Showing papers in "Investigative Ophthalmology & Visual Science in 2008"

Role of Near Work in Myopia: Findings in a Sample of Australian School Children

Abstract: PURPOSE. To examine the association of time spent in near work and reading with spherical equivalent refraction (SER) in a population-based sample of 12-year-old Australian schoolchildren. METHODS. Data on the time spent in near-work or outdoor activities per week and estimates for the duration of continuous reading and reading distances, were collected in questionnaires (2353 participants, 75.3% response) in the Sydney Myopia Study between 2004 and 2005; 2339 children underwent a comprehensive eye examination, including cycloplegia. RESULTS. Longer time spent on reading for pleasure and reports of close reading distance (30 cm) were associated with a more myopic refraction after adjustment for age, sex, ethnicity, and school type (Ptrend 0.02 and P 0.0003, respectively). Time spent in individual near-work activities, however, correlated poorly with SER (all r 0.2) and was not significant in multivariate analyses for myopia (SER 0.50 D), with adjustment for age, sex, ethnicity, parental myopia, school type, and outdoor activity. Children of European Caucasian ethnicity reported spending marginally less time in near work than children of East Asian ethnicity (26.0 h/wk vs. 32.5 h/wk, P 0.0001). East Asian ethnicity, however, was associated with substantially greater odds of having myopia (odds ratio [OR], 11.0; 95% confidence interval [CI], 7.0‐17.4). Near work such as close reading distance (30 cm) and continuous reading (30 minutes) independently increased the odds of having myopia in this sample of children. CONCLUSIONS. Although myopia was not significantly associated with time spent in near work after adjustment for other factors, there were significant independent associations with close reading distance and continuous reading. These associations may indicate that the intensity rather than the total duration of

Opa1 Mutations Induce Mitochondrial Dna Instability and Optic Atrophy Plus Phenotypes

Abstract: Mutations in OPA1, a dynamin-related GTPase involved in mitochondrial fusion, cristae organization and control of apoptosis, have been linked to non-syndromic optic neuropathy transmitted as an autosomal-dominant trait (DOA). We here report on eight patients from six independent families showing that mutations in the OPA1 gene can also be responsible for a syndromic form of DOA associated with sensorineural deafness, ataxia, axonal sensory-motor polyneuropathy, chronic progressive external ophthalmoplegia and mitochondrial myopathy with cytochrome c oxidase negative and Ragged Red Fibres. Most remarkably, we demonstrate that these patients all harboured multiple deletions of mitochondrial DNA (mtDNA) in their skeletal muscle, thus revealing an unrecognized role of the OPA1 protein in mtDNA stability. The five OPA1 mutations associated with these DOA 'plus' phenotypes were all mis-sense point mutations affecting highly conserved amino acid positions and the nuclear genes previously known to induce mtDNA multiple deletions such as POLG1, PEO1 (Twinkle) and SLC25A4 (ANT1) were ruled out. Our results show that certain OPA1 mutations exert a dominant negative effect responsible for multi-systemic disease, closely related to classical mitochondrial cytopathies, by a mechanism involving mtDNA instability.

Intracranial pressure in primary open angle glaucoma, normal tension glaucoma, and ocular hypertension: a case-control study.

Abstract: PURPOSE To compare intracranial pressure (ICP) in subjects with primary open-angle glaucoma (POAG), normal-tension glaucoma (NTG; subset of POAG), and ocular hypertension (OHT) with that in subjects with no glaucoma.

Oncogenic mutations in GNAQ occur early in uveal melanoma.

Abstract: Purpose Early/initiating oncogenic mutations have been identified for many cancers, but such mutations remain unidentified in uveal melanoma (UM). An extensive search for such mutations was undertaken, focusing on the RAF/MEK/ERK pathway, which is often the target of initiating mutations in other types of cancer.

Upregulated IL-23 and IL-17 in Behçet patients with active uveitis.

Abstract: PURPOSE. Behcet disease (BD) is a systemic inflammatory dis- ease presumably caused by an autoimmune response. The interleukin (IL)-23/IL-17 pathway has been demonstrated to be involved in the development and maintenance of certain in- flammatory diseases. This study was designed to investigate the role of IL-23 and IL-17 in BD. METHODS. IL-23p19 mRNA in peripheral blood mononuclear cells (PBMCs) was examined using RT-PCR. The levels of IL-23, IL-17, and IFN- in sera or PBMCs were detected by ELISA. Flow cytometry was used to evaluate the frequencies of IL-17- producing and IFN--producing T cells and the expression of CD45RO. RESULTS. Results showed that the expression of IL-23p19 mRNA, IL-23, IL-17, and IFN- was markedly elevated in BD patients with active uveitis. The frequencies of IL-17-produc- ing and IFN--producing T cells from PBMCs were significantly upregulated in BD patients with active uveitis. The increased IL-17 (3.10% 0.53%) in BD patients with active uveitis was primarily produced by CD45RO memory T cells. Recombi- nant (r) IL-23 could upregulate IL-17 production by poly- clonally stimulated PBMCs, whereas interferon (IFN)- down- regulated IL-17 production. CONCLUSIONS. These findings reveal that the levels of IL-23, IL-17, and IFN- are elevated in BD patients with active uveitis, and they suggest that the IL-23/IL-17 pathway together with IFN- is associated with the active intraocular inflammation in BD patients. (Invest Ophthalmol Vis Sci. 2008;49:3058 -3064)

Reduced retina microglial activation and improved optic nerve integrity with minocycline treatment in the DBA/2J mouse model of glaucoma.

Abstract: PURPOSE. In the context of the retinal ganglion cell (RGC) axon degeneration in the optic nerve that occurs in glaucoma, microglia become activated, then phagocytic, and redistribute in the optic nerve head. The authors investigated the potential contribution of retinal microglia activation to glaucoma progression in the DBA/2J chronic mouse glaucoma model. METHODS. The authors treated 6-week-old DBA/2J mice for 25 weeks with minocycline, a tetracycline derivative known to reduce microglia activation and to improve neuronal survival in other models of neurodegenerative disease. They quantified RGC numbers and characterized microglia activation, gliosis, and both axonal integrity and retrograde tracer transport by RGCs in mice systemically treated with minocycline or vehicle only. RESULTS. Minocycline reduced microglial activation and improved RGC axonal transport and integrity, yet it had no effect on the characteristic age-related ocular changes that

Ultrahigh-speed optical coherence tomography for three-dimensional and en face imaging of the retina and optic nerve head.

