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Turalba A, Payal AR, Gonzalez-Gonzalez LA, Cakiner-Egilmez T, Chomsky AS, Vollman DE, Baze EF, Lawrence M, Daly MK. Cataract Surgery Outcomes in Glaucomatous Eyes: Results From the Veterans Affairs Ophthalmic Surgery Outcomes Data Project. Am J Ophthalmol 2015;160(4):693-701.e1.Abstract

PURPOSE: To compare visual acuity outcomes, vision-related quality of life, and complications related to cataract surgery in eyes with and without glaucoma. DESIGN: Retrospective cohort study. METHODS: Cataract surgery outcomes in cases with and without glaucoma from the Veterans Affairs Ophthalmic Surgical Outcomes Data Project were compared. RESULTS: We identified 608 glaucoma cases and 4306 controls undergoing planned cataract surgery alone. After adjusting for age, pseudoexfoliation, small pupil, prior ocular surgery, and anterior chamber depth, we found that glaucoma cases were more likely to have posterior capsular tear with vitrectomy (odds ratio [OR] 1.8, P = .03) and sulcus intraocular lens placement (OR 1.65, P = .03) during cataract surgery. Glaucoma cases were more likely to have postoperative inflammation (OR 1.73, P < .0001), prolonged elevated intraocular pressure (OR 2.96, P = .0003), and additional surgery within 30 days (OR 1.92, P = .03). Mean best-corrected visual acuity (BCVA) and Visual Function Questionnaire (VFQ) scores significantly improved after cataract surgery in both groups (P < .0001), but there were larger improvements in BCVA (P = .01) and VFQ composite scores (P < .0001) in the nonglaucoma vs the glaucoma group. A total of 3621 nonglaucoma cases (94.1%) had postoperative BCVA 20/40 or better, compared to 466 glaucoma cases (89.6%) (P = .0003). CONCLUSIONS: Eyes with glaucoma are at increased risk for complications and have more modest visual outcomes after cataract surgery compared to eyes without glaucoma. Despite this, glaucoma patients still experience significant improvement in vision-related outcomes after cataract extraction. Further study is needed to explore potential factors that influence cataract surgery outcomes in glaucomatous eyes.

Turalba A, Cakiner-Egilmez T, Payal AR, Gonzalez-Gonzalez LA, Chomsky AS, Vollman DE, Baze EF, Lawrence MG, Daly MK. Outcomes after cataract surgery in eyes with pseudoexfoliation: Results from the Veterans Affairs Ophthalmic Surgery Outcomes Data Project. Can J Ophthalmol 2017;52(1):61-68.Abstract

OBJECTIVE: To compare clinical outcomes of cataract surgery in eyes with and without pseudoexfoliation (PXF). DESIGN: Retrospective deidentified data analysis. PARTICIPANTS: A total of 123 PXF and 4776 non-PXF eyes of patients who underwent cataract surgery. METHODS: We compared data on visual acuity, Visual Function Questionnaire (VFQ)-based quality of life, and complications in PXF and non-PXF eyes from the Veterans Affairs (VA) Ophthalmic Surgery Outcomes Data Project across 5 VA medical centres. RESULTS: Pupillary expansion devices were used in 31 (25.2%) PXF cases and 398 (8.4%) non-PXF cases (p < 0.0001). Capsular tension rings were used in 6 (4.9%) PXF cases and 55 (1.2%) non-PXF cases (p < 0.004). The following complications occurred more frequently in PXF cases: zonular dehiscence without vitrectomy (4 [3.3%] PXF cases vs 40 [0.8%] non-PXF cases p = 0.02), persistent inflammation (28 [24.1%] vs 668 [14.5%]; p = 0.007), and persistent intraocular pressure elevation (5 [4.3%] vs 68 [1.5%]; p = 0.03). Best corrected visual acuity (BCVA) improved in both groups after 1 month, but 87 (83.7%) PXF cases achieved postoperative BCVA better than or equal to 20/40 compared to 3991 (93.8%) non-PXF cases (p = 0.0003). There was no significant difference in the postoperative composite VFQ scores between PXF (82.1 ± 16.9) and non-PXF cases (84.2 ± 16.8, p = 0.09). CONCLUSIONS: Several complications occurred more frequently in the PXF group compared to the non-PXF group, and fewer PXF cases achieved BCVA better than or equal to 20/40. Despite this, both groups experienced similar improvement in vision-related quality of life after cataract surgery.

