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Microsurgeons require the ability to make precise and stable maneuvers in order to achieve surgical objectives and to minimize surgical risks during freehand microsurgical procedures. This work presents a novel common path swept source optical coherence tomography based smart surgical tool that suppresses hand tremor. It allows enhanced tool tip stabilization, more accurate targeting and may lower surgical risk. Here the one dimensional motion tremor of a surgeon's hand is assessed by the surgical instrument. The ability to accurately locate a surgical target and the ability to maintain tool tip offset distances in a chicken embryo model are significantly improved as compared to freehand use.

Purpose: To assess modifiable environmental risk factors and protective factors for age-related macular degeneration (AMD) in a native Japanese population., Design: A case-control study., Participants: We included 422 case-control samples composed of 279 consecutive AMD cases and 143 controls., Methods: Information regarding systemic conditions and lifestyle were documented in each subject by standardized questionnaire including age, gender, smoking history, body mass index (BMI), and history of cardiovascular disease, hypertension, and diabetes. Serum fatty acids profiles were analyzed by gas chromatography performed on blood samples taken from each study participant. Logistic regression and multiple comparison analyses were utilized in this study., Main Outcome Measures: Population-specific information assessing systemic conditions, lifestyle, and serum fatty acid profiles., Results: Among environmental factors analyzed cigarette smoking showed the most significant association with development of all AMD (P<0.00001; odds ratio [OR], 4.06; 95% confidence interval [CI], 2.22-7.43), typical neovascular AMD (P<0.0001, OR, 4.59; 95% CI, 2.29-9.18), and polypoidal choroidal vasculopathy (P<0.001; OR, 4.87; 95% CI, 1.96-12.1). Hypertension and BMI showed a mild association with AMD. Although male prevalence was significantly higher in all case groups than in controls with conventional Scheffe correction, there was no association of gender with AMD development when logistic regression analysis was used to adjust for cigarette smoking. There was no difference in fatty acid profiles, except for a mild association of eicosapentaenoic acid concentration in the all AMD group., Conclusions: In the Japanese population studied, cigarette smoking influenced the risk of AMD but fractionated serum fatty acid levels did not. Although prior reports indicate a male predominance in Japanese patients with AMD, this study demonstrates that cigarette smoking accounts for this confounding bias. In addition, our population-specific data do not demonstrate significant differences in serum fatty acid composition, including ω-3 and ω-6 long chain polyunsaturated fatty acids, in Japanese patients with and without AMD. These results are consistent with the high proportion of smokers in aged Japanese men and the high fish oil intake in this population., Financial Disclosure(s): The authors have no proprietary or commercial interest in any of the materials discussed in this article., (Copyright © 2011 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: To determine whether there is an association between complement factor H (CFH), high-temperature requirement A-1 (HTRA1), vascular endothelial growth factor (VEGF), and pigment epithelium-derived factor (PEDF) genotypes and response to treatment with photodynamic therapy (PDT) for age-related macular degeneration (AMD) in a Japanese population., Design: Prospective, case-control study., Participants: One hundred ten patients with exudative AMD treated by verteporfin PDT were recruited prospectively at the Department of Ophthalmology, Saitama Medical University Hospital, Saitama, Japan., Methods: The patients were genotyped for 4 single nucleotide polymorphisms (SNPs; rs800292, rs1061170, rs1410996, rs2274700) in the CFH gene, a rs11200638-SNP in the HTRA1 gene, 3 SNPs (rs699947, rs1570360, rs2010963) in the VEGF gene, and 4 SNPs (rs12150053, rs12948385, rs9913583, rs1136287) in the PEDF gene using a TaqMan assay., Main Outcome Measures: The treatment outcomes and genotypes of CFH, HTRA1, VEGF, and PEDF polymorphisms., Results: Best-corrected visual acuity 1 year after PDT was significantly increased in patients with the HTRA1-rs11200638 GG genotype as compared with patients with the GA or AA genotypes (P = 2.9 × 10⁻², 7.0 × 10⁻⁴, respectively). The rate of recurrence in the 12-month period after PDT was also associated with HTRA1-rs11200638 genotype (P = 3.12 × 10⁻²). Patients with the AA genotype of HTRA1-rs11200638 had an approximately 6-fold greater risk of the recurrence than patients with the GG genotype (P = 5.58 × 10⁻³). Significant differences were demonstrated in the mean time interval from the initial treatment to the time of recurrence for the genotypes of CFH-rs1410996/-rs2274700 (P = 8.50 × 10⁻³)., Conclusions: The HTRA1-rs11200638 and CFH-rs1410996/-rs2274700 variants were associated with response to PDT in this study population. These variants may be used for genetic biomarkers to estimate visual outcomes and recurrences in the response to PDT with significant predictive power., (Copyright © 2011. Published by Elsevier Inc.)

Purpose: To present outcomes of combined pars plana vitrectomy, silicone oil (SO) injection, and permanent keratoprosthesis (Kpro) procedure in prephthisical eyes., Methods: All 3 patients were monocular with chronic severe hypotony, aphakia, and total corneal opacity in their vital eye. Preoperative visual acuity ranged from light perception to counting fingers at 1 foot. Two patients had a history of failed corneal grafts because of SO, and one had funnel retinal detachment. Pars plana vitrectomy and long-term SO tamponade were performed, and a permanent Boston type 1 Kpro was used in lieu of a donor corneal transplantation., Results: No unexpected intraoperative complications were encountered. Patients were followed for a period of 11-13 months. All patients had anatomic success with an attached retina and a clear visual axis. The procedures resulted in increased visual function in all patients ranging from hand motions to 20/800. No case has progressed to phthisis bulbi during the follow-up period. At the last visit, biomicroscopic examination revealed clear Kpro with an attached retina., Conclusions: Boston type 1 Kpro implantation, as the primary corneal procedure with pars plana vitrectomy and intraocular SO, may be a viable option in selected patients with prephthisical eyes.

Objectives: To evaluate the redox state and the spatial distribution of mitochondria in malignant human brain cancer cells grown on different substrates., Methods: Cellular autofluorescence images were obtained through an inverted fluorescence microscope and the redox fluorometric ratio was evaluated (after the subtraction of background) as the net fluorescence signal through the DAPI filter divided by the net fluorescence signal through the FITC filter. Spatial mitochondria distribution patterns were evaluated by division of the cell area at the midpoint between the nuclear and cell membranes. The average fluorescence in the central area (CF) was divided by the average fluorescence from the peripheral area (PF). The CF/PF ratios were compared between cells cultured on either poly-D-lysine or collagen I substrates., Results: Glioblastoma cells seeded on the collagen-coated plates were observed to proliferate approximately 33-50% faster than the cells seeded on the poly-D-lysine-coated plates. Consistent with the proliferation findings, the redox ratios were lower for the cells seeded on the collagen-coated plates compared with poly-D-lysine. However, cell size and the percentage of cells with perinuclear mitochondrial distribution were not observed to be different in the cells seeded on the two surfaces., Conclusions: Redox ratio computation by using redox fluorometry is a useful predictor of cellular proliferation.

