0.46, 0.65) and 0.47 (95% CI: 0.39, 0.57), respectively. For the CFB polymorphisms, the minor A alleles at rs4151667 and rs614153 carried estimated risks of 0.54 (95% CI: 0.45, 0.64) and 0.41 (95% CI: 0.34, 0.51), respectively. These allele effects contributed to an absolute lowering of the risk of all AMD in Caucasian populations by 2.0%-6.0%. This meta-analysis provides a robust estimate of the protective association of C2/CFB with AMD.

A polymorphism in the CYP1B1 promoter is functionally associated with primary congenital glaucoma.

Primary congenital glaucoma (PCG) is a childhood autosomal-recessive disorder caused by developmental defects in the trabecular meshwork and anterior chamber angle. These defects cause raised intraocular pressure (IOP) that damages the optic nerve and if left untreated, results in irreversible blindness. Mutations in CYP1B1 gene at the GLC3A locus (2p21) are associated with PCG. However, there has been very limited exploration of its promoter region. We resequenced the CYP1B1 promoter in a large cohort (n = 835) that included patients with PCG (n = 301), other primary glaucomas (primary open-angle glaucoma: n = 115 and primary angle closure glaucoma: n = 100) and unaffected controls (n = 319). We functionally characterized one associated variant by luciferase reporter assay using the trabecular meshwork (TM3) cell line. We found evidence of strong (P = 6.01 × 10(-4)) association of rs2567206 (T2805C) SNP in PCG and not in other primary glaucomas. Luciferase assay indicated a ∼90% reduction in CYP1B1 promoter activity in the risk-allele (C) compared to the other allele (T). The association of the risk allele was stronger in cases harboring homozygous CYP1B1 mutations (P = 3.42 × 10(-12)). The risk haplotype 'C-C-G' in the promoter had a strong non-random association to the previously characterized risk haplotype 'C-G-G-T-A' in the coding region. The independent effect of genotype at the promoter T2805C locus (P = 0.001), and the interaction effect of genotypes at the promoter and coding region mutations loci (P = 0.001) were significant for the presenting IOP of the worst affected eye. This is the first study that unequivocally shows the functional involvement of a CYP1B1 promoter variant in PCG.

Immune defense single nucleotide polymorphisms and recruitment strategies associated with contact lens keratitis.

PURPOSE: To determine the association of single nucleotide polymorphisms (SNPs) of defensin 1B and toll-like receptor 4 with contact lens keratitis susceptibility and severity, and to understand the factors that influence study participation. DESIGN: Retrospective, case-control study. PARTICIPANTS: Ninety cases of keratitis and 185 controls recruited from studies conducted at Moorfields Eye Hospital and throughout Australia from 2003 to 2005 were analyzed for genetic associations. The reasons for participation of a subset of 146 participants from 1 site were also investigated. METHODS: Buccal swab samples were collected on Whatman FTA cards and mailed by post for analysis. DEFB1 (rs1799946, -52, rs1800972, -44, and rs11362, -20) and TLR4 (rs4986790, D299G) SNPs were screened by pyrosequencing and analyzed using a regression model for susceptibility (sterile, microbial keratitis [MK], controls) and severity. Study participation was investigated for age, gender, condition, and phone follow-up also using regression analysis. MAIN OUTCOME MEASURES: Relative risk of developing contact lens-related keratitis and more severe forms of the disease based on genetic profiles. RESULTS: Carriers of risk alleles of DEFB1 -52 and -20 showed a trend toward increased susceptibility to keratitis (-52: odds ratio [OR], 1.45; 95% confidence interval [CI], 0.99-2.11; P = 0.051; -20: OR, 1.37; 95% CI, 0.95-1.98; P = 0.088). A DEFB1 promoter haplotype (G-G-A) had a tendency toward decreased susceptibility of MK (OR, 0.68; 95% CI, 0.45-1.03; P = 0.062) and reduced severity (OR, 0.56; 95% CI, 0.30-1.07; P = 0.066). The TLR4 D299G was not associated with type and severity of keratitis. Older age (OR, 1.07; 95% CI, 1.05-1.08) and follow-up phone call (OR, 2.0; 95% CI, 1.2-3.5) were independent predictors of study participation. CONCLUSIONS: Genetic variation in DEFB1 that may lead to decreased protein expression of hBD-1 exhibits a tendency toward increased susceptibility and severity of contact lens-related keratitis. Investigation of these and other hBD genes that play important roles in animal models in a larger sample size is warranted. The approach of requesting samples from retrospective case series was generally feasible, although significant resources, including repeat phone calls, are required. More targeted strategies to recruit younger individuals to participate in genetic studies may be useful.

Association of single nucleotide polymorphisms of interleukins-1ß, -6, and -12B with contact lens keratitis susceptibility and severity.