Abstract: Optical coherence tomography (OCT) is an emerging optical imaging modality for biomedical research and clinical medicine.1 In the eye, OCT can provide detailed images of retinal diseases including diabetic retinopathy, age-related macular degeneration, and glaucoma.2,3 The commercial StratusOCT instrument (Carl Zeiss Meditec, Inc., Dublin, CA), with 8- to 10-μm axial image resolution, has become a standard in the diagnosis and monitoring of diseases such as wet age-related macular degeneration, diabetic macular edema, and glaucoma. Recent work in the field of ophthalmic OCT has demonstrated that spectral/Fourier domain detection methods4–7 enable OCT imaging with dramatically improved speed and sensitivity over conventional time domain methods.8–10 Spectral/Fourier domain OCT uses a broadband light source and a spectrometer to measure the interference spectrum. Light backscattered or backreflected from different positions in the sample is measured simultaneously, rather than sequentially, improving the imaging speed. The improved speed enables three-dimensional (3D) and high-definition imaging of the retina and enhanced visualization of retinal diseases.11–14 Several companies have developed clinical spectral/Fourier domain OCT instruments for retinal imaging. These instruments operate at speeds of 20,000 to 40,000 axial scans per second, 50 to 100 times faster than previous OCT instruments. Despite the dramatically improved performance of spectral/Fourier domain OCT, there are several important limitations. First, spectral/Fourier domain detection utilizes a high-speed camera and spectrometer for detection, which increases system size and complexity. Second, because of spectrometer and camera resolution limits, detection sensitivity and axial resolution change as a function of axial position or imaging depth.7,8,15 This sensitivity decrease may adversely affect the reproducibility of quantitative measurements. Finally, spectrometer losses and camera read out rates limit the maximum imaging speed. Therefore, although 3D imaging with spectral/Fourier domain OCT is possible, motion artifacts prevent reliable acquisition of 3D data sets when high sampling density is desired. For example, at an acquisition speed of 25,000 axial scans per second, a raster scan of the macula consisting of 512 × 512 axial scans would require >10 seconds. Current ophthalmic systems solve this problem by sacrificing sampling density along one axis of the raster scan or individually acquiring densely sampled data sets from different portions of the retina. The first approach does not adequately sample the retina, and the second approach requires the precise acquisition and registration of multiple data sets, which may be time-consuming and cumbersome in subjects with poor fixation. Some commercially available, high-end OCT instruments provide active eye motion tracking that enables long acquisition times and high-density data sets. However, eye tracking requires an additional optical system to determine eye motion and actuators to correct for eye motion, both of which increase system complexity and cost. Swept-source OCT10,16–19 is another approach for high-speed OCT imaging which possesses speed and sensitivity advantages similar to those of spectral/Fourier domain OCT. Both methods measure interference as a function of optical frequency or wavelength. However, swept-source OCT uses a frequency swept laser that enables measurement of interference at different optical frequencies or wavelengths sequentially over time. Swept-source OCT imaging of the retina was first demonstrated in 2006 at 18,800 axial scans per second and 14-μm axial resolution in air, using a 1050-nm frequency swept laser.20 Swept-source OCT imaging of the human retina was also demonstrated at 43,200 axial scans per second and 13 μm axial resolution in air, using an 850-nm frequency swept laser,21 and 16,000 axial scans per second and 9.5-μm axial resolution in air using an 850-nm frequency swept laser.22 Recently, swept-source OCT imaging of the human retina was demonstrated at 28,000 axial scans per second and 14.4-μm axial resolution in air with a 1050-nm frequency swept laser.23 Our group previously demonstrated swept-source OCT imaging at 236,000 axial scans per second and 19-μm axial resolution in air with a 1060-nm Fourier domain mode locked (FDML) frequency swept laser.24 In this article, OCT imaging of the retina and optic nerve head at 249,000 axial scans per second and 11-μm axial resolution in air (8 μm in the retina) are demonstrated.

Characterization of Outer Retinal Morphology with High-Speed, Ultrahigh-Resolution Optical Coherence Tomography

