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Liu Y, Garrett ME, Yaspan BL, Bailey JC, Loomis SJ, Brilliant M, Budenz DL, Christen WG, Fingert JH, Gaasterland D, Gaasterland T, Kang JH, Lee RK, Lichter P, Moroi SE, Realini A, Richards JE, Schuman JS, Scott WK, Singh K, Sit AJ, Vollrath D, Weinreb R, Wollstein G, Zack DJ, Zhang K, Pericak-Vance MA, Haines JL, Pasquale LR, Wiggs JL, Allingham RR, Ashley-Koch AE, Hauser MA. DNA copy number variants of known glaucoma genes in relation to primary open-angle glaucoma. Invest Ophthalmol Vis Sci 2014;55(12):8251-8.Abstract

PURPOSE: We examined the role of DNA copy number variants (CNVs) of known glaucoma genes in relation to primary open angle glaucoma (POAG). METHODS: Our study included DNA samples from two studies (NEIGHBOR and GLAUGEN). All the samples were genotyped with the Illumina Human660W_Quad_v1 BeadChip. After removing non-blood-derived and amplified DNA samples, we applied quality control steps based on the mean Log R Ratio and the mean B allele frequency. Subsequently, data from 3057 DNA samples (1599 cases and 1458 controls) were analyzed with PennCNV software. We defined CNVs as those ≥5 kilobases (kb) in size and interrogated by ≥5 consecutive probes. We further limited our investigation to CNVs in known POAG-related genes, including CDKN2B-AS1, TMCO1, SIX1/SIX6, CAV1/CAV2, the LRP12-ZFPM2 region, GAS7, ATOH7, FNDC3B, CYP1B1, MYOC, OPTN, WDR36, SRBD1, TBK1, and GALC. RESULTS: Genomic duplications of CDKN2B-AS1 and TMCO1 were each found in a single case. Two cases carried duplications in the GAS7 region. Genomic deletions of SIX6 and ATOH7 were each identified in one case. One case carried a TBK1 deletion and another case carried a TBK1 duplication. No controls had duplications or deletions in these six genes. A single control had a duplication in the MYOC region. Deletions of GALC were observed in five cases and two controls. CONCLUSIONS: The CNV analysis of a large set of cases and controls revealed the presence of rare CNVs in known POAG susceptibility genes. Our data suggest that these rare CNVs may contribute to POAG pathogenesis and merit functional evaluation.

Cronin T, Vandenberghe LH, Hantz P, Juttner J, Reimann A, Kacsó A-E, Huckfeldt RM, Busskamp V, Kohler H, Lagali PS, Roska B, Bennett J. Efficient transduction and optogenetic stimulation of retinal bipolar cells by a synthetic adeno-associated virus capsid and promoter. EMBO Mol Med 2014;6(9):1175-90.Abstract

In this report, we describe the development of a modified adeno-associated virus (AAV) capsid and promoter for transduction of retinal ON-bipolar cells. The bipolar cells, which are post-synaptic to the photoreceptors, are important retinal targets for both basic and preclinical research. In particular, a therapeutic strategy under investigation for advanced forms of blindness involves using optogenetic molecules to render ON-bipolar cells light-sensitive. Currently, delivery of adequate levels of gene expression is a limiting step for this approach. The synthetic AAV capsid and promoter described here achieves high level of optogenetic transgene expression in ON-bipolar cells. This evokes high-frequency (~100 Hz) spiking responses in ganglion cells of previously blind, rd1, mice. Our vector is a promising vehicle for further development toward potential clinical use.

