Genetic variants at 13q12.12 are associated with high myopia in the Han Chinese population.

High myopia, which is extremely prevalent in the Chinese population, is one of the leading causes of blindness in the world. Genetic factors play a critical role in the development of the condition. To identify the genetic variants associated with high myopia in the Han Chinese, we conducted a genome-wide association study (GWAS) of 493,947 SNPs in 1088 individuals (419 cases and 669 controls) from a Han Chinese cohort and followed up on signals that were associated with p < 1.0 × 10(-4) in three independent cohorts (combined, 2803 cases and 5642 controls). We identified a significant association between high myopia and a variant at 13q12.12 (rs9318086, combined p = 1.91 × 10(-16), heterozygous odds ratio = 1.32, and homozygous odds ratio = 1.64). Furthermore, five additional SNPs (rs9510902, rs3794338, rs1886970, rs7325450, and rs7331047) in the same linkage disequilibrium (LD) block with rs9318086 also proved to be significantly associated with high myopia in the Han Chinese population; p values ranged from 5.46 × 10(-11) to 6.16 × 10(-16). This associated locus contains three genes-MIPEP, C1QTNF9B-AS1, and C1QTNF9B. MIPEP and C1QTNF9B were found to be expressed in the retina and retinal pigment epithelium (RPE) and are more likely than C1QTNF9B-AS1 to be associated with high myopia given the evidence of retinal signaling that controls eye growth. Our results suggest that the variants at 13q12.12 are associated with high myopia.

Optic disc imaging with spectral-domain optical coherence tomography: variability and agreement study with Heidelberg retinal tomograph.

OBJECTIVE: To evaluate the agreement of optic disc measurements obtained with the Cirrus high-density optical coherence tomography (HD-OCT) and the Heidelberg retina tomograph (HRT) and compare the intervisit, test-retest variability between the instruments. DESIGN: Prospective, cross-sectional study. PARTICIPANTS: Two hundred seven subjects (109 glaucoma and 98 normal subjects). METHODS: One eye from each individual was selected randomly for optic disc imaging by the Cirrus HD-OCT and the HRT. Areas of the optic disc and the cup, cup volume, vertical cup-to-disc ratio and cup-to-disc area ratio were compared between the instruments. The OCT measurements were corrected for ocular magnification using the Littman's formula. The measurement agreement was evaluated with the Bland-Altman plots. The intervisit test-retest variability was examined in 17 randomly selected glaucoma patients who underwent optic disc imaging weekly for 8 consecutive weeks. The intraclass correlation coefficients (ICC) and the reproducibility coefficients of the optic disc parameters were computed. MAIN OUTCOME MEASURES: Measurement agreement, reproducibility coefficients, and ICCs of optic disc parameters. RESULTS: The OCT measured smaller optic disc and rim areas and greater cup volume, vertical cup-to-disc ratio and cup-to-disc area ratio than the HRT did (all with P<0.001). There were proportional biases in the Bland-Altman plots between OCT and HRT optic disc measurements except for rim area and cup-to-disc area ratio. The 95% limits of agreement of rim area ranged between -0.28 and 0.88 mm(2) before, and between -0.22 and 0.92 mm(2) after correction for ocular magnification. Both OCT and HRT showed high test-retest reproducibility with ICCs ≥ 0.921. Although the reproducibility coefficient of OCT rim area (0.093 mm(2); 95% confidence interval [CI], 0.081-0.105 mm(2)) was significantly smaller than that of the HRT (0.186 mm(2); 95% CI, 0.163-0.210 mm(2); P = .018), there were no differences in the ICCs between the instruments. CONCLUSIONS: Optic disc assessment by spectral-domain OCT and confocal scanning laser ophthalmoscopy demonstrates poor agreement but similarly low test-retest variability. The source of their disagreement and its effects on the detection of progression require further study.

Retinal nerve fiber layer imaging with spectral-domain optical coherence tomography: patterns of retinal nerve fiber layer progression.