Abstract: Optical coherence tomography (OCT) is a medical imaging technology that can perform high resolution, cross-sectional imaging of tissue morphology in situ and in real time.1 It can provide detailed images and quantitative information about retinal structure and has therefore become a standard diagnostic technique in ophthalmology.2,3 The StratusOCT system (Carl Zeiss Meditec, Inc., Dublin, CA), with an 8- to 10-μm axial image resolution, can provide detailed cross-sectional images of retinal disease and quantitative measurements, including macular thickness, nerve fiber layer thickness, and optic nerve head topography. Normative databases for macular thickness and nerve fiber layer thickness are available to aid in the interpretation of quantitative measurements. Analysis of finer intraretinal structures has been difficult because of speed and resolution limitations. However, segmentation of the inner retinal complex, outer plexiform layer, and outer retinal complex in StratusOCT images has recently been demonstrated.4 Another study demonstrated foveal photoreceptor layer changes in central serous chorioretinopathy that correlate with visual acuity loss.5 Recently, a correlation between macular hole postoperative visual acuity and photoreceptor thickness measured manually from StratusOCT images has been shown.6 Ultrahigh-resolution (UHR) OCT imaging with axial resolutions of 2 to 3 μm was demonstrated to improve significantly the visualization of normal retinal morphology7,8 and retinal disease9–14 Although conventional UHR OCT systems that use time domain detection enable visualization of structural detail in the outer retina, quantitative mapping with conventional UHR OCT technology has been difficult, because of imaging speed limitations and consequent motion artifacts. Nevertheless, several studies have been performed in which photoreceptor thickness was measured manually on a small number of images. One study correlated photoreceptor loss and foveal thickness with visual acuity in Stargardt disease-fundus flavimaculatus.12 In another study, central foveal photoreceptor thickness measured by UHR OCT was correlated with visual acuity in retinitis pigmentosa and degenerative diseases.14 To date, comprehensive quantitative mapping of the photoreceptors and RPE has not been possible with conventional UHR OCT, due to motion artifacts and limited retinal coverage. Recently, advances in OCT technology have enabled UHR OCT imaging with a ~50× increase in imaging speed over standard resolution OCT systems and a ~100× increase in imaging speed over previous UHR OCT systems.15–17 These techniques are based on “spectral” or “Fourier domain” detection, where echo time delays of light are measured by acquiring the interference spectrum of the light signal and taking its Fourier transform.18,19 Since all the light echoes from different axial depths are measured simultaneously, sensitivity and imaging speed are dramatically increased.20–22 With increased imaging speed, individual images are acquired within a fraction of a second and depict the true retinal topography. Multiple images may be rapidly acquired at different locations or orientations, enabling three-dimensional imaging.23,24 The number of axial scans per image may be increased significantly, to obtain high-definition images.25 Adaptive optics has also been combined with high-speed OCT, to visualize the cone photoreceptors in three dimensions.26 In this study, high-speed UHR OCT was used for the visualization, measurement and mapping of outer retinal morphology in normal subjects. OCT imaging was performed with a 3.5-μm axial-resolution prototype instrument using a radial scan pattern, and scattering bands in the outer retina were measured and mapped in 43 normal subjects of various ages. Quantitative maps were used to assign these bands to specific outer retinal layers or features based on comparison with the known topographic distribution of cones and rods in the macula. The cross-sectional relationships between layer measurements and age were also investigated. In addition, high-definition imaging was performed with a 2.8-μm axial-resolution prototype instrument to visualize finer bands in the outer retina. Anatomic correlates for these more subtle outer retinal features were determined.

Amyloid-β deposits lead to retinal degeneration in a mouse model of Alzheimer disease

Abstract: Purpose To compare the temporal and spatial expression patterns of amyloid precursor protein (APP), amyloid-β deposits, inflammatory chemokines, and apoptosis in the retina of a mouse model of Alzheimer disease (AD).

High-Resolution Spectral Domain-OCT Imaging in Geographic Atrophy Associated with Age-Related Macular Degeneration

Abstract: Purpose To describe morphologic variations in outer retinal layers in eyes with atrophic age-related macular degeneration (AMD) using high-resolution, spectral-domain optical coherence tomography (SD-OCT). Methods SD-OCT scans were obtained with a combined confocal scanning laser ophthalmoscope (cSLO) and SD-OCT for simultaneous tomographic and topographic in vivo imaging. A total of 81 eyes of 56 patients (mean age, 77.8 +/- 7.4 years) with geographic atrophy (GA) were examined. Morphologic alterations were analyzed and classified in the perilesional zone, at the junction between GA and nonatrophic retina, and in the atrophic area itself. Results In the perilesional zone, distinct morphologic alterations included elevations of the outer retinal layers, thickening, and spikes of the outer hyperreflective band as well as clumps at different neurosensory retinal levels. At the junction, highly variable transitions of the outer retinal layers were present with different degrees of loss of the normal hyperreflective bands. Within the actual GA, hyperreflective clumps at different retinal levels, segmented plaques of the outer band and elevations with variable reflectivity were visualized. Conclusions SD-OCT imaging in eyes with GA revealed a wide spectrum of morphologic alterations, both in the surrounding retinal tissue and in the atrophic area. These alterations may reflect different disease stages or, alternatively, heterogeneity on a cellular and molecular level. Longitudinal studies using in vivo SD-OCT imaging may allow evaluation of the relevance of these phenotypic changes as potential predictive markers for the progression of disease (i.e., enlargement rates of GA over time) and may be used for monitoring of future therapeutic interventions.

The prevalence and types of glaucoma in malay people: the Singapore Malay eye study.

Abstract: PURPOSE. To assess the prevalence and types of glaucoma in an Asian Malay population.METHODS. The Singapore Malay Eye Study is a population-based, cross-sectional survey that examined 3280 (78.7% response) persons aged 40 to 80 years. Participants underwent a standardized clinical examination including slit-lamp biomicroscopy, Goldmann applanation tonometry, and dilated optic disc assessment. Participants who were suspected to have glaucoma also underwent visual field examination (24-2 SITA standard, Humphrey Visual Field Analyzer II), gonioscopy, and repeat applanation tonometry. Glaucoma was defined according to International Society for Geographical and Epidemiologic Ophthalmology criteria.RESULTS. Of the 3280 participants, 150 (4.6%) had diagnosed glaucoma, giving an age- and sex-standardized prevalence of 3.4% (95% confidence interval [CI], 3.3%-3.5%). The age- and sex-standardized prevalence of primary open-angle glaucoma was 2.5% ( 95% CI, 2.4%-2.6%), primary angle-closure glaucoma 0.12% ( 95% CI, 0.10%-0.14%), and secondary glaucoma 0.61% ( 95% CI, 0.59%-0.63%). Of the 150 glaucoma cases, only 12 (8%) had a previous known history of glaucoma. Twenty-seven (18%) eyes had low vision ( based on best corrected visual acuity logarithm of the minimal angle of resolution [logMAR] > 0.30 to = 1.00).CONCLUSIONS. The prevalence of glaucoma among Malay persons 40 years of age and older in Singapore is 3.4%, comparable to ethnic Chinese people in Singapore and other racial/ethnic groups in Asia. As in Chinese, Caucasians, and African people, primary open-angle glaucoma was the main form of glaucoma in this population. More than 90% of glaucoma cases were previously undetected.

