Ravi K. Murthy

Normative Data for Macular Thickness by High-Definition Spectral-Domain Optical Coherence Tomography (Spectralis)

PURPOSE To establish normative data for the macular thickness by spectral-domain optical coherence tomography (SD-OCT) in subjects with no known retinal disease. DESIGN Prospective, observational study in an academic institutional setting. METHODS Fifty subjects (age range, 20 to 84 years) with no known retinal disease, best-corrected visual acuity 20/20, and normal intraocular pressure were enrolled. The subjects were divided into 3 age groups: group 1 included subjects 20 to 40 years of age; group 2 included subjects 41 to 60 years of age; and group 3 included subjects 61 years of age and older. All subjects underwent a complete ophthalmologic examination to rule out any retinal diseases or glaucoma. All the OCT scans were performed by a single operator, and data obtained from the right eyes were analyzed by default, unless the right eye did not meet the inclusion criteria, and then data from left eyes were analyzed (n = 4). Central point thickness (CPT) and retinal thickness (RT) in 9 Early Treatment Diabetic Retinopathy Study (ETDRS) subfields, including central subfield (CSF), were analyzed. Statistical analyses were carried out using the analysis of variance. RESULTS Overall, the mean CPT was 227.3 +/- 23.2 microm, and mean CSF was 270.2 +/- 22.5 microm. Among the ETDRS subfields, the outer nasal quadrant had the maximum thickness (mean +/- standard deviation, 339.5 +/- 16.9 microm). The RT did not show significant difference with age (P = .62) or with gender (P = .1). However, there was a suggestion of significant difference in RT of Black subjects as compared with White subjects (P = .007) in the present study. CONCLUSIONS Normative values for macular thickness in otherwise healthy eyes were measured to be 227.3 microm (CPT) and 270.2 microm (CSF) using commercially available Spectralis SD-OCT. Based on the data, the present study proposes the guidelines for normal CSF thickness to be 315 microm for future studies using macular thickness measurements with Spectralis SD-OCT (Heidelberg Engineering, Vista, California, USA).

Comparison of Retinal Thickness in Normal Eyes Using Stratus and Spectralis Optical Coherence Tomography

PURPOSE Spectral-domain optical coherence tomography (SD-OCT) is an advancement over time-domain OCT (TD-OCT) in the imaging of retinal disorders. Retinal thickness measured by SD-OCT differs from that measured by TD-OCT because the delineation of the outer boundary of the retina differs in the two instruments. The present study aims to evaluate this difference by comparing macular thickness, as obtained by Stratus and Spectralis OCT, in subjects without any known retinal disease. METHODS Thirty-six subjects with no history of retinal disease and with normal vision and normal intraocular pressure were enrolled in the study. Both Stratus and Spectralis OCT scanning were performed by the same operator on all subjects in one eye. Central point thickness (CPT) and retinal thickness in nine ETDRS subfields, including central subfield (CSF), were measured. Students t-test was used to determine statistical significance. RESULTS Mean CPT, as measured by the Stratus and Spectralis OCT, was 166.9 +/- 20.9 microm and 225.1 +/- 17.1 microm (P < 0.0001), and mean CSF was 202.3 +/- 19.6 microm and 271.4 +/- 19.6 microm (P < 0.0001), respectively. Although the mean difference in CSF thickness was 69.1 microm, it ranged from 61.9 to 74 microm in the other eight ETDRS subfields. CONCLUSIONS An increased measurement in retinal thickness of approximately 65 to 70 microm, as measured by Spectralis OCT compared with Stratus OCT, is consistent with the extent of axial retinal thickness measured by the two instruments. This increased measurement corresponds to the inclusion of the outer segment-RPE-Bruchs membrane complex by Spectralis OCT, which is relevant to studies using the newer SD-OCT for assessment of retinal thickness.