OBJECTIVE: To examine the use of the retinal nerve fiber layer (RNFL) thickness map generated by a spectral-domain optical coherence tomography (OCT) to detect RNFL progression and identify the pattern of progressive changes of RNFL defects in glaucoma. DESIGN: Prospective, longitudinal study. PARTICIPANTS: One hundred eighty-six eyes of 103 glaucoma patients. METHODS: Patients were followed at 4-month intervals for ≥ 36 months for RNFL imaging and visual field examination. Both eyes were imaged by the Cirrus HD-OCT (Carl Zeiss Meditec Inc., Dublin, CA) and had visual field testing at the same visits. We defined RNFL progression by Guided Progression Analysis (Carl Zeiss Meditec) of serial RNFL thickness maps. The pattern of RNFL progression was evaluated by comparing the baseline RNFL thickness deviation map and the RNFL thickness change map. Visual field progression was defined by trend analysis of visual field index and event analysis based on the Early Manifest Glaucoma Trial criteria. MAIN OUTCOME MEASURES: The presence and the pattern of RNFL progression. RESULTS: A total of 2135 OCT images were reviewed. Twenty-eight eyes (15.1%) from 24 patients (23.3%) had RNFL progression detected by RNFL thickness map analysis. Three RNFL progression patterns were observed: (1) widening of RNFL defects (24 eyes, 85.7%), (2) deepening of RNFL defects (2 eyes, 7.1%, both had concomitant widening of RNFL defects), and (3) development of new RNFL defects (5 eyes, 17.9%). The inferotemporal meridian (324°-336°) 2.0 mm away from the optic disc center was the most frequent location where RNFL progression was detected. Thirteen eyes (46.4%) had concomitant visual field progression; 61.5% (n = 8) of these had RNFL progression that preceded or occurred concurrently with visual field progression. Forty-two eyes from 37 patients (22.6%) had visual field progression by trend and/or event analyses without progression in the RNFL thickness map. CONCLUSIONS: Analysis of serial RNFL thickness maps generated by the spectral-domain OCT facilitates the detection of RNFL progression in glaucoma.

Evaluation of retinal nerve fiber layer progression in glaucoma: a comparison between spectral-domain and time-domain optical coherence tomography.

OBJECTIVE: To compare the performance of a spectral-domain optical coherence tomography (OCT) device and a time-domain OCT device to detect retinal nerve fiber layer (RNFL) progression in glaucoma patients. DESIGN: Prospective study. PARTICIPANTS: One hundred twenty-eight eyes of 81 glaucoma patients. METHODS: Patients were followed up at 4-month intervals for at least 24 months for RNFL imaging and visual field examination. Both eyes were imaged by the Cirrus HD-OCT (Carl Zeiss Meditec, Inc., Dublin, CA) and the Stratus OCT (Carl Zeiss Meditec, Inc.) and underwent visual field testing at the same visit. Linear regression analyses between circumpapillary RNFL measurements (average, superior, and inferior RNFL thicknesses), visual field index (VFI), and follow-up time were performed. RNFL progression and RNFL improvement were identified when a significant negative or positive trend was detected, respectively. The agreement between the OCT instruments for progression detection was analyzed with κ statistics. MAIN OUTCOME MEASURES: Number of eyes with RNFL progression and improvement, agreement of progression detection between RNFL measurements and VFI, and rate of change of average RNFL thickness. RESULTS: Twenty-two eyes (19 patients) and 4 eyes (4 patients) had progression, and 0 and 5 eyes (5 patients) had improvement detected by the Cirrus HD-OCT and the Stratus OCT average RNFL measurements, respectively. The agreement for detection of RNFL progression was poor between the 2 OCT instruments (κ = 0.188, 0.027, and 0.267 for average, superior, and inferior RNFL thicknesses, respectively). The respective agreement between VFI and average RNFL thickness progression determined by the Cirrus HD-OCT and the Stratus OCT was 0.125 and 0.047. The rate of average RNFL thickness progression ranged between -1.52 µm/year and -5.03 µm/year for the Cirrus HD-OCT and between -2.22 µm/year and -7.60 µm/year for the Stratus OCT. CONCLUSIONS: The Cirrus HD-OCT outperformed the Stratus OCT in detecting more eyes with RNFL progression and fewer eyes with RNFL improvement. Because of reduced measurement variability, the Cirrus HD-OCT could detect changes in RNFL thickness sooner than the Stratus OCT. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Evaluation of retinal nerve fiber layer progression in glaucoma a prospective analysis with neuroretinal rim and visual field progression.

OBJECTIVE: To evaluate the performance of progression detection and the rate of change of retinal nerve fiber layer (RNFL), neuroretinal rim, and visual field measurements in glaucoma. DESIGN: Prospective study. PARTICIPANTS: One hundred eight eyes of 70 glaucoma patients. METHODS: Patients were followed up every 4 months for at least 2.9 years (median, 3.2 years) for measurement of RNFL thickness with the Stratus optical coherence tomograph (OCT) (Carl Zeiss Meditec, Dublin, CA), neuroretinal rim area with the Heidelberg Retinal Tomograph (HRT 3; Heidelberg Engineering, GmbH, Dossenheim, Germany), and visual field with the Humphrey Field Analyzer II (Carl Zeiss Meditec). Linear regression analyses were performed between visual field index (VFI), RNFL, and neuroretinal rim measurements and age, with progression defined when a significant negative trend was detected. The agreement among structural and functional measurements was evaluated with κ statistics. The mean rate of change was estimated with linear mixed modeling. MAIN OUTCOME MEASURES: The agreement on progression detection and the rate of change of RNFL, neuroretinal rim, and VFI measurements. RESULTS: A total of 1105 OCT, 1062 HRT, and 1099 visual field measurements were analyzed. The agreement of progression detection among the 3 investigations was poor (κ≤0.09). Ten eyes (9.3%; 9 patients) showed progression by average RNFL thickness, 16 (14.8%; 14 patients) by global neuroretinal rim area, and 35 (32.4%; 31 patients) by VFI. Only 1 eye (0.9%) had progression detected by all 3 methods. There were large variations in the rate of change of VFI, average RNFL thickness, and global neuroretinal rim area, with a range between -0.63% and -4.97% per year, -2.32% and -10.12% per year, and -0.61% and -8.48% per year, respectively. The respective mean rate estimates were -1.15% per year (95% confidence interval [CI], -1.56% to -0.73%), -0.70% per year (95% CI, -1.19% to -0.21%), and -1.06% per year (95% CI, -1.56% to -0.55%). CONCLUSIONS: The agreement of progression detection among RNFL, neuroretinal rim, and visual field measurements was poor, and the rate of RNFL, neuroretinal rim, and visual field progression varied considerably within and between subjects. Given this variability, interpretation of RNFL, neuroretinal rim, and VFI progression always should be evaluated on an individual basis. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Evaluation of retinal nerve fiber layer progression in glaucoma: a comparison between the fast and the regular retinal nerve fiber layer scans.