We determined whether there is an association between complement factor H (CFH), high-temperature requirement A-1 (HTRA1), vascular endothelial growth factor (VEGF), and pigment epithelium-derived factor (PEDF) genotypes and the response to treatment with a single intravitreous injection of bevacizumab for age-related macular degeneration (AMD). Eighty-three patients with exudative AMD treated by bevacizumab injection were genotyped for three single nucleotide polymorphisms (SNPs; rs800292, rs1061170, rs1410996) in the CFH gene, a rs11200638-SNP in the HTRA1 gene, three SNPs (rs699947, rs1570360, rs2010963) in the VEGF gene, and four SNPs (rs12150053, rs12948385, rs9913583, rs1136287) in the PEDF gene using a TaqMan assay. The CT genotype (heterozygous) of CFH-rs1061170 was more frequently represented in nonresponders in vision than TT genotypes (nonrisk allele homozygous) at the time points of 1 and 3 months, while there was no CC genotype (risk allele homozygous) in our study cohort (p = 7.66 × 10(-3), 7.83 × 10(-3), respectively). VEGF-rs699947 was also associated with vision changes at 1 month and PEDF-rs1136287 at 3 months (p = 5.11 × 10(-3), 2.05 × 10(-2), respectively). These variants may be utilized for genetic biomarkers to estimate visual outcomes in the response to intravitreal bevacizumab treatment for AMD.

Purpose: To describe phenotype and genotype characteristics of age-related macular degeneration (AMD) in Japanese patients., Design: A case-control study., Participants: A total of 550 case-control samples composed of 408 consecutive AMD cases and 142 controls., Methods: Clinical information assessing age, gender, affected eyes, fundus features, and fluorescein/indocyanine green angiograms were systematically evaluated. Four single nucleotide polymorphisms (SNPs; rs800292, rs1061170, rs1410996, rs2274700) in the complement factor H (CFH) gene, 1 SNP (rs11200638) in the high-temperature requirement factor A1 (HTRA1) gene, 3 SNPs (rs699947, rs1570360, rs2010963) in the vascular endothelial growth factor (VEGF) gene, and 4 SNPs (rs12150053, rs12948385, rs9913583, rs1136287) in the pigment epithelium-derived factor (PEDF) gene were assessed using TaqMan technology., Main Outcome Measures: The clinical phenotype information and genotypes of CFH, HTRA1, VEGF, and PEDF polymorphisms., Results: Of Japanese patients with neovascular AMD (nAMD), 219 (58.7%) had typical nAMD and 154 (41.3%) had polypoidal choroidal vasculopathy (PCV). The frequency of bilateral exudative involvement was similar between typical nAMD (15.5%) and PCV (13.6%) (P = 0.613). Significant soft drusen were observed in the fellow eyes of 88 (47.6%) of 185 patients with unilateral typical nAMD and in 25 (18.8%) of 133 patients with unilateral PCV (P = 1.24x10(-7)). A serous pigment epithelium detachment was seen in 55 (25.1%) of 219 patients with typical nAMD and in 64 (41.6%) of 154 patients with PCV. A significant association was noted in CFH-rs800292, CFH-rs1410996, CFH-rs2274700, and HTRA1-rs11200638 with AMD development (P = 2.36x10(-5), 7.18x10(-5), 7.18x10(-5), 2.70x10(-7), respectively; population attributable risk = 57.3%, 57.8%, 57.8%, and 58.9%, respectively). We estimated the highest-risk group to have an approximately 70-fold greater risk of nAMD compared with the lowest-risk group when analyzing a combination of 4 SNPs in the CFH and HTRA1 genes., Conclusions: The Japanese AMD phenotype is characterized by a higher frequency of PCV, male predominance, and lower frequency of bilateral presentation compared with Caucasian AMD. Genotype analyses demonstrate a significant population attributable risk for SNPs in the CFH and HTRA1 genes and demonstrate joint effects for both genes. Gene variants in both CFH and HTRA1 contribute significantly to the AMD phenotype in a Japanese population., (Copyright 2010 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: To evaluate the role of indoleamine dioxygenase (IDO) in human retinal pigment epithelial cell (RPE)-mediated immune modulation., Methods: The immunosuppression of cultured human RPEs (ARPE-19 cells) was assayed using the mixed lymphocyte reaction (MLR) harvested from CBA/J and BALB/c mice. The expression of critical immune modulatory molecules, such as class II major histocompatibility complex (MHC) molecules, co-stimulatory molecules (B7-1 and B7-2), and indoleamine dioxygenase (IDO), was examined by reverse transcriptase polymerase chain reaction, flow cytometry, and immunofluorescence staining with and without IFN-gamma stimulation., Results: RPEs expressed IDO and class II MHC molecules under IFN-gamma stimulation. However, B7-1 and B7-2 were not expressed. RPE significantly suppressed MLR in the absence of IFN-gamma prestimulation. This phenomenon was enhanced by IFN-gamma stimulation. The addition of 1-methyl tryptophan (1-MT) into the culture medium successfully reversed RPE immunosuppression., Conclusions: The expression of IDO may in part explain RPE-mediated immune suppression effects.

Purpose: Ocular neovascularization is the primary cause of blindness in a wide range of prevalent ocular diseases including proliferative diabetic retinopathy, exudative age-related macular degeneration, and retinopathy of prematurity, among others. Antiangiogenic therapies are starting to give promising results in these diseases. In the present study the antiangiogenic potential of an 18-mer peptide derived from type 1 thrombospondin repeat-containing protein WISP-1 (wispostatin-1) was analyzed in vitro with human retinal endothelial cell proliferation and migration assays. The peptide was also tested in vivo in the corneal micropocket and the laser-induced choroidal neovascularization (CNV) mouse models., Methods: Human retinal endothelial cells were treated with the WISP-1 peptide and in vitro migration and proliferation assays were performed. Also evaluated was the antiangiogenic effect of this peptide in vivo using the corneal micropocket assay and the laser-induced CNV model., Results: Wispostatin-1 derived peptide demonstrated antimigratory and antiproliferative activity in vitro. Wispostatin-1 completely abolished bFGF-induced neovascularization in the corneal micropocket assay. The peptide also demonstrated significant inhibition of laser-induced CNV., Conclusions: An inhibitory effect of Wispostatin-1 on ocular neovascularization was found in vitro and in vivo. The identification of novel and potent endogenous peptide inhibitors provides insight into the pathogenesis of corneal and choroidal neovascularization. The results demonstrate potential for therapeutic application in prevalent ocular disease.