Temporal Macular Thinning on Spectral-Domain Optical Coherence Tomography in Proliferative Sickle Cell Retinopathy

Comment. Propionibacterium acnes is a slow-growing aerotolerant anaerobic gram-positive pathogen. Classically, it is associated with a chronic endophthalmitis, in which patients present with symptoms of anterior uveitis and a characteristic plaque is seen on the posterior lens capsule. In contrast, all 3 eyes described herein were completely quiet, without any clinical evidence of inflammation. We propose that the pronounced inflammatory reaction seen in chronic endophthalmitis secondary to P acnes is attributable to access of the organism to the anterior chamber, allowing the eye to react with an inflammatory response. In late CBS, there is an absence of inflammation, as the P acnes in the turbid fluid is sequestered within the capsular bag owing to the tight seal of the anterior capsule to the IOL. Interestingly, when this type of case is treated with an Nd:YAG posterior capsulotomy, the milky fluid can be visualized tracking into the vitreous cavity, and we have not yet observed inflammation or endophthalmitis. However, in 1988, Carlson and Koch did report a case of P acnes endophthalmitis after an Nd: YAG laser capsulotomy. Whether their patient had late CBS is unknown. In our experience with other such cases that were treated with an Nd:YAG laser, it seems that this turbid fluid, with its debris, is cleared quite effectively from the vitreous cavity without consequence. A possible explanation for this lack of inflammation is that the bacterial count is low, with a prolonged doubling time of P acnes, or perhaps the oxygenated vitreous serves as a poor culture medium for this anaerobic species. Alternatively, perhaps the anterior segment inflammation seen in chronic endophthalmitis is attributable to an immunologic phenomenon associated with P acnes infection. These cases bring into question the role of P acnes as a pathogen in late CBS and whether antibiotic therapy, topical or intravitreal, is actually indicated. In 1980, before the introduction of the Nd:YAG laser, Lindstrom and Harris introduced a technique for creating a posterior capsulotomy by inserting a needle through the pars plana to open the capsule. In our series, this technique was adapted for sampling the milky fluid and injecting an antibiotic into the capsular bag. It has the advantage of leaving the anterior chamber and the IOL undisturbed and can be easily performed at the slit lamp. Because intravitreal injections for macular degeneration have become commonplace, the technique can be performed safely, with minimal discomfort to the patient. On rare occasion, there may be a role for the application of this technique in other conditions. Late CBS typically presents with blurred vision months after uncomplicated phacoemulsification cataract surgery with an IOL. On examination, there is distention of the posterior lens capsule, which contains a milky fluid with particulate debris. We have shown that this fluid may contain P acnes; however, in our experience, it has not been associated with inflammation or an infectious process. Therefore, this condition may be treated with Nd:YAG capsulotomy, but caution is warranted, as there may be a slight risk for endophthalmitis.

Foveal structure defined by spectral domain optical coherence tomography correlates with visual function after macular hole surgery

Purpose. To study the correlation between final visual acuity after successful anatomic macular hole repair and features on spectral domain optical coherence tomography (SD-OCT). Methods. Retrospective review of charts of patients who underwent macular hole surgery. Data collection included pre- and postoperative best-corrected visual acuity (BCVA), central subfield foveal thickness (CSFT), and presence or absence of inner segment-outer segment (IS-OS) line changes on SD-OCT. Data collected from SD-OCT were correlated with Snellen BCVA, which was converted to logMAR score. Subjects were divided into 2 groups: group I had improvement in BCVA of 2 lines or more and group II improved less than 2 lines or had worsening of BCVA. Results. A total of 35 eyes of 32 patients had successful anatomic closure, which was documented both clinically and on SD-OCT. Mean age of the patients was 74.1 years and 71.2% (23/32) of patients were female. Overall, the mean BCVA changed from 1.01±0.38 preoperatively to 0.89±0.48 postoperatively (p=0.33). Based on the postoperative visual outcome, 16 eyes belonged to group I and 19 eyes belonged to group II. On the SD-OCT, the mean CSFT was 252.7±69.1 μm. No correlation was found between the mean CSFT and BCVA in either group. All the 16 patients in group I had a continuous IS-OS line on SD-OCT at the fovea in contrast to 26.3 % (5/19) of patients in group II (p=0.03). Conclusions. Establishment of continuity of IS-OS line is an important indicator of visual recovery in eyes with successful anatomic closure of macular hole.

