Idebenone Prevents Oxidative Stress, Cell Death and Senescence of Retinal Pigment Epithelium Cells by Stabilizing BAX/Bcl-2 Ratio.

PURPOSE: Age-related macular degeneration (AMD) is one of the leading causes of blindness. Degeneration of the retinal pigment epithelium (RPE) is pathognomonic for the disease, and oxidative stress plays an important role in the pathogenesis of this disease. This study investigates potential antiapoptotic and cytoprotective effects of idebenone on cultured RPE cells (ARPE-19) under conditions of oxidative stress. METHODS: ARPE-19 cells were treated with 1-100 µM idebenone. Cell viability (MTT assay), induction of intracellular reactive oxygen species (ROS) and histone-associated DNA fragments in mono- and oligonucleosomes, expression of proapoptotic BAX and antiapoptotic Bcl-2 as well as senescence-associated ß-galactosidase (SA-ß-Gal) activity were investigated under exposure to hydrogen peroxide (H2O2). RESULTS: Idebenone concentrations from 1 to 20 µM showed no toxic effects on ARPE-19 cells. When cells were treated with H2O2, pretreatment with 5, 7.5, 10, and 20 µM idebenone led to a significant increase in the viability of ARPE-19 cells. In addition, idebenone pretreatment significantly attenuated the induction of SA-ß-Gal and intracellular ROS as well as the amount of histone-associated DNA fragments after treatment with H2O2. The reduction of proapoptotic BAX and the elevation of antiapoptotic Bcl-2 under idebenone show that this process is rather mediated by inhibiting H2O2-induced apoptosis, not necrosis. CONCLUSION: In this study, idebenone increased survival of ARPE-19 cells and reduced cell death, senescence, and oxidative stress by stabilizing the BAX/Bcl-2 ratio.

Megahertz ultra-wide-field swept-source retina optical coherence tomography compared to current existing imaging devices.

BACKGROUND: To investigate the image quality of wide-angle cross-sectional and reconstructed fundus images based on ultra-megahertz swept-source Fourier domain mode locking (FDML) OCT compared to current generation diagnostic devices. METHODS: A 1,050 nm swept-source FDML OCT system was constructed running at 1.68 MHz A-scan rate covering approximately 70° field of view. Twelve normal eyes were imaged with the device applying an isotropically dense sampling protocol (1,900 × 1,900 A-scans) with a fill factor of 100 %. Obtained OCT scan image quality was compared with two commercial OCT systems (Heidelberg Spectralis and Stratus OCT) of the same 12 eyes. Reconstructed en-face fundus images from the same FDML-OCT data set were compared to color fundus, infrared and ultra-wide-field scanning laser images (SLO). RESULTS: Comparison of cross-sectional scans showed a high overall image quality of the 15× averaged FDML images at 1.68 MHz [overall quality grading score: 8.42 ± 0.52, range 0 (bad)-10 (excellent)] comparable to current spectral-domain OCTs (overall quality grading score: 8.83 ± 0.39, p = 0.731). On FDML OCT, a dense 3D data set was obtained covering also the central and mid-peripheral retina. The reconstructed FDML OCT en-face fundus images had high image quality comparable to scanning laser ophthalmoscope (SLO) as judged from retinal structures such as vessels and optic disc. Overall grading score was 8.36 ± 0.51 for FDML OCT vs 8.27 ± 0.65 for SLO (p = 0.717). CONCLUSIONS: Ultra-wide-field megahertz 3D FDML OCT at 1.68 MHz is feasible, and provides cross-sectional image quality comparable to current spectral-domain OCT devices. In addition, reconstructed en-face visualization of fundus images result in a wide-field view with high image quality as compared to currently available fundus imaging devices. The improvement of >30× in imaging speed over commercial spectral-domain OCT technology enables high-density scan protocols leading to a data set for high quality cross-sectional and en-face images of the posterior segment.

Microaneurysm formation rate as a predictive marker for progression to clinically significant macular edema in nonproliferative diabetic retinopathy.