OBJECTIVE: To compare the performance of the fast (256 A-scans in each scan circle) and the regular (512 A-scans in each scan circle) retinal nerve fiber layer (RNFL) scan protocols for detection of glaucoma progression using the Stratus optical coherence tomography (OCT) device (Carl Zeiss Meditec, Dublin, CA). DESIGN: Retrospective, longitudinal study. PARTICIPANTS: One hundred twenty-nine eyes from 72 glaucoma patients. METHODS: All patients had been followed up for 2.9 to 6.1 years with a median follow-up of 4 months. All eyes had at least 4 serial RNFL measurements obtained with both the fast and the regular RNFL scans. Visual field (VF) assessment was performed on the same day as RNFL imaging. Retinal nerve fiber layer thickness and VF progression were evaluated with linear regression analysis against age. The mean rate of average RNFL thickness reduction was estimated with linear mixed modeling. MAIN OUTCOME MEASURES: The agreement of progression detection and the rate of change of RNFL thicknesses. RESULTS: A total of 1373 fast and 1373 regular RNFL scans and 1236 VF tests were analyzed. With reference to the average RNFL thickness, the fast RNFL scan detected more eyes with progression (21 eyes from 19 patients vs. 15 eyes from 13 patients) than the regular scan at a comparable level of specificity (96.9% vs. 96.1%). More eyes were found to have increasing RNFL thickness with age at individual clock hours (except for 3, 5, 6, and 11 o'clock) when the measurements were obtained with the regular scan. The agreement between the fast and the regular scan for detection of RNFL progression was fair to moderate, with κ values ranging between 0.14 and 0.49. The rate of average RNFL thickness progression was -1.01 µm per year for the fast RNFL scan and -0.77 µm per year for the regular scan. CONCLUSIONS: The choice of scan protocols in the Stratus OCT has a significant impact in the evaluation of RNFL progression. The fast RNFL scan seems to be preferable to follow RNFL damage in glaucoma.

Willingness to pay for cataract surgery in baiyin district, northwestern China.

PURPOSE: To investigate willingness to pay for cataract surgery, and its associations, in Northwestern China. METHODS: Four hundred thirty-eight persons aged 50 years and above, diagnosed with cataract indicated for surgery, identified in an outreach screening program were included. Subjects were offered a willingness-to-pay interview for the maximal amount that the subjects would be willing to pay for a cataract surgery. Age, gender, literacy, education level, occupation, and annual household income were recorded. RESULTS: Among 328 (74.9%) subjects who completed the interview, 197 (60.1%) participants were willing to pay something for the cataract surgery (mean, 902.9 ± 856.7 renminbi[RMB], [US$ 145 ± 137]; median, 500RMB, US$ 78). Individuals with presenting visual acuity (PVA) in the worse eye ≤6/60 (OR: 2.1, 95% CI: 1.3-3.2) and a high annual household incomes (OR: 2.0, 95% CI: 0.9-4.6) were likely to be willing to pay for the surgery, as revealed in the regression models. Willingness to pay any amount for cataract surgery was more likely among literate persons (OR: 1.5, 95% CI: 1.0-2.4) and persons with non-agricultural occupation (OR: 1.8, 95% CI: 1.0-3.2). CONCLUSIONS: The amount that subjects were willing to pay is significantly less than the current cost of cataract surgery (5000 RMB, US$320) in the area. Providing low-cost cataract surgery to patients in a financially sustainable manner is important to increase uptake of cataract surgery among rural residents in Northwest China.

Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells.