Turalba AV, Pasquale LR. Hypertensive phase and early complications after Ahmed glaucoma valve implantation with intraoperative subtenon triamcinolone acetonide. Clin Ophthalmol 2014;8:1311-6.Abstract

OBJECTIVE: To evaluate intraoperative subtenon triamcinolone acetonide (TA) as an adjunct to Ahmed glaucoma valve (AGV) implantation. DESIGN: Retrospective comparative case series. PARTICIPANTS: Forty-two consecutive cases of uncontrolled glaucoma undergoing AGV implantation: 19 eyes receiving intraoperative subtenon TA and 23 eyes that did not receive TA. METHODS: A retrospective chart review was performed on consecutive pseudophakic adult patients with uncontrolled glaucoma undergoing AGV with and without intraoperative subtenon TA injection by a single surgeon. Clinical data were collected from 42 eyes and analyzed for the first 6 months after surgery. MAIN OUTCOME MEASURES: Primary outcomes included intraocular pressure (IOP) and number of glaucoma medications prior to and after AGV implantation. The hypertensive phase (HP) was defined as an IOP measurement of greater than 21 mmHg (with or without medications) during the 6-month postoperative period that was not a result of tube obstruction, retraction, or malfunction. Postoperative complications and visual acuity were analyzed as secondary outcome measures. RESULTS: Five out of 19 (26%) TA cases and 12 out of 23 (52%) non-TA cases developed the HP (P=0.027). Mean IOP (14.2±4.6 in TA cases versus [vs] 14.7±5.0 mmHg in non-TA cases; P=0.78), and number of glaucoma medications needed (1.8±1.3 in TA cases vs 1.6±1.1 in the comparison group; P=0.65) were similar between both groups at 6 months. Although rates of serious complications did not differ between the groups (13% in the TA group vs 16% in the non-TA group), early tube erosion (n=1) and bacterial endophthalmitis (n=1) were noted with TA but not in the non-TA group. CONCLUSIONS: Subtenon TA injection during AGV implantation may decrease the occurrence of the HP but does not alter the ultimate IOP outcome and may pose increased risk of serious complications within the first 6 months of surgery.

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Uchino Y, Mauris J, Woodward AM, Dieckow J, Amparo F, Dana R, Mantelli F, Argüeso P. Alteration of galectin-3 in tears of patients with dry eye disease. Am J Ophthalmol 2015;159(6):1027-1035.e3.Abstract
PURPOSE: To investigate the expression, release, and proteolytic degradation of galectin-3 in patients with dry eye disease. DESIGN: Observational case series with a comparison group. METHODS: Tear washes and conjunctival impression cytology specimens were collected through standard procedures from 16 patients with dry eye and 11 age-matched healthy subjects. Galectin-3 content in tears was analyzed by quantitative Western blot, using recombinant galectin-3 protein to generate a calibration curve. The relative expression of galectin-3 and matrix metalloproteinase 9 (MMP9) was evaluated by quantitative polymerase chain reaction. The cleavage of galectin-3 was studied in vitro using activated recombinant MMP9 and protease inhibitors. RESULTS: The concentration of galectin-3 protein in tears, but not galectin-3 expression in conjunctival epithelium, was significantly higher in tears of patients with dry eye (0.38 ng/μg total protein, range 0.04-1.36) compared to healthy subjects (0.12 ng/μg total protein, range 0.00-0.41) (P < .01). By Western blot, an intact (∼28.0 kDa) galectin-3 band was identified in tear samples from healthy subjects, whereas 50% of the dry eye samples were characterized by the additional presence of a partially degraded form (∼25.4 kDa). In our experiments, elevated expression of MMP9 in dry eye subjects correlated with the ability of active MMP9 to cleave galectin-3 from recombinant origin. Interestingly, cleavage of endogenous galectin-3 in tear samples was impaired using a broad-spectrum proteinase inhibitor cocktail, but not the pan-specific MMP inhibitor GM6001, suggesting the presence of proteases other than MMPs in promoting galectin-3 degradation in dry eye. CONCLUSIONS: Our results indicate that release of cellular galectin-3 into tears is associated with epithelial dysfunction in dry eye, and that galectin-3 proteolytic cleavage may contribute to impaired ocular surface barrier function.
Uchino Y, Uchino M, Yokoi N, Dogru M, Kawashima M, Komuro A, Sonomura Y, Kato H, Argüeso P, Kinoshita S, Tsubota K. Impact of Cigarette Smoking on Tear Function and Correlation between Conjunctival Goblet Cells and Tear MUC5AC Concentration in Office Workers. Sci Rep 2016;6:27699.Abstract