Objective: To evaluate the effects of phacoemulsification energy on the redox state and mitochondrial distribution of cultured human endothelial cells., Methods: Human corneal endothelial cells from fresh banked human donor tissue not suitable for transplantation were harvested and cultured. Cellular autofluorescence images were obtained using an inverted microscope. The redox fluorometric ratio, which can be related to oxidative stress, was calculated as the net value of fluorescence from the 4,6-diamidino-2-phenylindole channel divided by the net value of fluorescence from the fluorescein isothiocyanate-conjugated channel after subtraction of background. For determining the mitochondrial distribution patterns, the cell area was divided by drawing a line halfway between the nuclear and cell membranes. The average fluorescence in the central area was divided by the average fluorescence in the peripheral area. This ratio was compared., Results: Human corneal endothelial cells exposed to increasing phacoemulsification times and increasing ultrasonic energy levels displayed dose-dependent decreases in measured redox ratios. Lower redox ratios in response to phacoemulsification did not associate with decreases in cell size or altered patterns of mitochondrial localization., Conclusion: Redox fluorometry may serve as a useful indicator for the in vitro study of human corneal endothelial cell physiological response to ultrasonic stressors and potentially other nonoxidative stressors., Clinical Relevance: Redox fluorometry in combination with human corneal endothelial cell morphometric measurements has potential to serve as an indicator of human corneal endothelial cell injury resulting secondary to ultrasound phacoemulsification.

Purpose: To report the immunosuppressive property of dry human amniotic membrane (dHAM)., Methods: Mouse splenocytes harvested from Balb/c mice were stimulated using functional-grade anti-CD3e antibodies for 4 days either with or without either of 2 commercial types of dHAM (Ambiodry 1 & 2, IOP Inc., Costa Mesa, Calif., USA) added to the culture media. The cell proliferation assay was performed to analyze the extent of splenocyte proliferation., Results: dHAMs significantly suppressed mouse splenocyte proliferation compared to control. The suppression by dHAM with intact amniotic epithelium (Ambiodry 2) was significantly stronger than dHAM without epithelium (Ambiodry 1)., Conclusion: The immunosuppressive property of dHAM was demonstrated using an alloreactive splenocyte proliferation assay., ((c) 2008 S. Karger AG, Basel.)

The purpose of this study was to evaluate the effects of topical human amniotic fluid (HAF) and equine amniotic fluid (EAF) on corneal reepithelialization and stromal wound healing. New Zealand white rabbit corneas (n=52) were placed in an ex vivo air-interface organ culture. An 8.5mm-diameter mark in the center of the cornea was produced with a hand trephine to select the area for epithelial scraping. A number 15 surgical blade was used to remove the epithelial layer within the demarcated area in a standardized fashion. The corneas were assigned to one of four treatment groups (n=8): fetal bovine serum (FBS), HAF, EAF, and a control group that was exposed to phosphate buffer solution (PBS). Corneal epithelial defects were imaged every 8 h for 72 h after the application of a 30 microl drop of 0.015% fluorescein. Five corneas of each treatment group were used for histology, proliferation, and apoptosis assay at 72 h after the epithelial defect was created. There was no significant difference in the mean rate of closure of the corneal epithelial defect between FBS treated corneas and controls (P>0.06). The mean epithelial defect area (MEDA) was significantly smaller in the EAF group as compared to control corneas at 24 h (P=0.016), 40 h (P=0.032), 64 h (P=0.008) and 72 h (P=0.007) following epithelial scrape. The MEDA in the HAF group was significantly smaller at 16 h (P=0.008), 64 h (P=0.0072), and 72 h (P=0.016) compared to the control group. The MEDA in the HAF and EAF groups was smaller at all time points as compared to the FBS group, but the difference was not significant. At histology, the mean keratocyte density was significantly higher in the anterior stroma in the HAF (P<0.001) and EAF groups (P=0.001) as compared to control group. The number of BrdU positive keratocytes was significantly higher in the superficial and deep stromal sub-areas in the HAF group as compared to control (P<0.001 and P=0.002, respectively). EAF and FBS treated corneas also showed a higher number of BrdU positive cells compared to control, but this difference was not significant. Finally, we did not observe any difference in the amount of TUNEL positive keratocytes among the different groups. Our data indicates that the topical application of HAF and EAF is associated with accelerated reepithelialization in this cornea organ culture model. Similarly, corneal keratocyte density appears to be less affected after epithelial injury using this treatment.

Purpose: To quantify the effects of oxidant challenge on the redox state of adult human retinal pigment epithelial cells using microscopic autofluorescence spectroscopy and to determine whether treatment with the isothiocyanate sulforaphane protects these cells against oxidative stress., Methods: Oxidative stress was evoked in ARPE-19 cells by H2O2 and tert-butyl hydroperoxide. Reduced nicotinamide nucleotides NAD(P)H were assessed by excitation at 366 nm with measurement of fluorescence at 450 nm. Oxidized flavoproteins were assessed by excitation at 460 nm with measurement of fluorescence at 540 nm. The ratio of these measurements served as the index of cellular redox status., Results: Redox ratio and cell viability decreased in a dose-dependent manner after oxidant exposure. ARPE-19 cells treated with sulforaphane maintained significantly higher redox ratio and cell viability. The ratio for sulforaphane-treated cells after exposure to 0.64 mM H2O2 was 2.64 +/- 0.19 compared with 1.77 +/- 0.16 in untreated cells (P = 0.001). At 1.2 mM H2O2, the redox ratio of sulforaphane-treated cells was 2.30 +/- 0.18 compared with 1.76 +/- 0.13 in untreated cells (P = 0.02). Similar results were observed after insult with tert-butyl hydroperoxide., Conclusions: Redox fluorometry provides quantitative information on the redox status of living cells. Sulforaphane protects ARPE-19 cells from oxidative injury by induction of antioxidant phase 2 genes. The findings in this study describe a useful method for assessing antioxidant effects in live cells and support phase 2 gene induction as a potential treatment strategy for macular degeneration and diseases in which oxidative injury plays a causative role.

Background: There are few reports in the literature to describe the natural history of an occupational Nd:YAG laser-induced retinal injury., Methods: The chronological response to a Nd:YAG laser-induced injury was studied in a 48-year-old male using the techniques of color fundus photography, fluorescein angiography and optical coherence tomography. When he developed choroidal neovascularization his response to photodynamic therapy with verteporfin was recorded., Results: The patient initially developed a macular hole, which spontaneously closed. Three months after the injury a choroidal neovascular membrane developed. This stabilized after the use of photodynamic therapy., Conclusions: Surgical closure of Nd:YAG induced macular holes should be delayed to allow spontaneous closure. Photodynamic therapy was successful in treating choroidal neovascularization in this case.