PURPOSE: gammaD-Crystallin (CRYGD) is a major structural lens crystallin and its mutations result in congenital cataract formation. In this study, we attempted to correct the altered protein features of G165fsX8 CRYGD protein with small chemical molecules. METHODS: Recombinant FLAG-tagged mutants (R15C, R15S, P24T, R61C, and G165fsX8) of CRYGD were expressed in COS-7 cells and treated with small chemical molecules with reported protein chaperoning properties (sodium 4-phenylbutyrate [4-PBA], trimethylamine N-oxide [TMAO], and glycerol and DMSO [DMSO]). Protein solubility in 0.5% Triton X-100 and subcellular distribution was examined by western blotting and immunofluorescence, respectively. Apoptosis was assayed as the percentage of fragmented nuclei in transfected cells. Expression of heat-shock proteins (Hsp70 and Hsp90) was examined by reverse transcription-polymerase chain reaction analysis. RESULTS: Unlike WT and most mutants (R15C, R15S, P24T, and R61C) of CRYGD, G165fsX8 CRYGD was significantly insoluble in 0.5% Triton X-100. This insolubility was alleviated by dose-dependent 4-PBA treatment. The treatment relieved the mislocalization of G165fsX8 CRYGD from the nuclear envelope. Also, 4-PBA treatment reduced cell apoptosis and caused an upregulation of Hsp70. CONCLUSIONS: 4-PBA treatment reduced the defective phenotype of mutant G165fsX8 CRYGD and rescued the affected cells from apoptosis. This could be a potential treatment for lens structural protein and prevent lens opacity in cataract formation.

Ethnicity-based subgroup meta-analysis of the association of LOXL1 polymorphisms with glaucoma.

PURPOSE: To investigate the association and ethnic heterogeneity of lysyl oxidase-like 1 (LOXL1) single nucleotide polymorphisms (SNPs) with exfoliation syndrome (XFS)/exfoliation glaucoma (XFG) and other types of glaucoma. METHODS: We performed meta-analysis and ethnicity-based subgroup analyses according to published studies. Allele and genotype frequencies of SNPs rs1048661, rs2165241, and rs3825942 were extracted for analysis in Reviewer Manager: (1) comparison of the allelic distributions between XFS and XFG, (2) allelic association of LOXL1 SNPs with XFS/XFG, (3) associations in homozygote, heterozygote, and dominant and recessive models, and (4) allelic association with primary open angle glaucoma (POAG). RESULTS: In total 24 reported articles were retrieved, including Caucasian, African, Japanese, Indian, and Chinese populations. There was no significant difference in the distributions of rs1048661, rs2165241, and rs3825942 between XFS and XFG. The G allele of rs3825942 was the common at-risk allele for XFS/XFG in all populations with a total odds ratio (OR) of 10.89. The total homozygote OR of rs3825942 was 9.06 for XFS/XFG combined, but the total heterozygote OR was not significant. We also found that in the recessive model, the total OR was 14.70. There was no association of the three SNPs with POAG. CONCLUSIONS: The association of rs3825942, but not rs2165241 or rs1048661, with XFS/XFG is consistent in different ethnic populations in the recessive model. LOXL1 is not associated with POAG in all study populations.

Immunopanning purification and long-term culture of human retinal ganglion cells.

PURPOSE: To establish a robust method to isolate primary retinal ganglion cells (RGCs) from human fetal retina for long-term culture while maintaining neuronal morphology and marker protein expression. METHODS: A total of six human retinas were obtained from aborted fetuses at 10 to 12 weeks of gestation with informed consent from mothers. RGCs were isolated and purified by a modified two-step immunopanning procedure. The cells were maintained in a serum-free defined medium supplemented with brain-derived neurotrophic factor, ciliary neutrophic factor, and forskolin. The viable RGCs and the extent of neurite outgrowth were examined by calcein-acetoxymethylester assay. Expression of RGC markers was studied by immunocytochemistry. RESULTS: Primary RGCs from human fetal retinas were isolated and maintained in vitro for one month with substantial neurite elongation. In cell culture, almost 70% of the isolated cells attached, spread, and displayed numerous dendrites. They were immunoreactive to RGC-specific markers (Thy-1, TUJ-1, and Brn3a) and negative for glial fibrillary acidic protein and amacrine cells marker HPC-1. CONCLUSIONS: Human RGCs were successfully isolated and maintained in long-term culture. This can serve as an ideal model for biologic, toxicological, and genomic assays of human RGCs in vitro.

Retinal nerve fiber layer imaging with spectral-domain optical coherence tomography a study on diagnostic agreement with Heidelberg Retinal Tomograph.

