Lutein protects retinal pigment epithelium from cytotoxic oxidative stress.

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.

Recent Advances in Imaging Macular Atrophy for Late-Stage Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) is a leading cause of blindness worldwide. In late-stage AMD, geographic atrophy (GA) of dry AMD or choroidal neovascularization (CNV) of neovascular AMD eventually results in macular atrophy (MA), leading to significant visual loss. Despite the development of innovative therapies, there are currently no established effective treatments for MA. As a result, early detection of MA is critical in identifying later central macular involvement throughout time. Accurate and early diagnosis is achieved through a combination of clinical examination and imaging techniques. Our review of the literature depicts advances in retinal imaging to identify biomarkers of progression and risk factors for late AMD. Imaging methods like fundus photography; dye-based angiography; fundus autofluorescence (FAF); near-infrared reflectance (NIR); optical coherence tomography (OCT); and optical coherence tomography angiography (OCTA) can be used to detect and monitor the progression of retinal atrophy. These evolving diverse imaging modalities optimize detection of pathologic anatomy and measurement of visual function; they may also contribute to the understanding of underlying mechanistic pathways, particularly the underlying MA changes in late AMD.

Comparative in vitro safety analysis of dyes for chromovitrectomy: indocyanine green, brilliant blue green, bromophenol blue, and infracyanine green.

PURPOSE: Vital dyes such as infracyanine green (IfCG), brilliant blue green (BBG), and bromophenol blue (BPB) have been used as an alternative to indocyanine green (ICG) during chromovitrectomy. We compared the in vitro toxicity of IfCG, BBG, and BPB with ICG on the retinal pigment epithelial cells and retinal ganglion cells at various concentrations to optimize the safe dose and duration of exposure. METHODS: Cultured retinal ganglion cells (RGC-5) and human retinal pigment epithelial cells (ARPE-19) were exposed to 2 concentrations (0.25 and 0.5 mg/mL) of ICG, IfCG, BBG, and BPB at various time intervals (1, 5, 15, and 30 minutes). Cell viability was quantified with neutral red assay, and mode of cell death was evaluated with flow cytometry-based Annexin V and propidium iodide staining. RESULTS: Exposure to ICG resulted in 48%-74% reduction in neutral red uptake in both RGC-5 and ARPE-19 cells, after an exposure time of ≥5 minutes compared with control (P < 0.001). Infracyanine green, BBG, and BPB were significantly less toxic on the 2 cell lines at exposure times <15 minutes. (Reduction in cell viability ranged from 6.9% ± 3.3% to 29.3% ± 7.4% when compared with control, P > 0.5.) However, among the newer dyes, BBG caused necrosis in retinal pigment epithelial cells and retinal ganglion cells as the exposure time period increased beyond 5 minutes. CONCLUSION: Newer vital dyes, IfCG, BBG, and BPB, are significantly less toxic on retinal ganglion cells and retinal pigment epithelial cells' cell lines when compared with ICG. Infracyanine green was least toxic among the three newer dyes studied.

Evaluation of choroidal endothelial cell proliferation after exposure to varying doses of proton beam radiation.

PURPOSE: Focal epiretinal radiation has emerged as a promising tool in the management of choroidal neovascularization associated with age-related macular degeneration. However, the dosages tested are not backed by cell culture studies used in the clinical setting empirically. METHODS: Choroidal endothelial cells (RF6A) were maintained in a log scale and exposed to a single fraction of 2, 4, 8, and 12 cobalt gray-equivalent of proton radiation with an internal control. Cell viability was quantified using Vi-cell XR and neutral red assay at days 5, 9, and 12 after radiation. Mitochondrial viability using WST-1 and reactive oxygen species levels using dihydrorhodamine 123 were measured at similar intervals. RESULTS: By using neutral red assay, on day 12, the percentages of viable cells compared with control were 100.1 ± 5.7%, 96.7 ± 23.3%, 27.6 ± 6.6%, and 19.5 ± 3% at radiation doses of 2, 4, 8, and 12 cobalt gray-equivalent, respectively (P < 0.001). Increase in reactive oxygen species levels correlated with the number of dead cells implicating reactive oxygen species as an intermediary molecule (r = 0.85-0.96). CONCLUSION: Our study shows sensitivity of cultured choroidal endothelial cells to proton beam radiation at doses of 8 and 12 cobalt gray-equivalent in an in vitro model.

Incomplete Vogt-Koyanagi-Harada in a 14-year-Old African American female with bilateral disc edema.