We have previously identified a neuroprotective effect associated with empty (E1(-), E3(-), E4(-)) adenovirus vector delivery in a model of light-induced, photoreceptor cell death. In this study, we further characterize this protective effect in light-injured retina and investigate its molecular basis. Dark-adapted BALB/c mice, aged 6-8 weeks, were exposed to standardized, intense fluorescent light for 96 or 144 h. Prior to dark adaptation, all mice received intravitreous injection of 1 x 10(9) particles of an empty (E1(-), E3(-), E4(-)) adenovirus vector in one eye and vehicle in the other. Following light challenge of 96 or 144 h, histopathological analysis and quantitative photoreceptor cell counts were conducted. Semiquantitative assessment of messenger ribonucleic acid (mRNA) for the apoptosis related genes: p50, p65, IkBa, caspase-1, caspase-3, Bad, c-Jun, Bax, Bak, Bcl-2, c-Fos, and p53 using quantitative reverse transcriptase polymerase chain reaction was performed on eyes following 12 h of light exposure. Following 96 h of light exposure, the photoreceptor cell density for E1(-), E3(-), E4(-) adenovirus vector and vehicle-injected eyes were 87.5 +/- 9.5 and 79.3 +/- 10.1, respectively, (p = 0.79). After 144 h of light exposure, the photoreceptor cell density was preserved in vector-injected eyes as compared to vehicle treated eyes, 68.9 +/- 10.0 and 49.2 +/- 4.6, respectively (p = 0.016). Relative mRNA levels of c-Fos and c-Jun at 12-h light exposure after injection differed significantly between vector- and vehicle-injected eyes (p = 0.036, 0.016, respectively). The expression of the other apoptosis-related genes evaluated was not significantly affected. This study investigates the molecular basis of photoreceptor neuroprotective pathway induction associated with E1(-), E3(-), E4(-) adenovirus vectors. The results indicate that empty adenovirus vectors protect photoreceptors from light-induced degeneration by the modulation of apoptotic pathways. Gene expression changes suggest that the suppression of c-Fos and c-Jun upregulation contributes significantly to the neuroprotective effect. Understanding the molecular basis of the neuroprotective pathway induction in photoreceptors is critical to the development of novel therapies for retinal degenerations.

The peroxisome proliferator-activated receptors (PPAR's) are members of the steroid/thyroid nuclear receptor, superfamily of transcription factors. There are currently three known PPAR subtypes, alpha, beta, and gamma. The PPARs are now recognized participants in a number of biological pathways some of which are implicated in the pathogenesis of age-related macular degeneration (AMD). These include immune modulation, lipid regulation, and oxidant/antioxidant pathways important to the onset and progression of "dry" AMD, and vascular endothelial growth factor (VEGF) mediated pathways that stimulate choroidal neovascularization (CNV), characteristic of "wet" AMD. PPAR-alpha is found in retina and also on vascular cells important to formation of CNV. At this time, however, relatively little is known about potential contributions of PPAR-alpha to the pathogenesis of dry and wet AMD. This review examines current literature for potential roles of PPAR-alpha in the pathogenesis and potential treatment of AMD with emphasis on prevention and treatment of wet AMD.

Purpose: A noninvasive imaging technique was used for serial assessment of gene expression after intraocular gene transfer. Bioluminescence after intravitreous administration of an adenovirus vector containing the firefly luciferase gene was measured serially and noninvasively. The optical signal was then used as a bioassay to determine whether periocular immune modulation affects intraocular transgene expression., Methods: Sixty-two, 8-week-old, male BALB/c mice were used. The correlation of optical signal intensity was determined by tissue luciferase level after injecting 30 mice with one of three intravitreous doses of Ad-Luc-GFP (10(8), 5 x 10(8), or 10(9) particles in 1 microL). Ocular bioluminescence was measured at days 2, 5, 8, and 14. The bioluminescence was then directly compared with measured tissue luciferase levels. The remaining 32 mice were divided into two groups. One group (n = 16), was injected with periocular corticosteroid (400 mug in 10 microL). Two days later, Ad-Luc-GFP was administered by intravitreous injection (10(9) particles in 1 microL). The remaining mice (n = 16) were injected with the same dose of intravitreous Ad-Luc-GFP without corticosteroid pretreatment. Ocular bioluminescence was then assessed longitudinally on days 2, 4, 6, 8, 11, 14, 22, and 30 after intravitreous injection in n = 10 mice per group. The optical signal intensity in each group was compared over the study period. The remaining 12 mice (n = 6, each group) were used to assess histologic differences between the two groups., Results: In vivo measurement of ocular bioluminescence was well correlated with tissue luciferase levels (Spearman's correlation, r = 0.969, P < 0.001). Periocular TA injection markedly decreased the acute inflammatory reaction associated with intravitreous Ad-Luc-GFP and was associated with a significant increase in the duration of peak luciferase expression as well as the total period of luciferase expression., Conclusions: A significant enhancement of intraocular transgene expression is associated with periocular pretreatment with corticosteroid. Histologic evidence of immune cell reduction in ocular tissues in corticosteroid-treated eyes implies a local immune response. Periocular treatment with corticosteroids may enhance adenovirus-mediated gene expression in the eye.

Purpose: To determine the effect of keratoconjunctivitis sicca, induced by botulinum toxin-B (BTX-B), on the inflammatory cytokine gene expression in the lacrimal gland (LG). And to determine the effect of various topical anti-inflammatory agents on the resulting cytokine levels., Methods: Forty-two mice (eight-week-old, female, CBA/J) were divided into six groups. Four groups were injected with BTX-B into both lacrimal glands, one group was injected with saline into both LG (Sal, n=7), and one group served as an uninjected control (Con, n=7). The four groups of BTX-B injected mice were then assigned to a treatment group: 1. no additional treatment (BTX), 2. artificial tear treatment (AT), 3. Cyclosporine A (CSA) treatment, and 4. fluorometholone (FM) treatment (n=7 in each group). Corneal fluorescein staining was evaluated one, two, and four weeks after injection. LGs were harvested after two weeks (groups Con, Sal, and BTX) and four weeks (groups AT, CSA, and FM) after injection. Gene microarray analysis for inflammatory cytokines and their receptors, real time reverse-transcriptase polymerase chain reaction (RT-PCR), and immunofluorescent staining with anti-mouse CD3e monoclonal antibody were then performed on LG tissue., Results: BTX-B injection into the LG effectively induced dry eye in mice two and four weeks following injection. Microarray data identified the proinflammatory cytokines interleukin (IL)-1, tumor necrosis factor (TNF)-alpha, IL-12, and macrophage migration inhibitory factor (MIF) and the anti-inflammatory cytokines IL-10 and toll-interacting protein (Tollip) as candidates for validation by real time RT-PCR. MIF and IL-12 expression were elevated in BTX-B injected mice at weeks 2 and 4 regardless of treatment. Tollip and IL-1 expressions were increased in some groups after BTX-B injection regardless of the treatment type. Other cytokines showed no significant changes. LG structures were well maintained without significant T lymphocyte infiltration in all groups., Conclusions: Ocular surface change induced by BTX-B injection resulted in an altered expression of various inflammatory cytokines in our murine dry eye model. Alteration of the pathology-induced cytokine profile by topical therapy is reported.