Novel Hyaluronic Acid-Chitosan Nanoparticles for Ocular Gene Therapy

Abstract: Purpose Gene therapy offers a promising alternative for the treatment of ocular diseases. However, the implementation of this type of therapy is actually hampered by the lack of an efficient ocular gene delivery carrier. The main objective of the present work was to assess the effectiveness and investigate the mechanism of action of a new type of nanoparticle made of two bioadhesive polysaccharides, hyaluronic acid (HA) and chitosan (CS), intended for the delivery of genes to the cornea and conjunctiva. Methods The nanoparticles were obtained by a very mild ionotropic gelation technique. They were loaded with either the model plasmid pEGFP or pbeta-gal. Transfection and toxicological studies were conducted in human corneal epithelial (HCE) and normal human conjunctival (IOBA-NHC) cell lines. The mechanism of internalization of the nanoparticles by the corneal and conjunctival cells was investigated by using fluorescence confocal microscopy. Results The nanoparticles had a size in the range of 100 to 235 nm and a zeta-potential of -30 to +28 mV. The results of the transfection studies showed that HA-CS nanoparticles were able to provide high transfection levels (up to 15% of cells transfected), without affecting cell viability. The confocal images indicated that HA-CS nanoparticles were internalized by fluid endocytosis and that this endocytic process was mediated by the hyaluronan receptor CD44. Conclusions The results give evidence of the potential of HA-CS nanoparticles for the targeting and further transfer of genes to the ocular surface.

Corneal Hysteresis but Not Corneal Thickness Correlates with Optic Nerve Surface Compliance in Glaucoma Patients

Abstract: Purpose To investigate relationships between acute intraocular pressure (IOP)-induced optic nerve head surface deformation and corneal hysteresis and thickness in glaucomatous and nonglaucomatous human eyes. Methods This was a prospective experimental study of 100 subjects (38 with glaucoma, 62 without glaucoma). Data collected included spherical equivalent, optic disc diameter, central corneal thickness (CCT), axial length, cylinder, Goldmann IOP, Pascal IOP, and ocular pulse amplitude and ocular response analyzer (ORA) measurements of corneal hysteresis (CH). Elevation of IOP was induced in the right eye of each subject with a modified LASIK suction ring to an average of 64 mm Hg for less than 30 seconds. Heidelberg Retina Tomography II (HRT) was used to map the optic nerve surface before and during IOP elevation. Mean cup depth was calculated using built-in HRT data analysis software. Change in optic disc depth during IOP elevation was calculated for all right eyes, and tests for correlation with the parameters listed were performed. Results Both CH and CCT were lower in the glaucoma group (8.8 mm Hg and 532 microm) than in the control group (9.6 mm Hg, P = 0.012; 551 microm, P = 0.011, respectively). There were no statistically significant differences in spherical equivalent, cylinder, axial length, optic disc size, or ocular pulse amplitude between the glaucoma and the control groups. There was no difference between the amount of IOP elevation between the two groups (P = 0.41), and the average difference in mean cup depth between baseline (mean cup depth, 247 microm) and during IOP elevation was 33 microm (29.8 microm in glaucoma and 36.1 microm in control; P = 0.5). Multiple variable analysis, controlling for age and sex, showed that CH was correlated with mean cup depth increase (P = 0.032). This relationship persisted (P = 0.032) after controlling for glaucoma status in addition to age and sex. Other factors, including CCT (P = 0.3), axial length (P = 0.9), ocular pulse amplitude (P = 0.22), and spherical equivalent (P = 0.38), were not significant in this model. Conclusions In the glaucoma patients but not the control patients, CH but not CCT or other anterior segment parameters was associated with increased deformation of the optic nerve surface during transient elevations of IOP. (ClinicalTrials.gov number, NCT00328835.).

Interaction between bevacizumab and murine VEGF-A: a reassessment.

Abstract: Purpose Bevacizumab is a humanized anti-human VEGF-A monoclonal antibody (mAb) approved by the United States Food and Drug Administration for cancer therapy and used off label to treat neovascular age-related macular degeneration. Earlier studies characterized bevacizumab as species specific and lacking the ability to neutralize murine (m) VEGF-A. However, a recent study reported that bevacizumab is a potent inhibitor of hemangiogenesis and lymphangiogenesis in murine models. The authors sought to reassess the interaction between bevacizumab and mVEGF-A. Methods The authors performed Western blot analysis, plasmon resonance by BIAcore, and endothelial cell proliferation assays to characterize the interaction between bevacizumab and mVEGF-A. They also tested whether bevacizumab had any effects in two in vivo murine models, laser-induced choroidal neovascularization (CNV) and melanoma growth. Results Western blot detected a very weak interaction, but BIAcore detected no measurable interaction between mVEGF and bevacizumab. Bevacizumab failed to inhibit mVEGF-stimulated endothelial cell proliferation. In addition, bevacizumab was indistinguishable from the control antibody in the CNV and tumor models, whereas a cross-reactive anti-VEGF-A mAb had dramatic inhibitory effects. Conclusions Bevacizumab has an extremely weak interaction with mVEGF-A, which fails to result in immunoneutralization as assessed by several bioassays.

Antimicrobial efficacy of riboflavin/UVA combination (365 nm) in vitro for bacterial and fungal isolates: a potential new treatment for infectious keratitis.

Abstract: Purpose To demonstrate the antimicrobial properties of riboflavin/UVA (365 nm) against common pathogens. Methods One group of bacteria (Pseudomonas aeruginosa [PA], Staphylococcus aureus [SA], and Staphylococcus epidermidis [SE]) was tested by using Kirby-Bauer discs with (1) empty disc (Control - C); (2) riboflavin 0.1% (B2); (3) riboflavin 0.1% previously activated by UVA (B2'); (4) UVA alone (UVA); (5) group 2+additional UVA exposure (UVA+B2); and (6) group 3+additional UVA exposure (UVA+B2'). In addition, another group of microbes was tested with the same approach: methicillin-resistant S. aureus (MRSA), multidrug-resistant P. aeruginosa (MDRPA), drug-resistant Streptococcus pneumoniae (DRSP), and Candida albicans (CA). The mean growth inhibition zone (GIZ) in square millimeters was measured around the discs. The mean standard deviation (MSD) was calculated to be 3.65 when alpha = 0.01. A mean deviation (MD) > MSD indicates a significant difference. Results In the first group, the GIZ was significantly greater after UVA (MD = 14.30), UVA+B2 (MD = 39.61), and UVA+B2' (MD = 40.45) when compared with C, B2, and B2'. UVA alone was less effective than UVA+B2 (MD = 25.31) and UVA+B2' (MD = 26.15). The second group demonstrated increased GIZ in UVA (MD = 6.98), UVA+B2 (MD = 17.80), and UVA+B2' (MD = 21.15) when compared with C, B2, and B2'. UVA alone was less effective against the second group of bacteria than was UVA+B2 (MD = 10.82) and UVA+B2' (MD = 14.17). CA did not show any GIZ after treatment. Conclusions Riboflavin/UVA was effective against SA, SE, PA, MRSA, MDRPA, and DRSP, but was ineffective on CA and has the potential for use in treatment of microbial keratitis in the future.