Swaminathan SS, Oh D-J, Kang MH, Rhee DJ. Aqueous outflow: segmental and distal flow. J Cataract Refract Surg 2014;40(8):1263-72.Abstract
UNLABELLED: The elevated intraocular pressure (IOP) of primary open-angle glaucoma is caused by impaired outflow of aqueous humor through the trabecular meshwork. Within the juxtacanalicular region, alterations of both extracellular matrix homeostasis and the cellular tone of trabecular meshwork endothelial and the inner wall of Schlemm canal cells affect outflow. Newer pharmacologic agents that target trabecular meshwork and Schlemm canal cell cytoskeleton lower IOP. Aqueous drainage occurs nonhomogenously with greater flow going through certain portions of the TM and less going through other portions-a concept known as segmental flow, which is theoretically the result of outflow being dependent on the presence of discrete pores within Schlemm canal. The limited long-term success of trabecular meshwork bypass surgeries implicates the potential impact of resistance in Schlemm canal itself and collector channels. Additionally, others have observed that outflow occurs preferentially near collector channels. These distal structures may be more important to aqueous outflow than previously believed. FINANCIAL DISCLOSURE: Dr. Rhee is a consultant to Aerie Pharmaceuticals, Alcon Laboratories, Inc., Allegan, Inc., Aquesys, Inc., Glaukos Corp., Ivantis, Inc., Johnson & Johnson, Merck Sharp & Dohme Corp. and Santen, Inc., and has received research funding from Alcon Laboratories, Inc., Merck Sharp & Dohme Corp., and Ivantis, Inc. No other author has a financial or proprietary interest in any material or method mentioned.
Rezende FA, Lapalme E, Qian CX, Smith LE, SanGiovanni JP, Sapieha P. Omega-3 supplementation combined with anti-vascular endothelial growth factor lowers vitreal levels of vascular endothelial growth factor in wet age-related macular degeneration. Am J Ophthalmol 2014;158(5):1071-1078.e1.Abstract
PURPOSE: To determine the influence of omega-3 supplementation on vitreous vascular endothelial growth factor A (VEGF-A) levels in patients with exudative age-related macular degeneration (wet AMD) receiving intravitreal anti-VEGF treatment. DESIGN: Prospective, randomized, open-label, single-center, clinical trial, consecutive interventional case series. METHODS: The study included 3 cohorts with wet AMD and a control group with epiretinal membrane or macular hole. Twenty wet AMD patients being treated with anti-VEGF were randomized to daily supplementation of antioxidants, zinc, and carotenoids with omega-3 fatty acids (docosahexaenoic acid and eicosapentaenoic acid; group 1, n = 10) or without omega-3 fatty acids (group 2, n = 10). They were compared with an anti-VEGF treatment-naïve wet AMD group (group 3, n = 10) and an epiretinal membrane or macular hole group (group 4, n = 10). Primary outcome was vitreal VEGF-A levels (at the time of anti-VEGF injection). Secondary outcomes were plasma VEGF-A and central foveal thickness. Patients with new submacular hemorrhage or any other treatment within 3 months were excluded. Final analyses included 9, 6, 7, and 8 patients in groups 1 through 4, respectively. RESULTS: Patients receiving omega-3s (group 1) had significantly lower levels of vitreal VEGF-A (141.11 ± 61.89 pg/mL) when compared with group 2 (626.09 ± 279.27 pg/mL; P = .036) and group 3 (735.48 ± 216.43 pg/mL; P = .013), but similar levels to group 4 (235.81 ± 33.99 pg/mL; P = .215). All groups showed similar values for plasma VEGF-A and central foveal thickness measurements. CONCLUSIONS: This study demonstrated that omega-3 supplementation combined with anti-VEGF treatment is associated with decreased vitreal VEGF-A levels in wet AMD patients.
Emami-Naeini P, Dohlman TH, Omoto M, Hattori T, Chen Y, Lee HS, Chauhan SK, Dana R. Soluble vascular endothelial growth factor receptor-3 suppresses allosensitization and promotes corneal allograft survival. Graefes Arch Clin Exp Ophthalmol 2014;252(11):1755-62.Abstract
PURPOSE: To investigate the effect of VEGF-C and VEGF-D blockade via soluble VEGFR-3 (sVEGFR-3) on T cell allosensitization, corneal neovascularization, and transplant survival. METHODS: Corneal intrastromal suture placement and allogeneic transplantation were performed on BALB/c mice to evaluate the effect of sVEGFR-3 on corneal neovascularization. Soluble VEGFR-3 trap was injected intraperitoneally to block VEGF-C/D (every other day starting the day of surgery). Immunohistochemical staining of corneal whole mounts was performed using anti-CD31 (PECAM-1) and anti-LYVE-1 antibodies to quantify the levels of hem- and lymphangiogenesis, respectively. Mixed lymphocyte reaction (MLR) was performed to assess indirect and direct host T cell allosensitization and the frequencies of IFN-γ-producing T cells in the draining lymph nodes were assessed using flow cytometry. Graft opacity and survival was evaluated by slit-lamp biomicroscopy. RESULTS: Treatment with sVEGFR-3 resulted in a significant blockade of lymphangiogenesis 2 weeks post-transplantation and significantly prolonged corneal allograft survival compared to the control group at 8 weeks post-transplantation (87.5 % vs. 50 %), and this was associated with significant reduction in the frequencies of allosensitized T cells and decreased frequencies of IFN-γ-producing CD4 T cells. CONCLUSIONS: Soluble VEGFR-3 suppresses corneal lymphangiogenesis and allograft rejection and may offer a viable therapeutic modality for corneal neovascularization and corneal transplantation.