Purpose: Ethnic variation has been reported in age-related macular degeneration (AMD)-associated Y402H polymorphism in complement factor H (CFH). This variation is evident in the Japanese population. Recently a strong association between a novel single-nucleotide polymorphism (SNP; rs1410996) in the CFH gene and AMD has been identified in Caucasian patients. The present study was undertaken to investigate whether four coding and noncoding variants of the CFH gene, including rs1410996, are associated with AMD in native, unrelated Japanese patients., Methods: A total of 188 patients with AMD and 139 control subjects without AMD were recruited for the study. Four SNPs (rs800292, rs1061170, rs1410996, and rs2274700) in the CFH gene were assessed by genotyping assay. The information regarding systemic conditions and lifestyle including smoking were documented in each subject by standardized questionnaire., Results: The intronic SNP (rs1410996) and the synonymous SNP (rs2274700) were associated with a significant risk of AMD (P = 2.37 x 10(-5) and 3.52 x 10(-5), respectively). A significant association was also noted between a coding variant (rs800292, I62V) and AMD (P = 8.63 x 10(-6)). In contrast, the Y402H variant showed no significant association with AMD (P = 0.101). Two common haplotypes also demonstrated significant association with AMD (P = 1.08 x 10(-3) and 2.00 x 10(-5)). Among the environmental factors, smoking alone had a significant association with AMD (P = 1.17 x 10(-4))., Conclusions: Although the Y402H variant was not significantly associated with AMD, other coding and noncoding variants in the CFH gene including rs1410996 and smoking moderately influenced the risk of AMD in a Japanese population.

We report noninvasive autofluorescence mitochondrial imaging in cultured human corneal endothelial cells (HCECs). HCECs harvested from eye bank corneas were cultured in thin glass-bottom plates. Mitochondria were imaged with an autofluorescence microscope using a DAPI filter set (excitation: G365, emission: band pass 445/50) and then, after fixation with 4% paraformaldehyde, cells were stained with MitoTracker Green FM (MTG). Both images were aligned using a linear conformal algorithm for image mapping based on manually selected corresponding feature points, and then mathematically compared using two-dimensional spatial image correlation coefficients. Autofluorescence imaging provided highly resolved mitochondrial signals from living HCECs, comparable to those taken with MTG. Both techniques yielded very similar images at high magnification and high resolution, demonstrating the tubular morphology and cytoplasmic distribution that are characteristic of mitochondria. Image registration using a linear conformal mapping technique and cross-correlations showed high correlation of overlapping autofluorescence and MTG images. This study validates the novel use of autofluorescence vital imaging as a noninvasive, inexpensive and functional alternative to the mitochondria-specific dyes in cultured HCEC. This noninvasive mitochondrial imaging technique can be useful in future applications studying mitochondrial biology of ocular cells.

Purpose: To demonstrate the morphological and functional outcomes of intravitreous injection of an expansile gas bubble for the treatment of stage 2 macular holes., Design: Prospective interventional case series., Participants: Twenty eyes of 20 consecutive patients with unilateral stage 2 macular holes., Methods: The patients underwent intravitreous sulfur hexafluoride injection, followed by postoperative facedown positioning for 3 to 5 days. The patients were observed per protocol schedule with complete ophthalmological examination, including determination of corrected visual acuity (VA), slit-lamp biomicroscopic examination, funduscopic examination, and optical coherence tomography (OCT). Follow-up was greater than 12 months for all patients (mean, 19.5)., Main Outcome Measures: Posterior vitreous detachment, anatomical closure of the hole, VA, and hole diameter as measured by OCT., Results: Detachment of the posterior vitreous at the macula was achieved in 19 of 20 eyes (95%). Ten cases (50%) had anatomical closure of the hole with intravitreous gas injection alone. The remaining 10 cases (50%) achieved anatomical closure of the hole after subsequent vitreous surgery. There was a significant difference in hole diameter (P = 0.004) and in pretreatment vision (P = 4.5x10(-5)) in patients for whom gas alone resulted in hole closure and those proceeding to vitreous surgery. Hole closure by gas injection alone was achieved in 7 of 7 eyes (100%) with pretreatment vision better than 20/40 and in 6 of 7 eyes (86%) when the hole was smaller than 200 mum. There were no major complications in this series of patients. Successfully treated macular holes remained closed at all follow-up points during the study period., Conclusion: An intravitreous injection of an expansile concentration of the inert sulfur hexafluoride gas alone frequently induces detachment of the posterior vitreous in the aged eye. Anatomical closure of the hole without major complications is more likely in smaller holes with better pretreatment vision. This technique may have clinical application for stage 2 holes in selected cases. A number of potential advantages including decreased morbidity and a potential cost savings may result from successful utilization of this procedure.

The purpose of this investigation was to determine whether the high-temperature requirement A-1 (HTRA1) gene polymorphism is associated with age-related macular degeneration (AMD) in native, unrelated Japanese patients. A total of 123 patients with AMD and 133 control subjects without AMD were recruited for this study. The single-nucleotide polymorphism (SNP) rs11200638 in the HTRA1 gene was assessed using a TaqMan assay. The risk A allele frequencies in the AMD cases and control patients were 0.577 and 0.380, respectively, and were associated with a significant risk of developing AMD (p=7.75x10(-6)). The results were more significant in subtype analyses with wet AMD (p=5.96x10(-7)). We conclude that the rs11200638 variant in the HTRA1 gene is strongly associated with AMD in the Japanese population. This result supports the hypothesis that the HTRA1 gene may increase susceptibility to AMD development and can participate in a potential new molecular pathway for AMD pathogenesis by extending this association across diverse ethnicities.

Purpose: To evaluate the efficacy of topical human amniotic fluid (HAF) in the treatment of ocular acute alkali burns in mice., Design: Experimental study., Methods: A chemical burn with 2 microl of sodium hydroxide 0.15 mol/l was created in one eye of 30 mice. The animals were divided into gender- and age-matched groups according to the topical treatment that was administered: group 1 was treated with preterm HAF (n = 10 mice); group 2 was treated with term HAF (n = 10 mice), and group 3 was treated with saline solution (n = 10 mice). Treatment consisted of one drop that was applied to the burned eye five times per day (week one), and three times per day (week two). The epithelial defect was photographed and measured on days two and four. Ocular burn damage was assessed at days two, seven, and 14 after a pre-established classification. On day 14, both eyes of each mouse were enucleated and assessed histopathologically., Results: Median epithelial defect (interquartile range [IQR], 25th, 75th percentile) at day four was 9.93% (IQR, 8.57, 11.27) for group 1, 7.30% (IQR, 5.96, 8.97) for group 2, and 18.92% (IQR, 11.71, 27.64) for group 3 (P < .0076). The overall change (difference in slope) in ocular burn score between days 2 and 14 was -0.127 (P = .009) in group 1 vs 3, -0.134 (P = .012) in group 2 vs 3, and 0.007 (P = .88) in group 1 vs 2. On histologic examination saline solution-treated corneas had more inflammatory cells and blood vessels than HAF-treated corneas., Conclusion: Topical preterm/term HAF was an effective topical therapy for limiting the damage after acute alkali burns of the eye in this animal model.