OBJECTIVE: To evaluate and compare the diagnostic agreement and performance for glaucoma detection between a confocal scanning laser ophthalmoscope and a spectral-domain optical coherence tomograph (OCT). DESIGN: Prospective, cross-sectional study. PARTICIPANTS: One hundred fifty-five subjects (79 glaucoma and 76 normal subjects). METHODS: One eye from each individual was selected randomly for optic disc and retinal nerve fiber layer (RNFL) imaging by the Heidelberg Retinal Tomograph (HRT; Heidelberg Engineering, GmbH, Dossenheim, Germany) and the Spectralis OCT (Heidelberg Engineering), respectively. Glaucoma was defined based on the presence of visual field defects with the Humphrey visual field analyzer (Carl Zeiss Meditec, Dublin, CA). The agreement of the categorical classification ("within normal limits," "borderline," and "outside normal limits") at the temporal, superotemporal, superonasal, nasal, inferonasal and inferotemporal sectors of the optic disc were evaluated (kappa statistics). The diagnostic sensitivity and specificity between optic disc and RNFL assessment were compared (McNemar's statistics). Area under the receiver operating characteristic curve (AUC) of OCT RNFL and HRT optic disc parameters were computed after adjustment of age, axial length, and optic disc area. MAIN OUTCOME MEASURES: Agreement of categorical classification, AUC of optic disc, and RNFL parameters. RESULTS: The agreement of categorical classification between HRT and Spectralis OCT were fair to moderate (kappa ranged between 0.30 and 0.53) except for global (kappa = 0.63) and inferotemporal (kappa = 0.68) measurements. Defining glaucoma as having "outside normal limits" in the global and/or in >or=1 of the sectoral measurements, the respective sensitivities of Spectralis OCT and HRT were 91.1% and 79.8% (P = 0.012) at a similar level of specificity (97.4% and 94.7%). The AUC of OCT global RNFL thickness (0.978) was greater than those of HRT global rim area (0.905), vertical cup-disc ratio (0.857), rim-disc area ratio (0.897), and multivariate discriminant analysis (0.880-0.925; all with P

Trimethylamine N-oxide alleviates the severe aggregation and ER stress caused by G98R alphaA-crystallin.

PURPOSE: Crystallins are major functional and structural proteins in mammalian lens. Their expression, distribution, and protein-protein interaction affect lens development and fiber cell differentiation. Mutated crystallins lead to structural and functional changes of lens structure and could lead to opacity formation and cataract development. The purpose of this study was to investigate the biological effects of the cataract-causing G98R mutation on the alphaA-crystallin (CRYAA) protein and to test the capability of chemical chaperone trimethylamine N-oxide (TMAO) to reverse such effects. METHODS: Myc/His-tagged, human, full-length, wild-type (WT) or G98R CRYAA was expressed in human lens epithelial B3 cells and treated or not treated with TMAO. Triton X-100 (Tx) solubility and cellular localization of CRYAA were examined by western blotting and confocal immunofluorescence, respectively. Ubiquitin proteasome-associated degradation was assayed by MG132 treatment. Endoplasmic reticulum (ER) stress, unfolded protein response, and apoptosis were analyzed by the expression of phosphorylated protein kinase-like ER-kinase, binding immunoglobulin protein (BiP), C/EBP homologous protein/growth arrest and DNA damage-inducible gene 153 (CHOP/GADD153), and caspase-3 and immunocytochemistry. Changes in heat shock and stress signaling were investigated. RESULTS: When transfected in lens epithelial B3 cells, unlike WT CRYAA located in the cytoplasm, the G98R CRYAA mutant formed aggregates inside the ER and the protein was predominantly Tx-insoluble. ER stress was induced by G98R CRYAA expression, and cells underwent apoptosis, as shown by a more frequent appearance of fragmented nuclei. Treatment with TMAO reduced Tx-insoluble mutant protein in time- and dose-dependent manners. Other chemical chaperones, 4-phenylbutyric acid, dimethysulfoxide, and glycerol, were much less effective than TMAO. ER-associated aggregates were reduced after TMAO treatment, and the protein was degraded through the ubiquitin-proteasome pathway. This alleviated ER stress and resulted in less apoptosis. Moreover, TMAO treatment induced a moderate upregulation of heat shock protein 70, indicating its effect on heat-shock response to modulate protein folding and assembly. No change was found for nontransfected cells after TMAO treatment. CONCLUSION: The natural osmolyte and chemical chaperone TMAO reduced the aggregation of G98R CRYAA. This alleviated ER stress and rescued the affected cells from apoptosis. Our results showed that the chemical chaperone reduces mutant CRYAA aggregates in lens cells. We suggest a potential chemical-based strategy to reduce lens opacity formation.

An alphaA-crystallin gene mutation, Arg12Cys, causing inherited cataract-microcornea exhibits an altered heat-shock response.