Uchino Y, Uchino M, Yokoi N, Dogru M, Kawashima M, Okada N, Inaba T, Tamaki S, Komuro A, Sonomura Y, Kato H, Argüeso P, Kinoshita S, Tsubota K. Alteration of tear mucin 5AC in office workers using visual display terminals: The Osaka Study. JAMA Ophthalmol 2014;132(8):985-92.Abstract
IMPORTANCE: There are limited reports on the relationship between mucin 5AC (MUC5AC) concentrations in tears, working hours, and the frequency of ocular symptoms in visual display terminal (VDT) users. This investigation evaluated these relationships among patients with dry eye disease (DED) and individuals serving as controls. OBJECTIVE: To determine the relationship between MUC5AC concentration in the tears of VDT users based on the diagnosis of DED and frequency of ocular symptoms. DESIGN, SETTING, AND PARTICIPANTS: An institutional, cross-sectional study was conducted. Participants included 96 young and middle-aged Japanese office workers. Both eyes of 96 volunteers (60 men and 36 women) were studied. Participants working in a company that used VDTs completed questionnaires about their working hours and the frequency of ocular symptoms. Dry eye disease was diagnosed as definite or probable, or it was not present. Tear fluid was collected from the inferior fornix after instillation of 50 μL of sterilized saline. The MUC5AC concentration was normalized to tear protein content and expressed as MUC5AC (nanograms) per tear protein (milligrams). The differences in MUC5AC concentration between DED groups, between VDT working hours (short, intermediate, and long), and between symptomatic and asymptomatic groups were evaluated with 95% CIs based on nonparametric Hodges-Lehmann determination. MAIN OUTCOMES AND MEASURES: Ocular surface evaluation, prevalence of DED, and MUC5AC concentration. RESULTS: The prevalence of definite and probable DED was 9% (n = 9) and 57% (n = 55), respectively. The mean MUC5AC concentration was lower in the tears of VDT users with definite DED than in those with no DED (P = .02; Hodges-Lehmann estimator, -2.17; 95% CI, -4.67 to -0.30). The mean MUC5AC concentration in tears was lower in the group that worked longer hours than in the group that worked shorter hours (P = .049; estimated difference, -1.65; 95% CI, -3.12 to 0.00). Furthermore, MUC5AC concentration was lower in participants with symptomatic eye strain than in asymptomatic individuals (P = .001; estimated difference, -1.71; 95% CI, -2.86 to -0.63). CONCLUSIONS AND RELEVANCE: The data obtained in the present study suggest that office workers with prolonged VDT use, as well as those with an increased frequency of eye strain, have a low MUC5AC concentration in their tears. Furthermore, MUC5AC concentration in the tears of patients with DED may be lower than that in individuals without DED.