Purpose: To study gene expression changes in the rat retina and choroid following transpupillary thermotherapy (TTT) and to identify molecular mechanisms that may enhance treatment of choroidal neovascularization, complicating age-related macular degeneration., Methods: One fundus of Brown Norway rats was treated with an 810 nm diode laser while the contralateral fundus received no treatment. The mRNA was extracted and processed for cDNA microarray analysis. Genes with increased expression were validated by semiquantitative reverse transcription polymerase chain reaction (PCR) and quantitative real-time PCR (qRT-PCR)., Results: Of the 14,815 cDNA elements on the array, 12 genes were up-regulated in TTT treated eyes. Upregulation of eight of these 12 genes could be verified by semiquantitative RT-PCR. The eight verified genes were EPCR, IL-1beta, MCP-1, TSP-1, Fgl, Asns, MT-2, and NMDMC, which included 4 angiogenesis- and coagulation-related genes., Conclusions: This study demonstrates upregulation of angiogenesis- and coagulation-related genes following TTT. The response profile and its temporal relationships provide insight into the molecular mechanisms that lead to vascular occlusion and antiangiogenesis induced by TTT.

Purpose: To report optical coherence tomography (OCT) histologic correlations of three common peripheral retinal lesions and to explore the feasibility of the use of OCT to image retinal pathologic conditions in fixed tissue specimens., Design: Experimental study., Methods: We studied peripheral retinal lesions that were available to us in 11 formalin-fixed caps of seven human eyes that had been enucleated previously for choroidal melanoma. OCT scans were performed through these lesions, and the images were correlated with corresponding histologic sections., Results: Three peripheral retinal diseases that included cystoid degeneration, localized retinal detachment, and paving stone degeneration were imaged successfully, and their optical characteristics correlated well with their histologic data., Conclusion: We explored potential utility for OCT imaging of peripheral retinal pathologic conditions in fixed tissue specimens. That these OCT images correlate with findings from histologic sections suggests a possible role for OCT in the eye pathology laboratory.

Purpose: To further define the origin of the "outer red line" (ORL) present in retinal optical coherence tomograms., Design: Laboratory investigation., Methods: Eleven formalin-fixed caps of seven human eyes previously enucleated for choroidal melanoma at the Wilmer Eye Institute were studied. The ORL evaluation consisted of sequential, surgical elimination of the retinal pigment epithelium, Bruch's membrane (BM), and choroid in separate, but adjacent, areas. Optical coherence tomography (OCT) images through each of these areas were correlated with corresponding histologic sections. The relative contribution of the retinal pigment epithelium, BM, and choroid to the ORL in retinal OCT3 images was evaluated., Results: In the specimens examined, the ORL correlated anatomically with a highly reflective optical surface at the level of the outer retina, and did not appear to represent a specific anatomic layer. The reflecting layers appeared to be composed of the retinal pigment epithelium, BM, and inner choroid. The relative contribution of BM together with the inner choroid to the ORL appeared greater than that of the retinal pigment epithelium., Conclusions: This study suggests that the ORL present in retinal OCT3 images and generally associated with the retinal pigment epithelium/BM/inner choroidal complex appears to indicate a highly reflective optical surface which, when eye layers are normally configured, represents this complex of anatomic structures with a predominant contribution from BM and inner choroid. The findings of this study suggest a potential role for OCT in the eye pathology laboratory.

Purpose: To evaluate the transduction efficiency and localization of a reporter gene after intravitreous injection of adenovirus vector in laser photocoagulation (PC)- and photodynamic therapy (PDT)-treated eyes., Methods: Adult Lewis rats received fundus PC, fundus PDT, or no treatment. Intravitreous injection of an adenovirus vector containing the construct expressing beta-galactosidase (AdlacZ.11D) was performed in each group. All eyes were then enucleated for histochemistry and processed for quantitative image analysis., Results: In eyes with no treatment, there was moderate to intense staining for lacZ in the anterior segment, but little in the retina. In eyes treated with PC and PDT, there was significantly more LacZ staining in the retina. The increased staining corresponded closely with the sites treated with PC and PDT. Gene transduction in PC-treated eyes was enhanced and extended to at least 135 days after virus delivery, but not extended in PDT-treated eyes. Gene transfer and expression were targeted and enhanced at the site of laser burns, at all doses tested (3 x 10(5) to 3 x 10(9) particles per eye)., Conclusions: Compared with untreated eyes, eyes treated with PC and to a lesser extent PDT, manifest increased transduction efficiency, in areas of the retina that are targeted by laser treatment. This finding suggests a new and promising strategy for the treatment of retinochoroidal neovascularization. Adenovirus gene therapy in combination with PC or PDT would have the advantage of increased transduction efficiency; increased duration of transgene expression; targeted delivery; and, potentially, a lower effective dose of virus.

Purpose: To compare the angioarchitecture of choroidal arteries and veins in patients with age-related macular degeneration (AMD) to the angioarchitecture of age-matched normal subjects using indocyanine green (ICG) angiography., Methods: ICG angiography was performed in 35 consecutive AMD patients and 18 normal age-matched volunteers with a fundus ICG camera. ICG video images, including the arterial and venous phases, were quantitatively analyzed using image analyzing software., Results: In patients with AMD, the choroidal arterioles are dilated, fewer, run a straighter course, and possess fewer bifurcations. The number of choroidal arteries and the macular fluorescent intensity in the arterial phase of choroidal filling was significantly less in patients with AMD as compared to age-matched normal controls (P = 0.008). The mean and maximum caliber of choroidal veins in the macula was dilated in AMD eyes than in age-matched normal control eyes (P < 0.001). There was no statistically significant difference in arterial dye filling or venous caliber observed in AMD eyes, with or without choroidal neovascular membrane (CNV)., Conclusion: Choroidal arterial perfusion in the macula was significant decreased in eyes with AMD with and without CNV, and was associated with choroidal venous dilation. These observations implicate poor choroidal perfusion of the macula in the pathogenesis of AMD.

In this study, we investigated whether intraocular gene transfer of pigment epithelium-derived factor (PEDF) ameliorates the extent of light-induced photoreceptor cell death. Lewis rats received intravitreous injection of 3 x 10(9) particles of adenoviral vector expressing PEDF (AdPEDF.11) in one eye and 3 x 10(9) particles of empty adenoviral vector (AdNull.11) in the contralateral eye. The rats were then dark-adapted for 3 days after which they were continuously exposed to fluorescent light (2,500 lux) for 0, 6, 24, 96, and 168 h. Both eyes were then enucleated and processed for morphometric analysis. Cell death in the retina was examined using TUNEL staining with a propidium iodide counterstain. The photoreceptor cell counts in each of the three groups were significantly different (P < 0.001). Eyes that received intravitreous injection of AdNull.11 or no injection showed a greater number of pyknotic photoreceptor cells and a reduced photoreceptor cell density as compared to eyes treated with intravitreous AdPEDF.11 injection. AdNull.11 treated eyes showed a lesser but still significant protection of photoreceptor cells when compared to untreated eyes. Fewer TUNEL-positive photoreceptor cells were present in AdPEDF.11 treated eyes than in AdNull.11 treated or untreated eyes (P = 0.004). The amplitudes of the ERG a-wave, b-wave, and oscillatory potentials (OPs) were increased significantly by treatment (P < 0.05). These data suggest that adenovirus vector-mediated intraocular expression of PEDF significantly increases photoreceptor cell survival following excessive light exposure. Neuroprotection may result from inhibition of light-induced apoptotic processes. This study provides proof of concept for a gene transfer approach to modulating retinal cell death resulting from photo-oxidative damage and supports the hypothesis that gene transfer of PEDF is broadly applicable to modulating apoptosis in the retina., (2004 Wiley-Liss, Inc.)