PURPOSE: To report an unusual case of incomplete Vogt-Koyanagi-Harada (VKH) in a 14-Year-Old African American female. OBSERVATIONS: Here we present a 14-Year-Old African American Female with incomplete VKH who presented to the emergency department with a one-month history of malaise, fever, bilateral decreased vision and temporal headaches. At the time of presentation, she was found to have bilateral anterior uveitis and disc edema. The brain and orbit MRI were unremarkable, CSF analysis showed lymphocytic pleocytosis with negative cultures and gram stain. An extensive lab workup was unrevealing. CONCLUSION: This is an unusual case of incomplete VKH in a young African American female presenting with anterior uveitis and disc edema without serous retinal detachments. Initiation of high dose oral prednisone promptly after diagnosis resulted in rapid improvement of symptoms and improvement in visual acuity with the resolution of the disc edema. This case highlights the importance of considering VKH in the differential diagnosis of disc edema associated with uveitis, even in the absence of serous retinal detachment.

Bevacizumab neutralizes the protective effect of vascular endothelial growth factor on retinal ganglion cells.

PURPOSE: Vascular endothelial growth factor (VEGF) is well known for its role in pathologic neovascularization, including wet age-related macular degeneration. However, a growing body of evidence indicates that VEGF is also neuroprotective of non-vascular cells in various animal models through reduction of oxidative stress. In light of the widespread use of intraocular anti-VEGF therapies for age-related macular degeneration (AMD), we evaluated the impact of anti-VEGF agents on the neuroprotective effect of VEGF on retinal ganglion cells. METHODS: Staurosporine differentiated retinal ganglion cells were treated with increasing doses of VEGF in the presence of hydrogen peroxide. After optimization, an increasing concentration of bevacizumab was added to neutralize VEGF-mediated protection. The degree of oxidative damage was measured at various time points using buthionine sulfoxime (BSO), a glutathione reductase inhibitor. Cell viability was assessed using WST-1 and Crystal violet assays. RESULTS: VEGF (200 ng/ml) protected differentiated retinal ganglion cells (RGC)-5 against H(2)0(2)-mediated oxidative stress. This effect was eliminated by co-treatment with bevacizumab (2.0 mg/ml), which by itself was not cytotoxic. CONCLUSIONS: These results indicate an important role for VEGF in the maintenance of retinal ganglion cells.

In-vivo high resolution imaging of optic nerve head drusen using spectral-domain Optical Coherence Tomography.

BACKGROUND: Optic nerve head drusen (ONHD) are white calcareous deposits, seen either superficially on the optic nerve head or buried within it. Diagnosis of ONHD is made by one or more ways: clinical exam, autofluorescence, ultrasound of the optic nerve, CT scan and/or visual field examination. The present study describes features of ONHD based on another diagnostic modality, the spectral-domain OCT (Spectralis). METHODS: This is a retrospective case series of 5 patients with bilateral ONHD with a best-corrected visual acuity of 20/20 and no other posterior segment pathology. All the patients underwent fundus photography, fundus autofluorescence, B-scan ultrasonography, Spectralis OCT and Humphrey 30-2 threshold visual fields. RESULTS: All 5 patients had surface ONHD which were autofluorescent and echodense on B-scan ultrasonography. Spectralis OCT findings in the corresponding areas include 'scattered spots with high reflectivity' casting a shadow underneath. The reflectivity can be distinctly differentiated from the blood vessels on the optic nerve. Two patients had an arcuate scotoma on the Humphrey visual fields. No correlation was found between the changes on Spectralis OCT with that of visual field. CONCLUSIONS: Spectralis OCT is another useful ancillary investigation in the diagnosis of ONHD and we describe the features in the present study.

Fundus autofluorescence and spectral domain optical coherence tomography in early detection of Plaquenil maculopathy.

PURPOSE: To report fundus autofluorescence (FAF) and spectral-domain optical coherence tomography (SD-OCT) findings in a patient with early hydroxychloroquine maculopathy. METHODS: A 50-year-old man presented with complaints of ring-like shadows in front of his eyes. He had been on hydroxychloroquine (Plaquenil) 400 mg orally twice daily for 10 years (5.14 mg/kg/day). Fundus examination revealed granular hypopigmentation in an arcuate pattern in the inferior parafoveal area bilaterally. The patient underwent further evaluation with SD-OCT and FAF imaging. Functional changes were assessed by microperimetry. RESULTS: Hypopigmented lesions on the fundus were autofluorescent and SD-OCT corresponding to this area revealed loss of inner segment-outer segment, suggesting loss of photoreceptor layer in the affected area with underling dysfunctional retinal pigment epithelium. CONCLUSIONS: Findings on SD-OCT and FAF imaging support the histopathologic findings described in cases of hydroxychloroquine maculopathy.