Shah AS, Prabhu SP, Sadiq MAA, Mantagos IS, Hunter DG, Dagi LR. Adjustable nasal transposition of split lateral rectus muscle for third nerve palsy. JAMA Ophthalmol 2014;132(8):963-9.Abstract
IMPORTANCE: Third nerve palsy causes disfiguring, incomitant strabismus with limited options for correction. OBJECTIVE: To evaluate the oculomotor outcomes, anatomical changes, and complications associated with adjustable nasal transposition of the split lateral rectus (LR) muscle, a novel technique for managing strabismus associated with third nerve palsy. DESIGN, SETTING, AND PARTICIPANTS: Retrospective medical record review appraising outcomes of 6 consecutive patients with third nerve palsy who underwent adjustable nasal transposition of the split LR muscle between 2010 and 2012 with follow-up of 5 to 25 months at a tertiary referral center. INTERVENTION: Adjustable nasal transposition of the split LR muscle. MAIN OUTCOMES AND MEASURES: The primary outcome was postoperative horizontal and vertical alignment. Secondary outcomes were (1) appraising the utility of adjustable positioning, (2) demonstrating the resultant anatomical changes using magnetic resonance imaging, and (3) identifying associated complications. RESULTS: Four of 6 patients successfully underwent the procedure. Of these, 3 patients achieved orthotropia. Median preoperative horizontal deviation was 68 prism diopters of exotropia and median postoperative horizontal deviation was 0 prism diopters (P = .04). Two patients had preoperative vertical misalignment that resolved with surgery. All 4 patients underwent intraoperative adjustment of LR positioning. Imaging demonstrated nasal redirection of each half of the LR muscle around the posterior globe, avoiding contact with the optic nerve; the apex of the split sat posterior to the globe. One patient had transient choroidal effusion and undercorrection. Imaging revealed, in this case, the apex of the split in contact with the globe at an anterolateral location, suggesting an inadequate posterior extent of the split. In 2 patients, the surgical procedure was not completed because of an inability to nasally transpose a previously operated-on LR muscle. CONCLUSIONS AND RELEVANCE: Adjustable nasal transposition of the split LR muscle can achieve excellent oculomotor alignment in some cases of third nerve palsy. The adjustable modification allows optimization of horizontal and vertical alignment. Imaging confirms that the split LR muscle tethers the globe, rotating it toward primary position. Case selection is critical because severe LR contracture, extensive scarring from prior strabismus surgery, or inadequate splitting of the LR muscle may reduce the likelihood of success and increase the risk of sight-threatening complications. Considering this uncertainty, more experience is necessary before widespread adoption of this technique should be considered.
Jakobiec FA, Trief D, Rashid A, Rose MF, Minckler D, Vanderveen D, Mukai S. New insights into the development of infantile intraocular medulloepithelioma. Am J Ophthalmol 2014;158(6):1275-1296.e1.Abstract
PURPOSE: To define the maturational sequence of 3 infantile intraocular medulloepitheliomas. DESIGN: Retrospective clinicohistopathologic and immunohistochemical study. METHODS: Immunoreactivity of paraffin sections for CRX (cone-rod homebox transcription factor) and NeuN (biomarker for neuronal differentiation) were investigated together with other biomarkers, including S100, glial fibrillary acidic protein, epithelial membrane antigen, and various cytokeratins. RESULTS: Three infants (aged 1, 6, and 8 months) had iris neovascularization, 2 had anterior ciliary body tumors, and 1 a posterior tumor associated with a retinochoroidal coloboma. Each tumor displayed a premedullary monolayer of cuboidal epithelium that was S100(+), NeuN(-), and CRX(-) and that transitioned into a multilaminar medullary epithelium forming neurotubules with adluminal cells that were CRX(+). NeuN first appeared in ablumenal neurotubular cells in 1 tumor and was also discovered among neuroblast-appearing cells in another. The third tumor associated with a coloboma was CRX(-) and NeuN(-). CONCLUSIONS: A simple premedullary epithelial monolayer appears to be the fundamental source for the tumor and its multilaminar medullary epithelium. CRX(+) and NeuN(+) cells within the multilayered medullary layer approximate expression patterns similar to those found in retinal development and differentiation. Discovery of these biomarkers in the neoplastic ciliary epithelium in a small number of tumors indicates preliminarily that the most anterior layers of the optic cup have a retained retinal and neuroglial differentiation potentiality. The third case was CRX(-) and NeuN(-) and possibly arose from embryonic pigment epithelium at the edge of the retinochoroidal coloboma. These immunohistochemical findings offer histogenetic and potential diagnostic insights.