Purpose: To describe the angioarchitecture of choroidal veins in normal subjects using indocyanine green (ICG) angiography., Design: Consecutive observational case series., Participants: Thirty-six eyes of 33 healthy volunteers ranging in age from 21 to 81 years (50.0+/-18.2 [mean +/- standard deviation])., Methods: Indocyanine green angiography was performed with a modified Topcon fundus ICG camera. Montage images were obtained during the venous phase of the ICG angiogram., Main Outcome Measures: Indocyanine green angiographic montage images covering all areas of the observable fundus., Results: Eighteen of 36 eyes examined (50%) showed asymmetry of choroidal venous vascular patterns consistent with a preferential route of choroidal venous drainage of the macular region. Twelve of the 18 eyes with a preferred drainage route drained superotemporally; the remaining 6 eyes drained by either an inferotemporal or a superonasal route. A preferential choroidal venous drainage route in the macula was identified with equal frequency in young and old subjects. Of 36 eyes examined, 27 (75%) lacked ICG angiographic evidence of symmetric separation of outer choroidal veins., Conclusions: There is ICG angiographic evidence of asymmetry of choroidal venous drainage in one half of normal individuals. The relative frequency of a resulting preferential drainage route in a group of normal subjects and the equal distribution in young and old subjects indicate that this finding is neither pathologic nor attributable to aging. There is also ICG angiographic evidence for a lack of symmetric separation in the outer choroidal venous system in the majority of normal human subjects tested, indicating that classic watershed zones may not be present or are less prominent in this portion of the choroidal vasculature.

Purpose: To compare optical coherence tomography (OCT) images of idiopathic epiretinal membranes (ERMs) with those of secondary ERMs., Methods: OCT was performed on 70 eyes of 63 consecutive patients with biomicroscopic evidence of ERMs and 23 eyes of 23 healthy volunteers without ERMs. OCT findings were correlated with the clinical pathogenesis of the ERM., Results: Evaluation by OCT established that 48 of 70 ERMs were globally adherent to the retina and that 22 of 70 ERMs were focally adherent to the retina. When correlated to clinical pathogenesis, 20% of idiopathic membranes and 52% of secondary membranes were focally attached to the retina. There was a significant difference in the pattern of membrane attachment to the retina in the two pathogenic groups (P = 0.007). Eight of nine eyes with macular pseudoholes were associated with globally adherent membranes., Conclusion: Secondary ERMs are more likely to be characterized by focal retinal adhesion than are primary ERMs. Primary ERMs tend to be globally adherent. This finding may contribute to understanding the underlying mechanisms of ERM formation in different clinical settings.

Purpose: To determine whether intraocular gene transfer of pigment epithelium-derived factor (PEDF) protects the retina from ischemia-reperfusion injury., Methods: Four days before induction of pressure-induced ischemia, Lewis rats received intravitreous injection of 3 x 10(9) particles of an adenovirus vector expressing PEDF (AdPEDF.11) in one eye and 3 x 10(9) particles of an empty adenovirus vector (AdNull.11) in the contralateral eye. Seven days after reperfusion, eyes were enucleated and processed for morphometric analysis. Apoptotic cells stained by TdT-dUTP terminal nick-end labeling (TUNEL) in the retina were counted 12 hours after initiation of reperfusion. Retina levels of PEDF were measured by enzyme-linked immunosorbent assay., Results: PEDF levels in retinal homogenates from eyes receiving AdPEDF.11 injection were well above the background levels in the untreated baseline and control eyes (P = 0.04). Retinal thickness was preserved in AdPEDF.11-treated eyes. Retinal cell density was significantly greater in the ganglion cell layer (GCL; P = 0.014), inner nuclear layer (INL; P = 0.008), and outer nuclear layer (ONL; P = 0.008) of AdPEDF.11-treated eyes compared with the corresponding layers in AdNull.11-treated eyes. AdNull.11-treated eyes also had significantly more TUNEL-positive cells in these layers than AdPEDF.11-treated eyes (P < 0.05)., Conclusions: Adenoviral vector-mediated intraocular expression of PEDF significantly increases cell survival after ischemia-reperfusion injury of the retina. The protective effect may result from inhibition of ischemia-induced apoptosis. This study provides proof of concept for a gene transfer approach directed at interrupting programmed cell death induced by retinal ischemic insult.

Purpose: To compare the characteristics of residual fundus fluorescence observed in the ultra-late phase of indocyanine green (ICG) angiography, in normal subjects and in patients with age-related macular degeneration (ARMD)., Methods: ICG angiography was performed on 38 patients, 21 had ARMD, 9 were normal subjects aged >62, and 8 were normal subjects aged <36. The intensity and pattern of fluorescence from angiograms obtained in the ultra-late phase, 24 hours after dye injection, was also recorded and analyzed., Results: In the ultra-late phase, 95% of ARMD eyes with CNV showed geographic hypofluorescent lesions. All of the CNV that could be delineated with fluorescein and/or ICG angiography were located in these geographic lesions. In 73% of ARMD eyes without CNV, these hypofluorescent lesions occurred, while age-matched normal subjects had no hypofluorescent lesions. The mean intensity of fluorescence in the normal older subject group was significantly higher than that seen in the normal young subject group., Conclusions: Increased fluorescence, associated with older subjects, in the ultra-late phase of ICG angiography may reflect aging changes in the chorioretinal complex. Geographic hypofluorescent areas, demonstrated only in the ultra-late phase, associate with ARMD and may represent areas predisposed to CNV development.