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 microm. 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.

Bilateral optic nerve and retinal infiltration as an initial site of relapse in a child with T-cell acute lymphoblastic leukemia.

PURPOSE: To describe a case of leukemic infiltration of bilateral optic nerves and retina as a site of relapse in a child with T-Cell Acute Lymphoblastic Leukemia (ALL). OBSERVATIONS: We report a 7 year old female who presented one year following initial treatment for T-Cell ALL with visual acuity impairment, bilateral optic nerve infiltration and infiltration of the retina of both eyes. OCT demonstrated subretinal fluid in both eyes, which eventually resolved, and perivascular hyperreflectivity within the inner retinal layers. She was treated with systemic and intrathecal chemotherapy, total body and orbital radiation and eventual bone marrow transplantation with notable improvement in vision and regression of retinal and optic nerve findings. With continued remission, there was notable outer retinal thinning, specifically of the photoreceptors in the right eye. CONCLUSIONS AND IMPORTANCE: Leukemic abnormalities of the eye are not uncommon, however optic nerve and retinal infiltration are rare manifestations. Leukemic infiltrates of the retina can be detected by OCT despite normal funduscopic examination and monitored for improvement. The optic nerve and other ocular tissues are considered a pharmacologic sanctuary and thus, the optic nerve can be a site of relapse in leukemia. The use of radiation therapy is a helpful adjunct with systemic, intrathecal chemotherapy and stem cell transplantation in obtaining clinical remission and visual acuity improvement.

Evaluation of modified portable digital camera for screening of diabetic retinopathy.

AIMS: To describe a portable wide-field noncontact digital camera for posterior segment photography. METHODS: The digital camera has a compound lens consisting of two optical elements (a 90-dpt and a 20-dpt lens) attached to a 7.2-megapixel camera. White-light-emitting diodes are used to illuminate the fundus and reduce source reflection. The camera settings are set to candlelight mode, the optic zoom standardized to x2.4 and the focus is manually set to 3.0 m. RESULTS: The new technique provides quality wide-angle digital images of the retina (60 degrees ) in patients with dilated pupils, at a fraction of the cost of established digital fundus photography. CONCLUSIONS: The modified digital camera is a useful alternative technique to acquire fundus images and provides a tool for screening posterior segment conditions, including diabetic retinopathy in a variety of clinical settings.

Sinonasal lymphoma masquerading as persistent blepharoconjunctivitis.

Sinonasal lymphoma, a rare nonepithelial malignant tumor often originates from the nose and paranasal sinuses. Typically, these lymphomas present with symptoms of nasal obstruction, discharge, epistaxis and facial swelling. Orbital extension results in ocular symptoms like proptosis, blurred vision and diplopia. In this case report, we describe persistent blepharoconjunctivitis, an unusual ocular presentation of sinonasal lymphoma. The tumor was positively identified with immunoreactivity of biopsy specimen; Appropriate chemotherapy lead to successful remission of the disease.

Direct image-creating aspheric lens for indirect ophthalmoscopy.

AIMS: To describe a compact handheld optical lens that creates a real, upright image of the fundus with indirect ophthalmoscopy. METHODS: The compound lens consists of three double aspheric lenses of variable dioptric power mounted coaxially in a plastic housing mount. Lens A and B are separated by a distance of 30.96 mm while lens B and C are 19.38 mm apart. The length of the optical system is 79.63 mm. RESULTS: The lens design provides an upright aerial image of the fundus between the lens and the observer compared to the real inverted image formed by conventional indirect lenses during indirect ophthalmoscopy. CONCLUSIONS: The direct image-creating aspheric lens provides an upright image of the fundus with binocular indirect ophthalmoscopy, thus eliminating the need for mental reorientation of an inverted image.

Optical coherence tomography findings of bilateral foveal leukemic infiltration.

We report a case of a 59-year-old man with a history of atypical chronic myelogenous leukemia who presented with a several-week history of decreased vision in both eyes. His clinical examination revealed bilateral foveal infiltration, which was also demonstrated on optical coherence tomography. After a failed induction with imatinib (Gleevec(®)), he was treated with omacetaxine (Synribo(®)) with an appropriate hematologic response. As his leukemia improved with chemotherapy, his retinal lesions regressed as demonstrated by serial optical coherence tomography and fundus photographs, with near complete restoration of foveal architecture.

Infrared imaging enhances retinal crystals in Bietti's crystalline dystrophy.

Infrared imaging dramatically increased the number of crystalline deposits visualized compared with clinical examination, standard color fundus photography, and red free imaging in patients with Bietti's crystalline dystrophy. We believe that this imaging modality significantly improves the sensitivity with which these lesions are detected, facilitating earlier diagnosis and may potentially serve as a prognostic indicator when examined over time.