PURPOSE: To evaluate the predictive value of microaneurysm (MA) formation rate concerning the development of clinically significant macular edema (CSME) in patients with mild-to-moderate nonproliferative diabetic retinopathy as evaluated by an automated analysis of central field fundus 30° photographs. METHODS: Two hundred and eighty-seven eyes were included in the study. Photographs obtained at Day 0, at 6, and 12 months were analyzed using the RetmarkerDR software (Critical Health SA) in a masked manner, and the MA formation rate was documented. A threshold of a calculated MA formation rate of 2 or more was chosen to consider a patient "positive." The ability to predict CSME development was then calculated for a period of up to 5 years. HbA1c values, blood pressure, or duration of diabetes were also evaluated. RESULTS: The study population consisted of 89 male and 59 female patients with a mean age of 57.6 years, a mean HbA1c of 7.8, and a mean duration of diabetes of 12.3 years. Forty-seven of 287 eyes (16.4%) developed CSME during follow-up. An increased MA formation rate of >2 MA was clearly associated with development of CSME. Using the automated analysis and a threshold of 2 or more new MA, the authors were able to identify 70.2% of the eyes that developed CSME during follow-up ("true positive") and using a threshold of up to 2 new MA, 71.7% of the patients that did not develop CSME ("true negative"). No significant differences concerning baseline and 1-year HbA1c levels within patient eyes that developed CSME compared with patient eyes below or over the calculated threshold of 2 MA (P = 0.554 and P = 0.890, respectively) were seen. The positive and negative predictive value was calculated to be 33% versus 92.5%, sensitivity was 70%, and specificity was 72%. CONCLUSION: Using the RetmarkerDR software, the authors were able to identify patients with higher risk to develop CSME during follow-up using a threshold of 2 or more MA formation rate. Together with the high negative predictive value, the automated analysis may help to determine the individual risk of a patient to develop sight-threatening complications related to diabetic retinopathy and schedule individual screening intervals.

Microaneurysm turnover in diabetic retinopathy assessed by automated RetmarkerDR image analysis--potential role as biomarker of response to ranibizumab treatment.

PURPOSE: To evaluate the influence of a ranibizumab treatment on microaneurysm (MA) turnover in diabetic retinopathy. METHODS: Sixty-nine eyes were included in this retrospective study. We compared a group of 33 eyes with ranibizumab treatment for diabetic macular edema to 36 eyes with nonproliferative diabetic retinopathy only. Nonmydriatic ultra-widefield scanning laser ophthalmoscopy (Optomap) images were obtained at a mean 4.76 ± 1.69 days prior to the first ranibizumab injection (baseline) and again 35.94 ± 2.44 days after the third consecutive injection in a 4-week interval. In untreated controls, images were obtained at baseline and 97.81 ± 3.16 days thereafter. Images were analyzed using the RetmarkerDR software (Critical Health SA, Coimbra, Portugal), and the turnover of MAs was documented and analyzed. Thereafter, MA turnover was correlated with central retinal thickness (CRT) as assessed by OCT. RESULTS: At baseline, patients in the treatment group had 5.64 ± 0.75 MAs. One month after 3 ranibizumab injections, measured MAs decreased to 4.03 ± 0.66. In the untreated control group, the initial number of 3.36 ± 0.6 MAs remained almost unchanged over 3-4 months (2.89 ± 0.57 MAs). Dynamic analysis showed that after ranibizumab treatment 3.06 ± 0.5 new MAs appeared, while 5.09 ± 0.79 disappeared. In the control group, 2.11 ± 0.4 new MAs appeared and 2.61 ± 0.48 disappeared. MA turnover was significantly higher with ranibizumab compared to the control group (8.15 ± 1.14 vs. 4.72 ± 0.81, p < 0.001). Consistently, CRT decreased from 444 to 330 µm in the ranibizumab group, while there was no change in the control group (291 vs. 288 µm). CONCLUSION: The treatment of macular edema using ranibizumab does not only reduce macular thickness, but also has an impact on the turnover of MAs in diabetic retinopathy. RetmarkerDR analysis showed that more pre-existent MAs disappeared than new MAs developed, and the absolute number of MAs also decreased.