Intravitreal injection of erythropoietin protects both retinal vascular and neuronal cells in early diabetes.

Abstract: PURPOSE. To explore and evaluate the protective effect of erythropoietin (EPO) on retinal cells of chemically induced diabetic rats after EPO was injected intravitreally at the onset of diabetes. METHODS. Diabetes was induced in Sprague-Dawley rats by intraperitoneal injection of streptozotocin (STZ). At the onset of diabetes, a single intravitreal injection of EPO (0.05-200 ng/eye) was performed. In the following 6 weeks, the blood retinal barrier (BRB) was evaluated by Evans blue permeation (EBP). Retinal cell death in different layers was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. The retinal thickness and cell counts were examined at the light microscopic level. Electron microscopy (EM) was used to scrutinize retinal vascular and neuronal injury. Neurosensory retinas of normal and diabetic rats were used as the sources of reverse transcription-polymerase chain reaction (RT-PCR) and Western blot for the detection of EPO, EPO receptor (EpoR), and products of the extracellular signal-regulated kinase (ERK) and the signal transducers and activators of transcription 5 (STAT5) pathways. The distribution of EpoR in retinal layers was demonstrated by immunohistochemistry (IHC). RESULTS. In the diabetic rats, BRB breakdown was detected soon after the onset of diabetes, peaked at 2 weeks, and reached a plateau at 2 to 4 weeks. The number of TUNEL-positive cells increased in the neurosensory retina, especially, the outer nuclear layer (ONL) at 1 week after diabetes onset and reached a peak at 4 to 6 weeks. The retinal thickness and the number of cells in the ONL were reduced significantly. EM observations demonstrated vascular and photoreceptor cell death starting soon after the onset of diabetes. All these changes were largely prevented by EPO treatment. Upregulation of EpoR in the neurosensory retina was detected at both the transcriptional and protein levels 4 to 8 weeks after the onset of diabetes, whereas, the endogenous EPO levels of neurosensory retinas were essentially unchanged during the same period observed. In EPO-treated diabetic groups, EpoR expression remained at upregulated levels. Within 2 weeks of the onset of diabetes, activation of the ERK but not the STAT5 pathway was detected in the diabetic retina treated with EPO. CONCLUSIONS. These data demonstrate that apoptosis is an major contributor to neuronal cell death in the early course of diabetic retinopathy (DR). The upregulation of EpoR may be a compensatory response of retinal cells and tissue to diabetic stresses. The EPO/EpoR system as a maintenance-survival mechanism of retinal neurons responds to the insults of early diabetes other than ischemia. The protective function of EPO/EpoR at the least acts through the EpoR-mediated ERK pathway. Exogenous EPO administration by intravitreal injection in early diabetes may prevent retinal cell death and protect the BRB function. Therefore, this is a novel approach for treatment of early DR.

Comparison of macular thickness measurements between time domain and spectral domain optical coherence tomography.

Abstract: PURPOSE To compare macular thickness measurements obtained from time domain optical coherence tomography (OCT) and spectral domain OCT and to evaluate their repeatability and agreement. METHODS Thirty-five healthy normal subjects were included. In one randomly selected eye in each subject, three serial macular measurements were obtained from a time domain OCT (Stratus OCT, Carl Zeiss Meditec, Dublin, CA) and a spectral domain OCT (3D OCT; Topcon, Tokyo, Japan) by an experienced technician in random order. Total and regional macular thicknesses obtained by the two OCTs were compared. Their agreement was examined with Bland-Altman plots. Repeatability (2.77 x within subject SD [Sw]), coefficient of variation (CVw; Sw/overall mean), and intraclass correlation coefficient (ICC) were calculated to evaluate repeatability. RESULTS Low variability for macular thickness measurements was found in both time domain and spectral domain OCTs. The range of the respective CVw and ICC values were 1.6% to 3.2% and 0.85 to 0.91 for Stratus OCT and 0.6% to 2.4% and 0.92 to 0.99 for 3D OCT. 3D OCT demonstrated better repeatability for total and regional macular thicknesses (all with P

Light-Induced Decomposition of Indocyanine Green

Abstract: Purpose To investigate the light-induced decomposition of indocyanine green (ICG) and to test the cytotoxicity of light-induced ICG decomposition products. Methods ICG in solution was irradiated with laser light, solar light, or surgical endolight. The light-induced decomposition of ICG was analyzed by high-performance liquid chromatography (HPLC) and mass spectrometry. Porcine retinal pigment epithelial (RPE) cells were incubated with the light-induced decomposition products of ICG, and cell viability was measured by trypan blue exclusion assay. Results Independent of the light source used, singlet oxygen (photodynamic type 2 reaction) is generated by ICG leading to dioxetanes by [2+2]-cycloaddition of singlet oxygen. These dioxetanes thermally decompose into several carbonyl compounds. The decomposition products were identified by mass spectrometry. The decomposition of ICG was inhibited by adding sodium azide, a quencher of singlet oxygen. Incubation with ICG decomposition products significantly reduced the viability of RPE cells in contrast to control cells. Conclusions ICG is decomposed by light within a self-sensitized photo oxidation. The decomposition products reduce the viability of RPE cells in vitro. The toxic effects of decomposed ICG should be further investigated under in vivo conditions.