PURPOSE: To investigate the clinical features and molecular basis of inherited cataract-microcornea caused by an alphaA-crystallin gene (CRYAA) mutation in a Chinese family. METHODS: A three-generation Chinese family with members having autosomal dominant cataract and microcornea was recruited. Genomic DNA from peripheral blood or buccal swab samples of five affected and five unaffected members were obtained. Based on 15 genes known to cause autosomal dominant cataract, single nucleotide polymorphisms (SNPs) or microsatellite markers were selected and genotyped for two-point linkage analysis. Direct sequencing was performed to identify the disease-causing mutation. The expression construct coding for recombinant COOH-terminal myc-His-tagged wild type or R12C alphaA-crystallin protein (CRYAA) was expressed in COS-7 cells. Detergent solubility and subcellular distribution of wild type and R12C CRYAA were examined by western blotting and immunofluorescence, respectively. Heat-shock response was monitored by quantitative polymerase chain reaction (qPCR) of heat-shock proteins 70 and 90alpha (HSP70 and HSP90alpha). RESULTS: The five affected family members showed variable lens opacities and microcornea. Clinical features of cataract were asymmetric in two eyes of some affected subjects. A heterozygous missense substitution, c.34C>T, in CRYAA, which is responsible for the R12C amino acid change, segregated with autosomal dominant cataract (ADCC) in this family. This substitution was absent in 103 unrelated controls. When expressed in COS-7 cells, the R12C mutant CRYAA resembled the wild type protein in its solubility when extracted with 0.5% Triton X-100 and with its cytoplasmic localization. However, mutant cells exhibited an altered heat-shock response, evidenced by the delayed expression of HSP70, when compared to cells expressing wild type CRYAA. CONCLUSIONS: The R12C mutation in CRYAA was responsible for a variable type of inherited cataract associated with microcornea in this Chinese family. The altered heat-shock response of mutant cells suggested a change of chaperoning capacity and networking, which could be associated with the pathogenesis of hereditary cataract-microcornea syndrome.

A novel gammaD-crystallin mutation causes mild changes in protein properties but leads to congenital coralliform cataract.

PURPOSE: To identify the genetic lesions for congenital coralliform cataract. METHODS: Two Chinese families with autosomal dominant coralliform cataract, 12 affected and 14 unaffected individuals, were recruited. Fifteen known genes associated with autosomal dominant congenital cataract were screened by two-point linkage analysis with gene based single nucleotide polymorphisms and microsatellite markers. Sequence variations were identified. Recombinant FLAG-tagged wild type or mutant gammaD-crystallin was expressed in human lens epithelial cells and COS-7 cells. Protein solubility and intracellular distribution were analyzed by western blotting and immunofluorescence, respectively. RESULTS: A novel heterozygous change, c.43C>A (R15S) of gammaD-crystallin (CRYGD) co-segregated with coralliform cataract in one family and a known substitution, c.70C>A (P24T), in the other family. Unaffected family members and 103 unrelated control subjects did not carry these mutations. Similar to the wild type protein, R15S gammaD-crystallin was detergent soluble and was located in the cytoplasm. ProtScale and ScanProsite analyses revealed raised local hydrophobicity and the creation of a hypothetical casein kinase II phosphorylation site. CONCLUSIONS: A novel R15S mutation caused congenital coralliform cataract in a Chinese family. R15S possessed similar properties to the wild type gammaD-crystallin, but its predicted increase of hydrophobicity and putative phosphorylation site could lead to protein aggregation, subsequently causing opacification in lens.

AC and AG dinucleotide repeats in the PAX6 P1 promoter are associated with high myopia.

PURPOSE: The PAX6 gene, located at the reported myopia locus MYP7 on chromosome 11p13, was postulated to be associated with myopia development. This study investigated the association of PAX6 with high myopia in 379 high myopia patients and 349 controls. METHODS: High myopia patients had refractive errors of -6.00 diopters or greater and axial length longer than 26 mm. Control subjects had refractive errors less than -1.00 diopter and axial length shorter than 24 mm. The P1 promoter, all coding sequences, and adjacent splice-site regions of the PAX6 gene were screened in all study subjects by polymerase chain reaction and direct sequencing. PAX6 P1 promoter-luciferase constructs with variable AC and AG repeat lengths were prepared and transfected into human ARPE-19 cells prior to assaying for their transcriptional activities. RESULTS: No sequence alterations in the coding or splicing regions showed an association with high myopia. Two dinucleotide repeats, (AC)(m) and (AG)(n), in the P1 promoter region were found to be highly polymorphic and significantly associated with high myopia. Higher repeat numbers were observed in high myopia patients for both (AC)(m) (empirical p = 0.013) and (AG)(n) (empirical p = 0.012) dinucleotide polymorphisms, with a 1.327-fold increased risk associated with the (AG)(n) repeat (empirical p = 0.016; 95% confidence interval: 1.059-1.663). Luciferase-reporter analysis showed elevated transcription activity with increasing individual (AC)(m) and (AG)(n) and combined (AC)(m)(AG)(n) repeat lengths. CONCLUSIONS: Our results revealed an association between high myopia and AC and AG dinucleotide repeat lengths in the PAX6 P1 promoter, indicating the involvement of PAX6 in the pathogenesis of high myopia.