Evaluation of cytotoxic effects of bevacizumab on human corneal cells.

Purpose: Corneal neovascularization contributes to corneal opacification in inflammatory conditions of the cornea and severely compromises the success of corneal transplantation. Vascular endothelial growth factor (VEGF) plays an important role in stimulating and maintaining corneal neovascularization. Anti-VEGF therapy, especially the use of anti-VEGF antibody bevacizumab, has gained popularity in the management of retinal neovascularization and is being used topically for corneal neovascularization. The aim of this study was to investigate the safety profile of bevacizumab on human corneal cell lines. Methods: Human corneal epithelial and fibroblast cell lines and an umbilical vascular endothelial cell line were treated with increasing doses of bevacizumab. The effect of this treatment on cell viability was assessed by WST-1 and crystal violet staining assays. Cytotoxicity was also assessed by fluorescent microscopy and flow cytometric evaluation of propidium iodide-stained cells. Results: In the cytotoxicity experiments, there was no difference in cell numbers after 24-hour exposure compared with control in any of the cell lines at the concentrations tested (P > 0.05 to 0.98). Conclusion: Bevacizumab was nontoxic to human corneal epithelial and fibroblast cells at 3 different concentrations.

Evaluation of ultraviolet light toxicity on cultured retinal pigment epithelial and retinal ganglion cells

Purpose: Our study is aimed at evaluating the role of UVB light in inducing cytotoxicity in an in vitro model. Methods: RGC-5 and ARPE-19 cells were exposed to different time periods of UVB light: 0, 15, 30, and 45 min. They were subsequently examined for changes in cell morphology, cell viability (neutral red uptake assay), generation of reactive oxygen species (ROS), expression of bax, bcl-2 and cytochome C by reverse transcriptase polymerase chain reaction and western blot, respectively. Results: Dose-dependent reduction in cell viability to UVB light was demonstrated with parallel increase in ROS. Increased duration of exposure (>15 minutes), was associated with increased expression of bax and cytochrome C, and absence of bcl-2 expression. Conclusion: UVB light exposure results in cell cytotoxicity. The concomitant generation of ROS and expression of apoptotic markers suggests the role of oxidative stress in UVB-mediated apoptosis in an in vitro model of retinal ganglion and pigment epithelial cells.

Evaluation of Differential Toxicity of Varying Doses of Bevacizumab on Retinal Ganglion Cells, Retinal Pigment Epithelial Cells, and Vascular Endothelial Growth Factor–Enriched Choroidal Endothelial Cells

PURPOSE To evaluate in vitro the effects of bevacizumab, an anti-vascular endothelial growth factor (VEGF) antibody, on retinal pigment epithelial cells (RPE) and retinal ganglion cells (RGC), at doses that were inhibitory to VEGF-enriched choroidal endothelial cells (CEC). METHODS Monkey CEC (RF6A), human RPE cells (ARPE-19), and rat RGC (RGC-5) were exposed for 24 h to increasing doses of bevacizumab. Cell numbers were quantified with WST-1 assay. Cell death was assessed using propidium iodide (PI) staining via flow cytometry and fluorescent microscopy. RESULTS Bevacizumab was inhibitory to RF6A at 2.0 mg/mL (P < 0.005). No effect on cell viability was noted on ARPE-19 and RGC-5 cell lines at this particular dose of bevacizumab. These results were supported by fluorescent microscopy of PI-stained cells. CONCLUSIONS VEGF-stimulated proliferation of CEC was inhibited by bevacizumab. Bevacizumab was not cytotoxic to human RPE and rat RGC in vitro at a dose that is inhibitory to monkey CEC.