Prevalence and factors predictive of intraocular fungal infection in patients with fungemia at an academic urban tertiary care center.

OBJECTIVE: To report the prevalence and to identify factors predictive of intraocular infection in patients with fungemia receiving prophylactic antifungal therapy. METHODS: A retrospective review of patients who received prophylactic antifungal therapy and a dilated fundus examination at an academic urban tertiary care center from 2000 to 2007. Basic demographic information, fungal species grown, antifungal agent(s) used, number of positive blood culture specimens, visual acuity, visual symptoms, and known risks of disseminated candidiasis were noted. Logistic regression analysis was used to determine the factors significantly associated with intraocular fungal infection. RESULTS: A total of 132 patients with positive fungemia culture were requested to have ophthalmology consults. The prevalence of ocular infection was 6.9% (N=9). All nine patients were infected with Candida species. Undergoing gastrointestinal (GI) surgery within the prior 6 months was significantly related to developing intraocular infection, with an odds ratio of 18.5 (95% confidence interval, 15.1-24.3; P=0.002). Having ≥3 positive fungal blood cultures was also a significant risk factor, with an odds ratio of 2.6 (95% confidence interval, 1.8-3.7; P=0.03). Among 40 patients having GI surgery, eight (20.0%) had intraocular fungal disease, compared with one of 92 patients (1.1%) not having GI surgery. Among 125 patients with a negative baseline examination result, two of 32 patients (6.3%), who had recent GI surgery, subsequently developed fungal ocular disease, compared with 0 of 93 patients (0%), who did not have recent GI surgery. CONCLUSION: Recent GI surgery and higher numbers of positive fungal blood culture specimens may be predictive of candida ocular infections. Normal baseline fundoscopy examination results in patients with such risks may require repeat evaluations to detect delayed manifestations.

Human ciliary epithelium as a source of synthesis and secretion of vascular endothelial growth factor in neovascular glaucoma.

IMPORTANCE: Retinal ischemia-induced upregulation of vascular endothelial growth factor (VEGF) leads to endothelial proliferation of the anterior segment, resulting in neovascular glaucoma. OBJECTIVE: To investigate the ciliary epithelium as a possible source of VEGF in human eyes enucleated for intractable neovascular glaucoma. DESIGN, SETTING, AND PARTICIPANTS: In this proof-of-concept, laboratory-based study, 16 human enucleated eyes (8 with neovascular glaucoma and 8 as controls) were investigated. MAIN OUTCOMES AND MEASURES: Presence of VEGF by immunohistochemical analysis (VEGF protein) and in situ hybridization (VEGF messenger RNA). RESULTS: In eyes with neovascular glaucoma, strong VEGF immunoreaction in the nonpigmented epithelial cells of the ciliary processes and in the retina was noted. In situ hybridization for VEGF messenger RNA revealed a similar pattern, with positive stain results only in eyes with neovascular glaucoma. A minimal amount of VEGF immunostaining was seen in control eyes. CONCLUSIONS AND RELEVANCE: The nonpigmented ciliary epithelium is an important site of VEGF synthesis in patients with neovascular glaucoma. The ciliary epithelium may represent an additional focus of treatment in the management of neovascular glaucoma, especially in eyes that are nonresponsive to panretinal photocoagulation.

Infrared imaging and spectral-domain optical coherence tomography findings correlate with microperimetry in acute macular neuroretinopathy: a case report.

INTRODUCTION: Spectral-domain optical coherence tomography findings in a patient with acute macular neuroretinopathy, and correlation with functional defects on microperimetry, are presented. CASE PRESENTATION: A 25-year old Caucasian woman presented with bitemporal field defects following an upper respiratory tract infection. Her visual acuity was 20/20 in both eyes and a dilated fundus examination revealed bilateral hyperpigmentary changes in the papillomacular bundle. Our patient underwent further evaluation with spectral-domain optical coherence tomography, infrared and fundus autofluorescence imaging. Functional changes were assessed by microperimetry. Infrared imaging showed the classic wedge-shaped defects and spectral-domain optical coherence tomography exhibited changes at the inner segment-outer segment junction, with a thickened outer plexiform layer overlying these areas. Fluorescein and indocyanine green angiography did not demonstrate any perfusion defects or any other abnormality. Microperimetry demonstrated focal elevation in threshold correlating with the wedge-shaped defects in both eyes. CONCLUSION: Spectral-domain optical coherence tomography findings provide new evidence of the involvement of the outer plexiform layer of the retina in acute macular neuroretinopathy.