Retinal nerve fiber layer imaging with spectral-domain optical coherence tomography: a variability and diagnostic performance study.

OBJECTIVE: To evaluate and compare the retinal nerve fiber layer (RNFL) measurement variability, diagnostic sensitivity and specificity for glaucoma detection, and strength of the structure-function association obtained with a spectral-domain optical coherence tomography (OCT) device (Cirrus HD-OCT; Carl Zeiss Meditec, Inc., Dublin, CA) and a time-domain OCT device (Stratus OCT; Carl Zeiss Meditec, Inc.). DESIGN: Prospective, cross-sectional study. PARTICIPANTS: Ninety-seven normal subjects and 83 glaucoma patients. METHODS: One eye from each subject was imaged with Cirrus HD-OCT and Stratus OCT. Sixteen and 31 normal eyes were selected randomly to evaluate intravisit repeatability and intervisit reproducibility, respectively. The agreement of RNFL measurements was evaluated with Bland-Altman plots. The diagnostic sensitivity and specificity was examined with the area under the receiver operating characteristic curve (AUC). The association between average RNFL thickness and visual field sensitivity was evaluated with a second-order regression model. MAIN OUTCOME MEASURES: Retinal nerve fiber layer measurement variability, AUC, and coefficient of determination (R(2)). RESULTS: The intravisit repeatability of Cirrus HD-OCT ranged between 5.12 and 15.02 mum, and the intervisit reproducibility ranged between 4.31 and 22.01 mum. The intervisit variabilities of sectoral and average RNFL thicknesses were lower in Cirrus HD-OCT compared with Stratus OCT with significant differences at 1, 3, 4, and 8 to 11 o'clock (P< or =0.021). There were proportional biases of RNFL measurements between the 2 OCT devices. The difference of RNFL thicknesses increased with the means. The average (AUC, 0.962 for Cirrus HD-OCT and 0.956 Stratus OCT), superior (AUC, 0.963 and 0.950, respectively), and inferior (AUC, 0.949 and 0.931, respectively) RNFL thicknesses demonstrated the greatest AUCs in both OCT devices with no significant difference detected between the respective measurements (P> or =0.120). The strength of the structure-function association was comparable between Cirrus HD-OCT (R(2) = 0.580) and Stratus OCT devices (R(2) = 0.623; P = 0.918). CONCLUSIONS: Although the diagnostic performance and the strength of the structure-function association were comparable between Cirrus HD-OCT and Stratus OCT RNFL measurements, Cirrus HD-OCT demonstrated lower measurement variability compared with Stratus OCT with significant differences at 1, 3, 4, and 8 to 11 o'clock. The poor agreement was likely related to the different inherent characteristics of the 2 OCT systems.

Chronic ocular hypertension in rabbits induced by limbal buckling.

AIM: To establish a rabbit model of chronic ocular hypertension (OHT) by limbal buckling. METHODS: Eighteen New Zealand White rabbits were involved and divided into three groups. A latex encircling band of 20, 25 or 35 mm was implanted behind the limbus in the right eye of each animal. The intraocular pressure (IOP) was monitored for 8 weeks, after which optic nerve damage was evaluated by fundus photography, optical coherence tomography (OCT) retrograde labelling and histology. Meanwhile, the anterior chamber angle (ACA) was examined by OCT and gonioscopy. RESULTS: OHT was induced in all animals after surgery. The IOP peaked at 38.0±3.7, 32.0±3.9 and 24.1±6.5 mm Hg in groups 1, 2 and 3, respectively, and remained elevated for 22, 25 and 39 days on average, respectively. The elevated IOPs showed good consistency within 2 weeks, although the durations of high IOP varied moderately. The area ratio between the optic cup and disc (cup to disc area) was increased in 73% of the treated eyes, and the average changes were 0.10±0.13, 0.11±0.08 and 0.09±0.02 in groups 1, 2 and 3, respectively. The depth of the optic cup was also increased in the treated eyes, and the density of the retinal ganglion cells was reduced. Additionally, the ACA showed a dynamic change with IOP after the latter was reduced by paracentesis. CONCLUSION: Limbal buckling provides an effective method of producing chronic OHT and glaucomatous optic neuropathy in rabbits.

Gene expression profiles of human trabecular meshwork cells induced by triamcinolone and dexamethasone.