Lutein protects retinal pigment epithelium from cytotoxic oxidative stress

Abstract Context: Lutein (LUT) and zeaxanthin (ZEA) are currently under investigation in clinical trials as prophylactic nutritional agents for age-related macular degeneration (AMD). However, dose used in these trials is empirical and not been investigated in in vitro studies. Objective: In this study, we investigated the dose–response effect of LUT and ZEA in protecting retinal pigment epithelium (RPE) from oxidative stress, a common underlying pathology in AMD. Methods: Three thousand cultured human retinal pigment epithelial cells (ARPE-19) were plated in 72-well plate and after 24 h were exposed to increasing concentrations of hydrogen peroxide (H2O2). ARPE-19 cells were exposed to four different concentrations of LUT (0.5, 1, 2 and 4 µg/mL) and ZEA (0.1, 0.2, 0.4 and 0.8 µg/mL). After 24 h incubation, cells were subjected to oxidative stress induced with H2O2. Cultures containing saline solution and dichloromethane served as controls. Cell viability was assessed using the WST-1 assay. Pathophysiological pathways were evaluated by measuring caspase-3 levels as an indicator of apoptosis induction. Reactive oxygen species (ROS) levels were measured using dihydrorhodamine-123. Results: Cell viability as a percentage of control was 81.3%, 81.1%, and 88.8% at 0.5, 1, and 2 µg/ml, respectively of LUT (p < 0.001). The maximum cytoprotective effect was seen with LUT at 2 μg/mL. ZEA did not show any cytoprotective effect at all concentrations used in the study. Caspase-3 showed a corresponding decrease in levels with LUT (1 and 2 µg/ml). Significant decrease in ROS levels were measured only with LUT at 4 µg/ml (p = 0.02). Discussion and conclusions: Results from our study provide in vitro data to support the epidemiologic studies, which are currently underway to provide evidence that lutein may act as cofactor that modulates processes implicated in AMD pathogenesis.

Sequential spectral domain OCT documentation of retinal changes after branch retinal artery occlusion

Branch retinal artery occlusions (BRAO) are characterized histopathologically by inner retinal edema initially and atrophy in the presence of persistent ischemia. The duration of ischemia leading to irreversible atrophic retinal changes is not clear. Spectral-domain optical coherence tomography (SD-OCT) provides non-invasive detailed in-vivo histological changes in the retina. In this case report, we show sequential in vivo pathological changes seen in the inner retinal layers, in spite of clinical improvement, following the migration of an intraretinal embolus on the optic nerve head, which had previously resulted in symptomatic BRAO.

Aqueous Interleukin-6 Levels Are Superior to Vascular Endothelial Growth Factor in Predicting Therapeutic Response to Bevacizumab in Age-Related Macular Degeneration

Objective. To prospectively evaluate the effect of intravitreal bevacizumab on aqueous levels of interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) in patients with exudative age-related macular degeneration (AMD) and correlate clinical outcomes with cytokine levels. Methods. 30 eyes of 30 patients with exudative AMD underwent intravitreal injection of bevacizumab three times at monthly intervals. The aqueous samples prior to the 1st injection (baseline) and 3rd injection were analyzed for VEGF and IL-6 levels. Subjects were subgrouped based upon change in the central subfield (CSF) macular thickness on SD-OCT at 8 weeks. Group 1 included patients (n = 14) with a decrease in CSF thickness greater than 10% from the baseline (improved group). Group 2 included patients (n = 16) who had a decrease in CSF thickness 10% or less (treatment-resistant). Results. In subgroup analysis, in both groups 1 and 2 patients, compared to aqueous VEGF, aqueous IL-6 levels showed a better correlation with CSF thickness on SD-OCT (r = 0.72 and 0.71, resp.). Conclusions. Aqueous IL-6 may be an important marker of treatment response or resistance in wet macular degeneration. Future therapeutic strategies may include targeted treatment against both VEGF and IL-6, in patients who do not respond to anti-VEGF treatment alone.