Nonmydriatic ultra-wide-field scanning laser ophthalmoscopy (Optomap) versus two-field fundus photography in diabetic retinopathy.

The purpose of this study was to investigate the diagnostic properties of a 2-laser wavelength nonmydriatic 200° ultra-wide-field scanning laser ophthalmoscope (SLO) versus mydriatic 2-field 45° color fundus photography (EURODIAB standard) for assessing diabetic retinopathy (DR). A total of 143 consecutive eyes of patients with different levels of DR were graded regarding DR level and macular edema based on 2-field color photographs or 1 Optomap Panoramic 200 SLO image. All SLO images were nonmydriatic and all photographs mydriatic. Grading was performed masked to patient and clinical data. Based on photography, 20 eyes had no DR, 44 had mild, 18 moderate and 42 severe nonproliferative DR, and 19 eyes had proliferative DR. Overall correlation for grading DR level compared to Optomap SLO was moderate with kappa 0.54 (p < 0.001), fair-to-moderate in macular edema grading with kappa 0.39 (p < 0.001), and substantial for grading clinically significant macular edema (kappa 0.77). The wide-field SLO offers a wider field of view and can potentially better differentiate lesions by applying the 2 laser wavelengths. However, these advantages over 2-field fundus photography need to be confirmed in further studies.

Time required for navigated macular laser photocoagulation treatment with the Navilas.

BACKGROUND: Navilas laser is a novel technology combining photocoagulation with imaging, including fluorescein angiographic (FA) images which are annotated and aligned to a live fundus view. We determine the time necessary for planning and treatment of macular edema utilizing the Navilas. METHODS: The screen recordings during treatments were retrospectively analyzed for treatment type, number of laser shots, the duration of planning (measured from the time the planning image was selected to time of marking the last planned treatment spot), and total time of laser application. RESULTS: A total of 93 treatments (30 grid, 30 focal and 33 combined treatments) by four physicians from three sites were included. An average of 125 spots were applied to each eye. The total time spent for each focal treatment - including the planning was 7 min 47 s (±3 min and 32 s). CONCLUSIONS: Navilas is a novel device providing a time efficient platform for evaluating FA images and performing threshold macular laser photocoagulation.

Pars plana vitrectomy and internal limiting membrane peeling in epimacular membranes: correlation of function and morphology across the macula.

PURPOSE: To analyze the correlation between morphological and functional results 12 months after epiretinal membrane (ERM) surgery. METHODS: 31 eyes from 31 consecutive patients with metamorphopsia and best corrected visual acuity (BCVA) below 20/32 underwent a transconjunctival 23- gauge vitrectomy with ERM and internal limiting membrane peeling. Preoperatively and 3, 6 and 12 months postoperatively, we assessed BCVA, microperimetry (MP-1) and spectral domain optical coherence tomography (SD-OCT). Photoreceptor inner and outer segment (IS/OS) was graded on SD-OCT images and correlated with microperimetry measurements in the fovea and parafoveal region. RESULTS: The postoperative BCVA was significantly better in eyes with an intact IS/OS junction (p < 0.01). In addition, the mean defect depth was postoperatively decreased in the foveal and parafoveal area in eyes with an intact IS/OS junction. A correlation of SD-OCT IS/OS images and microperimetry in eyes with improvement in BCVA of at least 2 lines revealed a statistically significant result for the parafoveal quadrants (p < 0.011 for SD-OCT and p < 0.005 for microperimetry) but not for the foveal area alone. CONCLUSIONS: The IS/OS regeneration in the parafoveal quadrants contributes significantly to the recovery of BCVA following ERM surgery. Consequently, functional and morphological tests of the macular area should not be limited to the fovea but should be extended to the parafoveal region.

Axitinib modulates hypoxia-induced blood-retina barrier permeability and expression of growth factors.