Sriram S, Gibson DJ, Robinson P, Pi L, Tuli S, Lewin AS, Schultz G. Assessment of anti-scarring therapies in ex vivo organ cultured rabbit corneas. Exp Eye Res 2014;125:173-82.Abstract
The effects of a triple combination of siRNAs targeting key scarring genes were assessed using an ex vivo organ culture model of excimer ablated rabbit corneas. The central 6 mm diameter region of fresh rabbit globes was ablated to a depth of 155 microns with an excimer laser. Corneas were excised, cultured at the air-liquid interface in defined culture medium supplemented with transforming growth factor beta 1 (TGFB1), and treated with either 1% prednisolone acetate or with 22.5 μM cationic nanoparticles complexed with a triple combination of siRNAs (NP-siRNA) targeting TGFB1, TGFB Receptor (TGFBR2) and connective tissue growth factor (CTGF). Scar formation was measured using image analysis of digital images and levels of smooth muscle actin (SMA) were assessed in ablated region of corneas using qRT-PCR and immunostaining. Ex vivo cultured corneas developed intense haze-like scar in the wounded areas and levels of mRNAs for pro-fibrotic genes were significantly elevated 3-8 fold in wounded tissue compared to unablated corneas. Treatment with NP-siRNA or steroid significantly reduced quantitative haze levels by 55% and 68%, respectively, and reduced SMA mRNA and immunohistostaining. This ex vivo corneal culture system reproduced key molecular patterns of corneal scarring and haze formation generated in rabbits. Treatment with NP-siRNAs targeting key scarring genes or an anti-inflammatory steroid reduced corneal haze and SMA mRNA and protein.
Peacock ZS, Boulos T, Miller JB, Gardiner MF, Chuang S-K, Troulis MJ. Orbital fractures and ocular injury: is a postoperative ophthalmology examination necessary?. J Oral Maxillofac Surg 2014;72(8):1533-40.Abstract
PURPOSE: To determine whether formal ophthalmology evaluation is necessary after operative repair of orbital fractures and the association of an ocular injury to the severity of facial injury. PATIENTS AND METHODS: This was a retrospective cohort study of patients with orbital fractures undergoing operative repair from 2005 to 2013. Subjects were included if they had undergone reconstruction of orbital floor fractures and had data from pre- and postoperative examinations by the oral and maxillofacial surgery and ophthalmology services available. The predictor variables included the service performing the ocular examination (oral and maxillofacial surgery or ophthalmology) and the number of fractures present. The outcome variables were the presence of pre- and postoperative ocular injuries. Logistic regression models were used to determine the relationship of the fracture number to ocular injury. RESULTS: A total of 28 subjects had undergone repair of orbital fractures with preoperative and postoperative examinations performed by both services. Preoperative ocular injuries were found in 17 of the 28 subjects. Those detected by oral and maxillofacial surgeons were limited to changes in visual acuity, pupillary response, and extraocular muscle dysfunction in 11 subjects. Two subjects had new postoperative ocular findings that were considered minor and did not alter management. An increasing number of facial fractures was associated with an increased risk of ocular trauma. Those with 3 or more fractures had an odds ratio of 14.625 (95% confidence interval, 2.191 to 97.612, P = .006) for the presence of ocular injury. CONCLUSIONS: Operative repair of orbital fractures did not lead to new ocular injuries that would change the management. Thus, those without preoperative ocular injuries will not require a formal postoperative ophthalmology examination. However, the subjects with more fractures had an increased likelihood of ocular injuries.
Kwon MY, Bao P, Millin R, Tjan BS. Radial-tangential anisotropy of crowding in the early visual areas. J Neurophysiol 2014;112(10):2413-22.Abstract
Crowding, the inability to recognize an individual object in clutter (Bouma H. Nature 226: 177-178, 1970), is considered a major impediment to object recognition in peripheral vision. Despite its significance, the cortical loci of crowding are not well understood. In particular, the role of the primary visual cortex (V1) remains unclear. Here we utilize a diagnostic feature of crowding to identify the earliest cortical locus of crowding. Controlling for other factors, radially arranged flankers induce more crowding than tangentially arranged ones (Toet A, Levi DM. Vision Res 32: 1349-1357, 1992). We used functional magnetic resonance imaging (fMRI) to measure the change in mean blood oxygenation level-dependent (BOLD) response due to the addition of a middle letter between a pair of radially or tangentially arranged flankers. Consistent with the previous finding that crowding is associated with a reduced BOLD response [Millin R, Arman AC, Chung ST, Tjan BS. Cereb Cortex (July 5, 2013). doi:10.1093/cercor/bht159], we found that the BOLD signal evoked by the middle letter depended on the arrangement of the flankers: less BOLD response was associated with adding the middle letter between radially arranged flankers compared with adding it between tangentially arranged flankers. This anisotropy in BOLD response was present as early as V1 and remained significant in downstream areas. The effect was observed while subjects' attention was diverted away from the testing stimuli. Contrast detection threshold for the middle letter was unaffected by flanker arrangement, ruling out surround suppression of contrast response as a major factor in the observed BOLD anisotropy. Our findings support the view that V1 contributes to crowding.