Report from the NEI/FDA Ophthalmic Clinical Trial Design and Endpoints Symposium.

Abstract: The National Eye Institute (NEI) of the National Institutes of Health (NIH) and the U.S. Food and Drug Administration (FDA) held an open symposium on November 28–29, 2006, in Washington, DC, where representatives from both federal agencies, the Center for Medicare and Medicaid Services (CMS), university scientists and clinicians, and others conferred on endpoints and clinical trial strategies for evaluating new treatments for age-related macular degeneration (AMD), diabetic retinopathy, and other retinal disorders. The symposium was organized as a forum for discussion by the Association for Research in Vision and Ophthalmology (ARVO). Cohosts of the symposium were Janice M. Soreth, MD, Director of the Division of Anti-infective and Ophthalmology Products of the FDA’s Center for Drug Evaluation and Research (CDER), and Karl G. Csaky, MD, PhD, NEI Senior Investigator. The symposium evolved from a series of meetings between members of the eye and vision research community and CDER. The research community is looking to the FDA for information that will help with obtaining timely approval of new products arising from ophthalmic clinical research. The National Alliance for Eye and Vision Research (NAEVR) was the host of the initial meetings. NAEVR is a nonprofit advocacy organization working on behalf of professional, consumer, and industry organizations involved in eye and vision research. Stephen J. Ryan, MD, NAEVR Board President and President of the Doheny Eye Institute of the University of Southern California, in his keynote speech to symposium attendees, noted that the need for more efficient and cost-effective clinical trials “. . . is especially important in ophthalmology where new technologies are enabling better quantitative measurements of outcomes, which subsequently expedites the translation of clinical trials into improved practice patterns.” The format of the NEI/FDA Ophthalmic Clinical Trial Design and Endpoints Symposium was a roundtable discussion moderated by Dr. Csaky. Presenters and discussants sat before an auditorium of approximately 250 observers. Audience members, mainly from industry, federal agencies, academia, and vision advocacy groups, had the opportunity to submit questions to the symposium organizers before the discussion. In essence, the symposium addressed developing standards for clinical trials in ophthalmology and focused largely on the type and duration of clinical trials or postsurveillance studies in addition to the clinical importance of the endpoints used in vision research clinical trials. The discussion consisted of five sections for which speakers and discussants addressed questions they had received in advance. The five sections were as follows:

Essential Roles of the PI3 Kinase/Akt Pathway in Regulating Nrf2-Dependent Antioxidant Functions in the RPE

Abstract: Purpose To investigate functional interactions between the phosphatidylinositol 3-kinase (PI3K)/Akt pathway and the nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent antioxidant system in cultured human retinal pigment epithelium (RPE) cells. Methods Cultured ARPE-19 cells were treated with different concentrations of PI3K inhibitors, followed by exposure to sulforaphane, an Nrf2 inducer. Akt phosphorylation was detected by Western blot analysis. Intracellular glutathione (GSH) content was measured by HPLC. Expression of genes downstream of Nrf2, including glutamate-cysteine ligase (GCL) and glutathione S-transferase, was measured by quantitative RT-PCR. Nrf2 activity was measured by a dual luciferase assay after transfection of a reporter plasmid containing the antioxidant response element (ARE). The small interference RNA approach was used to knock down Nrf2 in the RPE. Nrf2 localization was determined by subcellular fractionation and Western blot analyses. Results PI3K inhibitors wortmannin and LY294002 caused dose-dependent cellular and mitochondrial GSH depletion and downregulation of the modulatory subunit of GCL in cultured RPE cells. Both the basal and the induced Nrf2 activities were inhibited by wortmannin and LY294002. Overexpression of a constitutively active form of Akt potentiated Nrf2 activation, and the effect of Akt was blocked by siRNA that knocked down Nrf2. LY294002 also inhibited sulforaphane-induced Nrf2 nuclear translocation. Conclusions The PI3K/Akt pathway plays key roles in regulating Nrf2-ARE-dependent protection against oxidative stress in the RPE.

Visual impairment, causes of vision loss, and falls: the singapore malay eye study.

Abstract: PURPOSE: To report associations of visual impairment and the main causes of vision loss with falls in an older Asian population. METHODS: The population-based Singapore Malay Eye Study examined 3280 (78.7% response rate) Malay adults 40 to 80 years of age. Details about any fall in the previous 12 months and personal and sociodemographic information were collected. Presenting visual acuity (PVA) was measured. Mild or moderate visual impairment (0.3/= 1.0), and the primary causes of visual impairment were determined by ophthalmologists at examination. RESULTS: Of the 3280 participants, 3266 (99.6%) provided information about falls. Of these, 14.7% (n = 480) reported having fallen in the past 12 months. After adjustment for gender, age, body mass index, history of angina, heart attack, stroke, hypertension, diabetes, and self-rated health, the results showed that severe visual impairment in the worse eye significantly increased the risk of falling (60%; OR = 1.6; 95% CI 1.1 to 2.3). Severe visual impairment in one eye and mild or moderate visual impairment in the other also doubled the risk of falls (OR = 2.1; 95% CI 1.4-3.1). Having glaucoma (n = 21) increased the risk of falling by more than fourfold (OR = 4.2; 95% CI 1.2-12.3) after adjustment for visual acuity. Although mild or moderate visual impairment was not significantly associated with falls, odds ratios tended toward the direction of risk. CONCLUSIONS: Findings from this Asian population provide further evidence in support of the association between severe visual impairment and falls in older persons. Language: en

A viscoelastic biomechanical model of the cornea describing the effect of viscosity and elasticity on hysteresis.