This study investigates the effects of the multikinase inhibitor axitinib on the expression of vascular endothelial growth factor (VEGF) receptors 1/2 (VEGFR-1/2) and platelet-derived growth factor (PDGF) receptor beta (PDGFR-ß), hypoxia-induced increased tissue permeability, occludin, zonula occludens protein 1 (ZO-1), VEGF-A, and PDGF expression of human retinal pigment epithelial (RPE) cells and human umbilical vein endothelial cells (HUVECs). Primary human RPE cells and HUVECs were exposed to hypoxia and axitinib. Viability of cells, tissue permeability, and expression of occludin, ZO-1, VEGF, PDGF, VEGFR-1/2 and PDGFR-ß, and their mRNAs, were investigated by reverse transcription-polymerase chain reaction, enzyme-linked immunosorbent assay, western blotting, and immunohistochemistry. Treatment with axitinib reduced expression of VEGFR-1/2 and PDGFR-ß. Hypoxia decreased cell viability, occludin, and ZO-1 expression and increased tissue permeability, expression, and secretion of VEGF and PDGF. Axitinib significantly reduced hypoxia-induced effects on HUVEC and RPE cells. Our in vitro results suggest that axitinib may have promising properties as a potential treatment for diabetic macular edema.

A randomized placebo-controlled trial of idebenone in Leber's hereditary optic neuropathy.

Major advances in understanding the pathogenesis of inherited metabolic disease caused by mitochondrial DNA mutations have yet to translate into treatments of proven efficacy. Leber's hereditary optic neuropathy is the most common mitochondrial DNA disorder causing irreversible blindness in young adult life. Anecdotal reports support the use of idebenone in Leber's hereditary optic neuropathy, but this has not been evaluated in a randomized controlled trial. We conducted a 24-week multi-centre double-blind, randomized, placebo-controlled trial in 85 patients with Leber's hereditary optic neuropathy due to m.3460G>A, m.11778G>A, and m.14484T>C or mitochondrial DNA mutations. The active drug was idebenone 900 mg/day. The primary end-point was the best recovery in visual acuity. The main secondary end-point was the change in best visual acuity. Other secondary end-points were changes in visual acuity of the best eye at baseline and changes in visual acuity for both eyes in each patient. Colour-contrast sensitivity and retinal nerve fibre layer thickness were measured in subgroups. Idebenone was safe and well tolerated. The primary end-point did not reach statistical significance in the intention to treat population. However, post hoc interaction analysis showed a different response to idebenone in patients with discordant visual acuities at baseline; in these patients, all secondary end-points were significantly different between the idebenone and placebo groups. This first randomized controlled trial in the mitochondrial disorder, Leber's hereditary optic neuropathy, provides evidence that patients with discordant visual acuities are the most likely to benefit from idebenone treatment, which is safe and well tolerated.

Synthesis, staining properties, and biocompatibility of a new cyanine dye for ILM peeling.

BACKGROUND: The aim of this work is to investigate the biocompatibility and staining properties of DSS: 3,3'-Di-(4-sulfobutyl)-1,1,1',1'-tetramethyl-di-1H-benz[e]indocarbocyanine (DSS). METHODS: Dye concentrations of 0.5, 0.25, and 0.1% were evaluated (290 and 295 mOsm). Toxicity was assessed using a colorimetric test measuring the inhibition of ARPE 19 cell, human primary RPE cell, and human Müller cell proliferation. Exposure time was 30, 60, 120, and 300 s. Indocyanine green (ICG) (0.5, 0.25, and 0.1%) served as a control. Cells were also illuminated with plain white light (750 mW/cm(2)) for 10 min to assess phototoxic effects. Besides staining of porcine and human lens capsule, internal limiting membrane (ILM)-staining was assessed by applying 0.25 and 0.5% DSS over the macula in two human post-mortem eyes. RESULTS: DSS of 0.25 and 0.1% showed no toxic effect on primary RPE cells and MIO-M1cells, and 0.5, 0.25, and 0.1% for ARPE-19 cells. In MIO-M1cells, 0.5% dye showed a significant reduction of mitochondrial dehydrogenase activity only following an exposure time of 300 s. Following illumination, ICG showed a significantly more pronounced effect on cell viability in primary RPE cells and MIO-M1cells compared to DSS. The absorption maximum is found at 591 nm; the even more bathochromic fluorescence proceeds with a common Stokes' shift where maxima at 620 and 660 nm with a quantum yield of 32% were found. The fluorescence is sufficiently hypsochromic and the fluorescence quantum yield high enough for an easy visual detection. The contrast and staining properties at the ILM were excellent and allowed for a controlled removal of the ILM during surgery. No penetration into deeper retinal layers was noted. CONCLUSIONS: Our results indicate that this new cyanine dye DSS may represent an alternative for ILM staining due to its matched absorption concerning visibility and fluorescence qualities as well as its good biocompatibility.