Cunningham CA, Wolfe JM. The role of object categories in hybrid visual and memory search. J Exp Psychol Gen 2014;143(4):1585-99.Abstract
In hybrid search, observers search for any of several possible targets in a visual display containing distracting items and, perhaps, a target. Wolfe (2012) found that response times (RTs) in such tasks increased linearly with increases in the number of items in the display. However, RT increased linearly with the log of the number of items in the memory set. In earlier work, all items in the memory set were unique instances (e.g., this apple in this pose). Typical real-world tasks involve more broadly defined sets of stimuli (e.g., any "apple" or, perhaps, "fruit"). The present experiments show how sets or categories of targets are handled in joint visual and memory search. In Experiment 1, searching for a digit among letters was not like searching for targets from a 10-item memory set, though searching for targets from an N-item memory set of arbitrary alphanumeric characters was like searching for targets from an N-item memory set of arbitrary objects. In Experiment 2, observers searched for any instance of N sets or categories held in memory. This hybrid search was harder than search for specific objects. However, memory search remained logarithmic. Experiment 3 illustrates the interaction of visual guidance and memory search when a subset of visual stimuli are drawn from a target category. Furthermore, we outline a conceptual model, supported by our results, defining the core components that would be necessary to support such categorical hybrid searches.
Birsner AE, Benny O, D'Amato RJ. The corneal micropocket assay: a model of angiogenesis in the mouse eye. J Vis Exp 2014;(90)Abstract
The mouse corneal micropocket assay is a robust and quantitative in vivo assay for evaluating angiogenesis. By using standardized slow-release pellets containing specific growth factors that trigger blood vessel growth throughout the naturally avascular cornea, angiogenesis can be measured and quantified. In this assay the angiogenic response is generated over the course of several days, depending on the type and dose of growth factor used. The induction of neovascularization is commonly triggered by either basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF). By combining these growth factors with sucralfate and hydron (poly-HEMA (poly(2-hydroxyethyl methacrylate))) and casting the mixture into pellets, they can be surgically implanted in the mouse eye. These uniform pellets slowly-release the growth factors over five or six days (bFGF or VEGF respectively) enabling sufficient angiogenic response required for vessel area quantification using a slit lamp. This assay can be used for different applications, including the evaluation of angiogenic modulator drugs or treatments as well as comparison between different genetic backgrounds affecting angiogenesis. A skilled investigator after practicing this assay can implant a pellet in less than 5 min per eye.
Tang H, Buia C, Madhavan R, Crone NE, Madsen JR, Anderson WS, Kreiman G. Spatiotemporal dynamics underlying object completion in human ventral visual cortex. Neuron 2014;83(3):736-48.Abstract
Natural vision often involves recognizing objects from partial information. Recognition of objects from parts presents a significant challenge for theories of vision because it requires spatial integration and extrapolation from prior knowledge. Here we recorded intracranial field potentials of 113 visually selective electrodes from epilepsy patients in response to whole and partial objects. Responses along the ventral visual stream, particularly the inferior occipital and fusiform gyri, remained selective despite showing only 9%-25% of the object areas. However, these visually selective signals emerged ∼100 ms later for partial versus whole objects. These processing delays were particularly pronounced in higher visual areas within the ventral stream. This latency difference persisted when controlling for changes in contrast, signal amplitude, and the strength of selectivity. These results argue against a purely feedforward explanation of recognition from partial information, and provide spatiotemporal constraints on theories of object recognition that involve recurrent processing.

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