Age-Related Macular Degeneration (AMD) is, together with Diabetic Retinopathy, the most common cause of vision loss among adults in the U.S. and other developed countries. In the U.S., at least 1.7 million people have impaired vision due to AMD. Every year, more than 165,000 people contract AMD and 16,000 go blind from it, predominantly from a rapidly progressing form of the disease called "wet" AMD. Wet AMD is characterized by serious or hemorrhagic detachment of the retinal pigment epithelium and choroidal neovascularization. The macula has the highest concentration of photoreceptors facilitating central vision and permitting high-resolution visual acuity. The damage caused by the leakage and fibrovascular scarring in wet AMD leads to profound loss of central vision and severe loss of visual acuity (usually 20/200 or worse). People with wet AMD have several limitations, including inability to read, inability to recognize faces or drive, and the disease often leads to blindness. The onset of severe visual changes in wet AMD can occur suddenly. More than 400,000 Americans are currently affected by this form of the disease, and the incidence is rising rapidly with the aging of the population. Therefore, the serious consequences of this disease along with the limited treatment options and their effectiveness make this a very good candidate for a gene transfer treatment approach. The investigational agent, Ad(GV)PEDF.11D, is an E1-, partial E3-, E4- deleted replication-deficient, adenovirus serotype 5, gene transfer vector. The transgene in this vector is the cDNA for human pigment epithelium-derived factor (PEDF). PEDF is one of the most potent known antiangiogenic proteins found in humans. While Ad(GV)PEDF.11D is able to transduce many somatic cell types, the natural barrier to other tissues created by the retina limits the ability of Ad(GV)PEDF.11D to affect tissues other than in the eye. Intravitreal administration of Ad(GV)PEDF.11D provides a convenient means of delivering PEDF to the relevant cells within the eye likely to result in a more prolonged duration of effect versus administration of the PEDF protein alone. In three murine disease models (the laser-induced choroidal neovascularization model, the VEGF transgenic model, and the retinopathy of prematurity model) significant inhibition of neovascularization (up to 85%) was demonstrated with doses of Ad(GV)PEDF vectors ranging from 1 x 10(8) to 1 x 10(9) pu. In toxicology studies performed in Cynomolgus monkeys, a dose-related inflammatory response was observed. A dose of 1 x 10(8) pu caused no adverse effects, while the inflammatory response observed at 1 x 10(9) pu was minimal and fully reversible. The observed inflammatory response at doses of 1 x 10(10) and 5 x 10(10) pu were increasingly severe. The proposed clinical trial is an open-label, dose-escalation, phase I study to investigate the safety, tolerability and potential activity of intravitreal injection of Ad(GV)PEDF.11D in patients with wet AMD. Ad(GV)PEDF.11D will be injected once via intravitreal injection into the eye with the most advanced AMD based on visual acuity. Subjects will be age 50 or over and have severe wet AMD in at least one eye defined by a best-corrected vision of 20/200 or worse. The primary objectives of this investigation are: (1) to assess the safety, tolerability and feasibility of intravitreal injection of Ad(GV)PEDF.11D in patients with severe, neovascular AMD, (2) to identify the maximum tolerated dose (MTD) of Ad(GV)PEDF.11D, and (3) to get some indication of potential activity of Ad(GV)PEDF.11D.

Purpose: To report a series of infants who progressed from mild retinopathy of prematurity (ROP) to severe ROP with retinal detachment without demonstrating detectable threshold disease., Methods: Between January 1993 and August 1998, seven infants at Oregon Health Sciences University, followed in accordance with the Cryotherapy for Retinopathy of Prematurity Study (CRYO-ROP) protocol, progressed to retinal detachment despite documentation that threshold had not been reached. This outlying subset of patients was analyzed and compared to the cohort in the CRYO-ROP study., Results: Six of 7 patients were male, 6 (86%) patients had symmetric disease, and all patients were born outside the study hospital. Mean birth-weight was 877 g and mean gestational age was 26 weeks. Mean postconceptual age at the time of retinal detachment was 41 weeks. Because of bilateral detachment in 3 patients, the total number of study eyes is 10. Failure to achieve threshold resulted from insufficient clock hours or insufficient stage in 2 eyes and lack of plus in 8 eyes. Zone I disease was present in 1 eye., Conclusion: Rarely, despite adhering to ROP examination protocol, the retina may detach without demonstrating antecedent threshold disease. Very low birthweight is a factor that may lead to a less predictable course. This study found a lack of plus disease results in failure to reach threshold more often than the occurrence of insufficient clock hours of stage 3 disease. Further study is needed to determine if selected cases of subthreshold ROP may benefit from ablative therapy.

Increased intraocular pressure and vascular ligation models are often used in studies of global ocular ischemia. The purpose of this study is to perform a paired comparison of retinal recovery in these paradigms. Our data indicate that ERG b-wave recovery profiles, following identical periods of ischemia, differ significantly between models. We propose that increased intraocular pressure models induce greater retinal injury than vascular ligation models. We suggest that pressure or another aspect of the increased intraocular pressure model induces injury beyond that caused by ischemia alone and caution against direct comparison of results obtained using these two models.

Purpose: The iron chelating agent deferoxamine mesylate USP (Desferal, Ciba, Summit, NJ) is commonly used in the treatment of acute iron intoxication and chronic iron overload (associated with the transfusion-dependent anemias). When used for prolonged periods of time or in high doses deferoxamine is attended by a range of ocular toxicities. The visual symptoms associated with deferoxamine administration often limit effective iron chelation therapy and can result in permanent vision loss. Deferoxamine has recently been conjugated to certain high molecular weight biocompatible polymers without altering its iron-binding properties. Here the effect of conjugation of deferoxamine to hydroxyethyl starch on retinal toxicity is examined., Methods: An albino rat model of electroretinographically determined, deferoxamine-induced retinal toxicity has been previously described. We use this model to evaluate and compare both native deferoxamine and hydroxyethyl starch conjugated deferoxamine., Results: Our data show that retinal function, as assessed by the electroretinogram b-wave, is significantly depressed 1 day after a single dose of native deferoxamine, while the b-waves of rats receiving a single dose of hydroxyethyl starch-deferoxamine, are not significantly depressed at any time during the study. In addition, the administered dose of hydroxyethyl starch-deferoxamine resulted in plasma deferoxamine concentrations up to five times greater than those achieved with native deferoxamine., Conclusion: These results suggest that hydroxyethyl starch conjugated deferoxamine is associated with less retinal toxicity than native deferoxamine and that it may be a safer alternative for iron chelation therapy.

Purpose: To examine a photoelectric artifact associated with the ERG-jet corneal contact lens electrode (Universe SA, La Chaux-de-Fons, Switzerland)., Methods: An artifact associated with the ERG-jet, gold foil corneal contact lens electrode was reproduced in vitro and in vivo using 50 msec light flashes. In vitro responses were examined using light flashes that varied in intensity, duration, and wavelength. Ionic strength of the bathing solution and temperature dependence were also examined. In vivo responses were compared to similarly recorded signals using the Burian-Allen bipolar electrode., Results: The artifact is not apparent with microsecond light flashes, as with the Grass PS22 Photo-stimulator connected to a Ganzfeld. Longer light flashes and increasing light intensities, however, elicit graded responses that may resemble the late PIII component of the ERG in profile and in magnitude. The artifact varies with temperature, ionic concentration of the bathing medium, and wavelength of stimulating light. The artifact also varies in magnitude and polarity from one disposable electrode to the next. Light flashes of shorter wavelengths elicit greater responses than light flashes of equal radiant energy but of longer wavelengths., Conclusions: The artifact derives from electrode polarization occurring at the interface between the gold foil and its ionic medium. Caution is required when using light stimuli longer than 2-3 msec with this and similar types of intrinsically polarizable metal electrodes.