Abstract: Purpose To develop a method for evaluating viscosity and elasticity of the cornea and to examine the effect that both properties have on hysteresis. Methods A three-component spring and dashpot model was created in Simulink in Matlab to represent the purely elastic and viscoelastic behavior of the cornea during a measurement using device called an ocular response analyzer (ORA). Values for elasticity and viscosity were varied while sinusoidal stress was applied to the model. The simulated stresses were used to determine how hysteresis is affected by the individual components of elasticity, viscosity, and maximum stress. To validate the model, high-speed photography was used to measure induced strain in a corneal phantom during ORA measurement. This measured strain was compared with the strains simulated by the model. Results When the spring in the viscoelastic portion of the model was stiffened, hysteresis decreased. When the spring in the purely elastic element was stiffened, hysteresis increased. If both springs were stiffened together, hysteresis peaked strongly as a function of the viscosity of the viscoelastic element. Below the peak value, lower elasticity was associated with higher hysteresis. Above the peak value, higher elasticity was associated with higher hysteresis. In addition, hysteresis increased as the air maximum pressure was increased. Measurements from phantom corresponded to predictions from the model. Conclusions A viscoelastic model is presented to illustrate how changing viscosity and elasticity may affect hysteresis. Low hysteresis can be associated with either high elasticity or low elasticity, depending on the viscosity, a finding consistent with clinical reports.

Evaluation of Time Domain and Spectral Domain Optical Coherence Tomography in the Measurement of Diabetic Macular Edema

Abstract: Optical coherence tomography (OCT) evaluation of macular structure is important in the diagnosis and monitoring of patients with diabetic macular edema (DME). Automated measurements of retinal thickness and volume in the macula from retinal tomograms using image-processing software have been useful in observing patients longitudinally in clinical practice and making comparisons in clinical trials. To date, time domain OCT using the Stratus OCT (Carl Zeiss Meditec, Inc., Dublin, CA) has been the most widely used technique for obtaining macular measurements1-5 and has been shown in previous studies to generate measurements with high repeatability.6-9 This instrument acquires images at a rate of 400 axial scans per second, with an axial resolution of 10 μm. Recently, a new class of OCT instruments employing spectral (Fourier) domain technology has been developed. One such instrument that is currently commercially available is the Cirrus HD-OCT (Carl Zeiss Meditec, Inc.). Although both time domain and spectral domain OCT use the same basic working principles, the scan rate of spectral domain OCT is at least 20,000 axial scans per second, with an improved axial resolution of 5 μm. Both the Stratus OCT and the Cirrus HD-OCT systems employ intrinsic software algorithms to calculate retinal thicknesses averaged across standardized subfields in the macula. Data for thickness calculations are collected from an array of A-scans distributed across the macula. This array differs between the Stratus OCT and Cirrus HD-OCT systems. In the Cirrus HD-OCT system, significantly more A-scans are used, and these are evenly distributed over the scanned area. The Stratus OCT employs fewer A-scans, and these are more heavily weighted toward the center of the scan. The Stratus OCT system corrects for this effect by extrapolating across areas not sampled; but, depending on the nature of the macular anatomy in those areas, quantitative differences in macular thickness computations may arise between the OCT systems. Also, although both OCT software systems perform automated delineations of retinal boundaries, each system uses different anatomic landmarks in the specification of the outer retinal boundary (information from the manufacturer, Carl Zeiss Meditec, Inc.). These differences may contribute to a different absolute value for thickness measurements, and may also affect how retinal boundaries are reliably delineated between the two OCT systems. For both clinical practice and in the conduct of clinical trials, the repeatability of OCT measurements has an important impact on how the data may be interpreted. The multiple differences between the time domain Stratus OCT and the spectral domain Cirrus HD-OCT systems may influence how repeatable macular measurements are in the context of DME where there may be considerable deviation from normal anatomy. Spectral domain OCT has been demonstrated to improve visualization of retinal structures in various macular diseases.10-12 The reliability of macular measurements obtained by using spectral domain OCT, however, has not been determined. Furthermore, a comparison of the reliability of these two OCT technologies in the determination of macular measurements has not been performed. The purpose of this study was to evaluate and compare the reliability of macular measurements obtained using time domain versus spectral domain OCT in patients with DME. The results of this study will provide information on how OCT macular measurements using both time domain and spectral domain technology can be compared and will further our understanding of how different modes of data capture and analysis in OCT systems can influence the measurements.

Role of NADPH oxidase in retinal vascular inflammation.

Abstract: PURPOSE In another study, it was demonstrated that NADPH oxidase-derived reactive oxygen species (ROS) are important for ischemia–induced increases in vascular endothelial growth factor (VEGF) and retinal neovascularization Diabetes–induced increases in retinal ROS, VEGF expression, and vascular permeability are accompanied by increases in the NADPH oxidase catalytic subunit NOX2 within the retinal vessels The goal of this study was to evaluate the potential role of NOX2 and NADPH oxidase activity in the development of retinal vascular inflammation METHODS Studies were performed in wild-type mice, mice lacking NOX2, and mice treated with the NADPH oxidase inhibitor apocynin in models of endotoxemia and streptozotocin-induced diabetes Intracellular adhesion molecule (ICAM)-1 expression was determined by Western blot analysis Leukocyte adhesion was assessed by labeling adherent leukocytes with concanavalin A Vascular permeability was assessed by extravasation of FITC-conjugated albumin ROS production was determined by dichlorofluorescein imaging RESULTS Both endotoxemia- and diabetes-induced increases in ICAM-1 expression and leukostasis were significantly inhibited by deletion of NOX2, indicating that this enzyme is critically involved in both conditions Moreover, apocynin treatment and deletion of NOX2 were equally effective in preventing diabetes-induced increases in ICAM-1, leukostasis, and breakdown of the blood–retinal barrier, suggesting that NOX2 is primarily responsible for these early signs of diabetic retinopathy CONCLUSIONS These data suggest that NOX2 activity has a primary role in retinal vascular inflammation during acute and chronic conditions associated with retinal vascular inflammatory reactions Targeting this enzyme could be a novel therapeutic strategy for treatment of the retinopathies associated with vascular inflammation (Invest Ophthalmol Vis Sci 2008; 49:3239 –3244) DOI:101167/iovs08-1755

Clinicopathologic Findings in Polypoidal Choroidal Vasculopathy

Abstract: Purpose To elucidate the pathogenic mechanism of polypoidal choroidal vasculopathy (PCV) based on histopathologic findings. Methods Specimens obtained by surgical excision of PCV from five eyes of five patients (mean age, 75.6 +/- 3.1 years) were studied histopathologically. Immunohistochemical studies were also performed to identify CD34, vascular endothelial growth factor (VEGF), CD68, alpha-smooth muscle actin (alpha-SMA) and hypoxia-inducible factor (HIF)-1alpha. Results Hyalinization of choroidal vessels and massive exudation of fibrin and blood plasma were observed in all the specimens of PCV lesions. Some blood vessels were located above the RPE in two of the five eyes. Immunohistochemically, CD68-positive cells were detected around the hyalinized vessels. There were no alpha-SMA-positive cells in the vessels of PCV. CD34 staining showed endothelial discontinuity. Vascular endothelial cells within the PCV specimens were negative for VEGF. HIF-1alpha positive inflammatory cells were located in the stroma of specimens. Conclusions Hyalinization of choroidal vessels, like arteriosclerosis, is characteristic of PCV.