PURPOSE: Triamcinolone acetonide (TA) and dexamethasone (DEX) are corticosteroids commonly used for ocular inflammation, but both can cause ocular hypertension. In this study, the differential gene expression profile of human trabecular meshwork (TM) cells in response to treatment by TA in comparison with DEX was investigated. METHODS: Total RNA was extracted from cultured human TM cells treated with TA or DEX and used for microarray gene expression analysis. The microarray experiments were repeated three times. Differentially expressed genes were identified by an empiric Bayes approach and confirmed by real-time quantitative PCR. RESULTS: TA (0.1 mg/mL) treatment resulted in 15 genes upregulated and 12 genes downregulated, whereas 1 mg/mL TA resulted in 36 genes upregulated and 21 genes downregulated. These genes were mainly associated with acute-phase response, cell adhesion, cell cycle and growth, growth factor, ion binding, metabolism, proteolysis and transcription factor. Two genes, MYOC and GAS1, were upregulated, and three genes, SENP1, ZNF343, and SOX30, were downregulated by both TA and DEX treatment. Eight differentially expressed genes were located in known primary open-angle glaucoma (POAG) loci, including MYOC, SOAT1, CYP27A1, SPOCK, SEMA6A, EGR1, GAS1, and ATP10A. CONCLUSIONS: Differential gene expression profiles of human TM cells treated by TA and DEX, and a dosage effect by TA, were revealed by microarray technology. TA and DEX treatment shared several differentially expressed genes, suggesting a common mechanism to cause ocular hypertension. Some differentially expressed genes located in the known POAG loci are potential candidates for glaucoma genes.

Relationship between retinal nerve fiber layer measurement and signal strength in optical coherence tomography.

PURPOSE: To examine the relationship between signal strength and retinal nerve fiber layer (RNFL) thickness measured by optical coherence tomography (OCT). DESIGN: Observational cross-sectional study. PARTICIPANTS: Forty normal subjects were recruited. METHODS: Retinal nerve fiber layer (RNFL) thickness was measured by Stratus OCT (Carl Zeiss Meditec, Dublin, CA). In each eye, the focusing knob was adjusted to obtain 6 images with different signal strengths ranging from 5 to 10. The relationships between signal strength and RNFL thickness were examined using the Spearman correlation coefficient. The differences of RNFL thicknesses were compared with repeated-measures analysis of variance. MAIN OUTCOME MEASURES: Retinal nerve fiber layer thicknesses measured at different signal strengths. RESULTS: Significant differences were observed between measurements obtained at signal strength of 10 and those obtained with signal strength of less than 10 at the superior, nasal, and temporal clock hours. RNFL thickness generally increased with the signal strength, with significant correlations found with the total average, superior, and nasal clock hours RNFL thicknesses. CONCLUSIONS: Optical coherence tomography RNFL measurements vary significantly with signal strength. Obtaining the maximal possible signal strength is recommended for RNFL thickness measurement.

Comparative study of central corneal thickness measurement with slit-lamp optical coherence tomography and visante optical coherence tomography.

PURPOSE: To evaluate the repeatability and reproducibility of central corneal thickness (CCT) measurements obtained by 2 anterior segment optical coherence tomography (OCT) imaging systems and to examine their agreements with ultrasound pachymetry. DESIGN: Observational cross-sectional study. PARTICIPANTS: Fifty eyes from 50 healthy normal subjects were recruited. METHODS: In one randomly selected eye in each subject, CCT was measured by slit-lamp OCT (SLOCT), Visante OCT, and ultrasound pachymetry. For anterior segment OCT measurements, both automatic and manual CCTs were obtained. Twenty-five of the 50 subjects were invited for 2 more visits within a week to evaluate repeatability and reproducibility of CCT measurement. MAIN OUTCOME MEASURES: Central corneal thickness measurement obtained by the 3 methods and their agreements. Intrasession and intersession within-subject standard deviation (S(w)), precision (1.96xS(w)), coefficient of variation (CV(w)) (100xS(w)/overall mean), and intraclass correlation coefficient (ICC) were calculated to evaluate repeatability and reproducibility. RESULTS: Good repeatability and reproducibility were found for both automatic and manual CCT measurements obtained by SLOCT and Visante OCT. For intrasession repeatability, CV(w) and ICC values ranged between 0.9% and 1.2% and 0.96 and 0.98, respectively. For intersession reproducibility, the respective CV(w) and ICC values ranged between 1.2% and 1.4% and 0.94 and 0.96. Although no significant difference was found between automatic/manual SLOCT measurements and ultrasound pachymetry, automatic Visante OCT CCT (535.7+/-30.2 microm) was significantly less than CCT with ultrasound pachymetry (550.3+/-31.14 microm) (P<0.001). In contrast, manual Visante OCT measurement (558.8+/-32.8 microm) was slightly higher than ultrasound pachymetry (P<0.001). Nevertheless, SLOCT and Visante OCT measurement of CCT had 95% limits of agreement comparable to that of ultrasound pachymetry. The best agreement was observed in the manual SLOCT measurement (95% limits of agreement between -15.5 and 11.7 microm). CONCLUSIONS: Both SLOCT and Visante OCT automatic and manual CCT measurements were reliable and showed comparable agreement with ultrasound pachymetry. Although the 2 anterior segment OCT imaging systems have similar design and working principles, clinicians should be aware of the differences in CCT measurement between the 2 anterior segment OCTs.