Inhibitory activity of ranibizumab, sorafenib, and pazopanib on light-induced overexpression of platelet-derived growth factor and vascular endothelial growth factor A and the vascular endothelial growth factor A receptors 1 and 2 and neuropilin 1 and 2.

BACKGROUND: Cumulative light exposure is significantly associated with progression of age-related macular degeneration. Growth factors and growth factor receptor signaling are known to have a substantial impact on the development of age-related macular degeneration. This study explored the effects of ranibizumab, sorafenib, and pazopanib on vascular endothelial growth factor A (VEGF) receptors 1 and 2 and neuropilin 1 and 2 expression in human retinal pigment epithelial cells. In addition, their effects on light-induced overexpression of VEGF and platelet-derived growth factor were investigated. METHODS: Primary human retinal pigment epithelial cells were exposed to white light and then treated with ranibizumab (0.125 mg/mL), sorafenib (1 µg/mL), or pazopanib (1 µg/mL). Viability of cells, expression of VEGF receptors 1 and 2 and neuropilin 1 and 2 and their mRNA, and secretion of VEGF and platelet-derived growth factor were investigated by reverse transcription-polymerase chain reactions, immunohistochemistry, and enzyme-linked immunosorbent assays. RESULTS: Treatment with sorafenib or pazopanib reduced the expression of VEGF receptors 1 and 2 and neuropilin 1, and sorafenib also reduced neuropilin 2. Light exposure decreased cell viability and increased expression and secretion of VEGF and platelet-derived growth factor. Sorafenib and pazopanib significantly reduced light-induced overexpression and secretion of VEGF and platelet-derived growth factor. Ranibizumab reduced secreted VEGF in cell culture supernatants only. CONCLUSION: Our in vitro results suggest that multikinase inhibitors have promising properties as a potential antiangiogenic treatment for age-related macular degeneration.

Filtering blue light reduces light-induced oxidative stress, senescence and accumulation of extracellular matrix proteins in human retinal pigment epithelium cells.

BACKGROUND: Cumulative light exposure is significantly associated with ageing and the progression of age-related macular degeneration. To prevent the retina from blue-light damage in pseudophakia, blue light-absorbing intraocular lenses have been developed. This study compares the possible protective effects of a blue light-absorbing intraocular lens to an untinted ultraviolet-absorbing intraocular lens with regard to light-induced oxidative stress and senescence of human retinal pigment epithelium. METHODS: As primary human retinal pigment epithelium cells were exposed to white light, either an ultraviolet- and blue light-absorbing intraocular lens or ultraviolet-absorbing intraocular lens was placed in the light beam. After 60 min of irradiation, cells were investigated by electron microscopy for viability, induction of intracellular reactive oxygen species, and senescence-associated ß-galactosidase activity. Expression and secretion of matrix metalloproteinases 1 and 3 and their mRNA were determined by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay. RESULTS: Light exposure induced structural damage, decreased retinal pigment epithelium cell viability, and increased reactive oxygen species, senescence-associated ß-galactosidase activity and matrix metalloproteinases 1 and 3 expression and secretion. Although both types of intraocular lens significantly reduced these effects, the protective effects of the ultraviolet- and blue light-absorbing intraocular lens were significantly stronger than those of the ultraviolet-absorbing intraocular lens. CONCLUSIONS: The ultraviolet- and blue light-absorbing intraocular lens demonstrated significantly better protection against light-induced oxidative stress, senescence and structural damage than the ultraviolet-absorbing intraocular lens. These in vitro findings support the hypothesis that the ultraviolet- and blue light-absorbing intraocular lens may prevent retinal damage in clinical use.