Factors affecting perceptual thresholds in epiretinal prostheses.

Abstract: Purpose—The goal was to evaluate how perceptual thresholds are related to electrode impedance, electrode size, the distance of electrodes from the retinal surface, and retinal thickness in six subjects blind as a result of retinitis pigmentosa, who received epiretinal prostheses implanted monocularly as part of a U.S. Food and Drug Administration (FDA)–approved clinical trial. Methods—The implant consisted of an extraocular unit containing electronics for wireless data, power recovery, and generation of stimulus current, and an intraocular unit containing 16 platinum stimulating electrodes (260- or 520-μm diameter) arranged in a 4 × 4 pattern. The electrode array was held onto the retina by a small tack. Stimulation was controlled by a computer-based external system that allowed independent control over each electrode. Perceptual thresholds (the current necessary to see a percept on 79% of trials) and impedance were measured for each electrode on a biweekly basis. The distance of electrodes from the retinal surface and retinal thickness were measured by optical coherence tomography on a less regular basis. Results—Stimulation thresholds for detecting phosphenes correlated with the distance of the electrodes from the retinal surface, but not with electrode size, electrode impedance, or retinal thickness. Conclusions—Maintaining close proximity between the electrode array and the retinal surface is critical in developing a successful retinal implant. With the development of chronic electrode arrays that are stable and flush on the retinal surface, it is likely that the influence of other factors such as electrode size, retinal degeneration, and subject age will become more apparent.

The effect of amblyopia on fine motor skills in children.

Abstract: PURPOSE. In an investigation of the functional impact of amblyopia in children, the fine motor skills of amblyopes and agematched control subjects were compared. The influence of visual factors that might predict any decrement in fine motor skills was also explored. METHODS. Vision and fine motor skills were tested in a group of children (n 82; mean age, 8.2 1.7 [SD] years) with amblyopia of different causes (infantile esotropia, n 17; acquired strabismus, n 28; anisometropia, n 15; mixed, n 13; and deprivation n 9), and age-matched control children (n 37; age 8.3 1.3 years). Visual motor control (VMC) and upper limb speed and dexterity (ULSD) items of the Bruininks-Oseretsky Test of Motor Proficiency were assessed, and logMAR visual acuity (VA) and Randot stereopsis were measured. Multiple regression models were used to identify the visual determinants of fine motor skills performance. RESULTS. Amblyopes performed significantly poorer than control subjects on 9 of 16 fine motor skills subitems and for the overall age-standardized scores for both VMC and ULSD items (P 0.05). The effects were most evident on timed tasks. The etiology of amblyopia and level of binocular function significantly affected fine motor skill performance on both items; however, when examined in a multiple regression model that took into account the intercorrelation between visual characteristics, poorer fine motor skills performance was associated with strabismus (F1,75 5.428; P 0.022), but not with the level of binocular function, refractive error, or visual acuity in either eye. CONCLUSIONS. Fine motor skills were reduced in children with amblyopia, particularly those with strabismus, compared with control subjects. The deficits in motor performance were greatest on manual dexterity tasks requiring speed and accuracy. (Invest Ophthalmol Vis Sci. 2008;49:594‐603) DOI:10.1167/ iovs.07-0869

Computerized Analysis of Retinal Vessel Width and Tortuosity in Premature Infants

Abstract: PURPOSE: To determine, with novel software, the feasibility of measuring the tortuosity and width of retinal veins and arteries from digital retinal images of infants at risk of retinopathy of prematurity (ROP). METHODS: The Computer-Aided Image Analysis of the Retina (CAIAR) program was developed to enable semiautomatic detection of retinal vasculature and measurement of vessel tortuosity and width from digital images. CAIAR was tested for accuracy and reproducibility of tortuosity and width measurements by using computer-generated vessel-like lines of known frequency, amplitude, and width. CAIAR was then tested by using clinical digital retinal images for correlation of vessel tortuosity and width readings compared with expert ophthalmologist grading. RESULTS: When applied to 16 computer-generated sinusoidal vessels, the tortuosity measured by CAIAR correlated very well with the known values. Width measures also increased as expected. When the CAIAR readings were compared with five expert ophthalmologists' grading of 75 vessels on 10 retinal images, moderate correlation was found in 10 of the 14 tortuosity output calculations (Spearman rho = 0.618-0.673). Width was less well correlated (rho = 0.415). CONCLUSIONS: The measures of tortuosity and width in CAIAR were validated using sequential model vessel analysis. On comparison of CAIAR output with assessments made by expert ophthalmologists, CAIAR correlates moderately with tortuosity grades, but less well with width grades. CAIAR offers the opportunity to develop an automated image analysis system for detecting the vascular changes at the posterior pole, which are becoming increasingly important in diagnosing treatable ROP.

Myopia and the urban environment: findings in a sample of 12-year-old Australian school children.

Abstract: PURPOSE To examine associations between myopia and measures of urbanization in a population-based sample of 12-year-old Australian children. METHODS Questionnaire data on sociodemographic and environmental factors including ethnicity, parental education, and time spent in near work and outdoor activities were collected from 2367 children (75.0% response) and their parents. Population density data for the Sydney area were used to construct five urban regions. Myopia was defined as spherical equivalent refraction