PURPOSE: To investigate whether single nucleotide polymorphisms (SNPs) in interleukin (IL)-1ß, IL-6, and IL-12ß are associated with the susceptibility and severity of contact lens-related keratitis. DESIGN: Retrospective, case control study. PARTICIPANTS: One hundred twelve cases of keratitis and 225 controls were recruited from studies conducted at Moorfields Eye Hospital and in Australia during 2003 through 2005. METHODS: Buccal swab samples were collected on Whatman FTA cards and were mailed by post for analysis. IL-1ß (-31), IL-6 (-174, -572, -597), and IL-12B (3'+1158) genotypes were analyzed with pyrosequencing and analyzed using a regression model for susceptibility (sterile, microbial keratitis, controls) and severity. Statistical significance was set at 0.05. MAIN OUTCOME MEASURES: The relative risk of developing contact lens-related keratitis and more severe forms of the disease based on allele, genotype, and haplotype associations. RESULTS: Carriers of IL-6 SNPs were more likely to experience moderate and severe events compared with those with nonmutated genotypes (-174 heterozygous: odds ratio [OR], 3.1; 95% confidence interval [CI], 1.1-8.3; homozygous: OR, 6.4; 95% CI, 1.4-28.4; -174/-597: OR, 4.1; 95% CI, 1.6-11.0). More severe keratitis and microbial keratitis were less likely to occur in wearers with the nonmutated IL-6 haplotype (severity OR, 0.4 [95% CI, 0.2-0.7]; microbial OR, 0.6 [95% CI, 0.4-0.9]). Wearers carrying an IL-12B SNP had an increased risk of sterile keratitis (OR, 9.7; 95% CI, 1.2-76.9) compared with controls. CONCLUSIONS: The IL-6 SNPs are known to reduce protein expression of this cytokine and thus ocular immune defense, and carriers of these SNPs were more likely to experience more severe and microbial keratitis, suggesting that IL-6 decreases the severity and susceptibility of contact lens-related keratitis. Carriers of a functional SNP of IL-12B that is known to increase IL-12 expression and stability are more likely to experience sterile keratitis, suggesting that this is associated with the intense inflammatory reaction that occurs in this condition.

Phenotypic variations in ocular features among siblings with oculocutaneous albinism.

Purpose: To assess the clinical profiles, presenting ocular features, and variations in the phenotypic features in siblings with oculocutaneous albinism (OCA). Methods: Electronic medical records of consecutive siblings diagnosed with albinism from January 2016 to December 2020 were reviewed to identify the affected siblings. The variations in their phenotypic characteristics were studied. Results: Significant variations were observed in the clinical features between the siblings (n = 42). A difference of >2 lines in visual acuity was observed in 50% (n = 21) of the sibling pairs. Compound hyperopic astigmatism was the commonest refractive error. The refractive status was different in 80.95% (n = 34) pairs. Although individually strabismus and abnormal head posture were observed in one-third and one-fourth of individual children, respectively, both siblings with similar strabismus were seen in only 16.67% (n = 7) and with a similar abnormal head posture in 13.33% (n = 5). Nystagmus was the most consistent finding across these siblings with a similar nature of horizontal jerk or pendular in 65% of sibling pairs. Conclusion: This study observed significant variations in phenotypic presentations among siblings with OCA. Such differences in clinical manifestations and severity would be helpful in appropriate counseling of these families as the need for rehabilitation services is likely to vary across siblings.

Association of Single-Nucleotide Polymorphisms in Interleukin Genes with Microbial Keratitis in a South Indian Population.

BACKGROUND: To examine the relationship between single-nucleotide polymorphisms (SNPs) in interleukin (IL) genes and keratitis and its clinical manifestations. METHODS: SNPs in IL1B, IL6, CXCL8, IL10, and IL12B were analysed. Differences in frequencies of alleles, genotypes and haplotypes between cases and controls as well as associations between SNPs and clinical variables were calculated by χ2 tests with odds ratios. RESULTS: The minor homologous genotype in IL1B rs16944 (p = 0.036; odds ratio (OR) = 2.063, 95% confidence interval (CI): 1.048-4.061) and CXCL8 rs4073 (p = 0.041; OR = 0.463, 95% CI: 0.224-0.956) and the heterologous genotypes in IL6 rs1800795 (p = 0.046; OR = 0.563, 95% CI: 0.326-0.972) and IL12B rs2569254 (p = 0.0446; OR = 0.557, 95% CI: 0.314-0.989) or rs730691 (p = 0.0051; OR = 0.451, 95% CI: 0.260-0.784) were associated with keratitis. The minor genotype of rs16944 was associated with severe infection (p = 0.046). The heterologous genotype in rs2569254 was associated with hospital admission, photophobia, and mode of contact lens wear (p ≤ 0.041). The heterologous genotype in rs730691 was associated with blurred vision, discharge, anterior chamber reaction, and mode of wear (p ≤ 0.047). CONCLUSIONS: This study demonstrates that SNPs in IL1B and CXCL8 are associated with risk of developing keratitis. The study also found relationships between SNPs and clinical measures of keratitis. The potential for ethnic differences in frequency of SNPs and their association with keratitis should be followed up using different populations.