Submandibular neck lifting with platysma resection.

A novel method for cervicofacial rejuvenation by submandibular neck lifting with platysma resection is presented. Between 2003 and 2009 we performed submandibular neck lifting in 18 cases. This technique is an isolated but efficient procedure for lifting of the neck. An additional advantage might be the possibility of doing the procedure with tumescence local anesthesia. In all cases, satisfactory aesthetic results were achieved.

The NEI VFQ-25 vision-related quality of life and prevalence of eye disease in a working population.

PURPOSE: To apply the National Eye Institute 25-item Visual Function Questionnaire (VFQ-25) to a large group of working people and assess the prevalence of eye disease. METHODS: In this cross-sectional study, 619 employees of two large enterprises in Munich, Germany, underwent complete ophthalmological examination and assessment of the vision-related quality of life by the Visual Function Questionnaire (VFQ-25, German translation). Statistical analysis including binary logistic regression to investigate predictive factors of eye disease was performed. RESULTS: The age of the participants was mean 42 years. In 108 patients (17.4%) an ophthalmological disease was present. The mean VFQ-25 composite score was 91.1, and was not significantly different between the group with (88.8) or without (91.6) eye disease. A clear age dependency of VFQ-25 scores was seen. Most reduced subscales in eye disease were ocular pain and role difficulties. Patients with amblyopia did not show any reduction in visual quality of life (composite score 93.4). In the binary regression model, considering the clinical parameters intraocular pressure (IOP), cup/disc ratio (CDR), and spherical equivalent, as well as some of the VFQ-25 subscales, the presence of eye disease could be predicted with a high specificity of 98.7% but a low sensitivity of 12.2%. CONCLUSIONS: Normal values of the VFQ-25 performance in a large number of working people in Germany are given with a clear age dependency. A combination of refraction, IOP, CDR and some VFQ subscales could reasonably well exclude any prevalent eye disease. For screening purposes, neither variable nor combination was sufficiently sensitive and specific.

Inhibition of human retinal pigment epithelial cell attachment, spreading, and migration by the human lectin galectin-1.

PURPOSE: Adhesion and migration of dislocated retinal pigment epithelial (RPE) cells are initial steps in the pathogenesis of proliferative vitreoretinopathy (PVR). The role of the endogenous lectin, galectin-1, in attachment, spreading, and migration of human RPE cells was investigated from a therapeutic perspective. METHODS: Human RPE cells were treated with galectin-1 concentrations that ranged 0-250 microg/ml. Cell viability was tested by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazoliumbromide (MTT) assay. Galectin-1 binding to the RPE cells was investigated by immunocytochemistry. Attachment of RPE cells was assessed on 96-well plates coated with laminin, or fibronectin, or galectin-1, or the glycoprotein-lectin combinations and subsequent MTT-testing. RPE migration in the absence or presence of galectin-1 on the respective substrates was tested using a modified Boyden chamber assay with platelet derived growth factor (PDGF)-BB as the chemoattractant. Cellular spreading was characterized by cytoplasmic halo formation of RPE cells after three hours in contact with the surface coating. RESULTS: Galectin-1 bound to the cell surface. Binding could be inhibited by a beta-galactoside. MTT assays revealed no toxicity within control limits for the concentration range tested. When added to the medium, galectin-1 dose-dependently inhibited RPE cell attachment, spreading, and migration by more than 70%, irrespective of the substratum tested. When coated onto the plastic surface, galectin-1 alone impaired spreading and migration of RPE cells, and reduced attachment to and migration on fibronectin by up to 80%. CONCLUSIONS: Galectin-1 inhibits RPE cell attachment, migration, and spreading in vitro with no apparent cytotoxicity. This activity of the endogenous effector deserves consideration as a potential therapeutic agent for the prevention of PVR.

Intravitreal voriconazole: in vitro safety-profile for fungal endophthalmitis.

INTRODUCTION: Fungal endophthalmitis is a rare but sight-threatening disease. Despite an expanding range of fungal pathogens, there are only few therapeutic agents for its treatment available. Voriconazole is a second-generation synthetic triazole with a broad action against yeasts and molds. The current study investigates the safety of Voriconazole for intravitreal application in a cell culture model. METHODS: Primary human retinal pigment epithelium cells (RPE) and primary human optic nerve head astrocytes were treated with concentrations of Voriconazole ranging from 25 microg/mL to 10 mg/mL. Possible toxic effects and IC50 were evaluated after 24 hours and under conditions of oxidative stress. By treating the RPE cell lines with tumor-necrosisfactor alpha (TNF-alpha), lipopolysaccharides (LPS), and interleukin-6 (IL-6) the effects of Voriconazole on cellular viability under conditions of inflammation were investigated. Toxicity was evaluated by colorimetric measuring the inhibition of RPE cell proliferation (MTT). Additionally cell viability was quantified by a microscopic live-dead-assay. RESULTS: Concentrations <250 microg/mL Voriconazole had no influence neither on RPE nor on optic nerve head astrocytes cell proliferation and cell viability when administered for 24 hours and under oxidative stress. When preincubated with tumor-necrosis-factor alpha, lipopolysaccharides and interleukin-6 for 24 hours and subsequently treated with Voriconazole at concentrations up to 250 microg/mL for 24 hours no significant decrease in proliferation and viability was observed. CONCLUSIONS: This study showed that no significant toxicity existed for Voriconazole in vitro on primary RPE and optic nerve head astrocytes when administered in therapeutic concentrations up to 250 microg/mL.

In vitro safety of intravitreal moxifloxacin for endophthalmitis treatment.

PURPOSE: To investigate the safety of moxifloxacin for intravitreal application in a cell culture model. SETTING: Department of Ophthalmology, Ludwig-Maximilians-Universität, Munich, Germany. METHODS: Primary human retinal pigment epithelium (RPE) cells, ARPE19 cells, and primary optic nerve head astrocyte (ONHA) cells were treated with concentrations of moxifloxacin ranging from 10 to 750 microg/mL. Possible toxic effects and median inhibitory concentration were evaluated after 24 hours as well as under conditions of oxidative stress. After treating the RPE and ONHA cell lines with tumor necrosis factor-alpha (TNF-alpha; 10 microg/mL), lipopolysaccharides (LPS; 20 microg/mL), and interleukin-6 (IL-6; 20 microg/mL), the effects of moxifloxacin on cellular viability under conditions of inflammation were investigated. Toxicity was evaluated by measuring the inhibition of RPE cell proliferation with the tetrazolium dye-reduction assay (MTT; 3-[4,5-dimethylthiazol- 2-yl]-2,5-diphenyl tetrazolium bromide). Cell viability was quantified by a microscopic live/dead assay. RESULTS: At concentrations higher than 150 microg/mL, moxifloxacin had adverse effects on primary RPE, ARPE19, and ONHA cell proliferation and viability. Lower concentrations did not affect RPE or ONHA cell proliferation and viability when administered for 24 hours. No significant decrease in proliferation and viability was observed after preincubation with TNF-alpha, LPS, and IL-6 for 24 hours and subsequent treatment with moxifloxacin concentrations of 10 to 150 microg/mL for 24 hours. Hydrogen peroxide exposure did not increase cellular toxicity. CONCLUSIONS: No significant toxicity of moxifloxacin was seen on primary RPE cells, ARPE19 cells, or ONHA cells at concentrations up to 150 microg/mL. Intravitreal use of moxifloxacin up to this concentration may be safe and effective for the treatment of endophthalmitis.