The first aim of this study was to clarify whether cigarette smoking affects tear secretion, goblet cell density, and tear MUC5AC concentration. The second purpose was to evaluate the correlations of conjunctival goblet cell density with tear MUC5AC concentration and other ocular surface evaluation factors. This cross-sectional study included 88 office workers. All subjects were required to fill in dry eye and smoking questionnaires, in addition to ocular surface evaluation. Tear wash fluid was collected from inferior fornix, and conjunctival epithelium was obtained by impression cytology. Tear MUC5AC concentration was quantified using enzyme-linked immunoassay, and conjunctival goblet cell density was counted after Periodic-acid Schiff staining. Tear MUC5AC concentration had significant positive correlation with conjunctival goblet cell density (r = 0.181, P = 0.03). In current smokers, Schirmer I test value, goblet cell density and tear MUC5AC concentration were significantly lower than non-smokers. Pack-years of smoking have significant negative correlation with goblet cell density (r = -0.174, P = 0.036) and tear MUC5AC concentration (r = -0.183, P = 0.028). We concluded that smoking might decrease tear secretion, goblet cell density and tear MUC5AC concentration. In addition, MUC5AC concentration in tears depends on goblet cell density in the conjunctiva among office workers.

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.

Uchino Y, Woodward AM, Mauris J, Peterson K, Verma P, Nilsson UJ, Rajaiya J, Argüeso P. Galectin-3 is an amplifier of the interleukin-1β-mediated inflammatory response in corneal keratinocytes. Immunology 2018;154(3):490-499.Abstract
Interleukin-1β (IL-1β) is a potent mediator of innate immunity commonly up-regulated in a broad spectrum of inflammatory diseases. When bound to its cell surface receptor, IL-1β initiates a signalling cascade that cooperatively induces the expression of canonical IL-1 target genes such as IL-8 and IL-6. Here, we present galectin-3 as a novel regulator of IL-1β responses in corneal keratinocytes. Using the SNAP-tag system and digitonin semi-permeabilization, we show that recombinant exogenous galectin-3 binds to the plasma membrane of keratinocytes and is internalized into cytoplasmic compartments. We find that exogenous galectin-3, but not a dominant negative inhibitor of galectin-3 polymerization lacking the N-terminal domain, exacerbates the response to IL-1β by stimulating the secretion of inflammatory cytokines. The activity of galectin-3 could be reduced by a novel d-galactopyranoside derivative targeting the conserved galactoside-binding site of galectins and did not involve interaction with IL-1 receptor 1 or the induction of endogenous IL-1β. Consistent with these observations, we demonstrate that small interfering RNA-mediated suppression of endogenous galectin-3 expression is sufficient to impair the IL-1β-induced secretion of IL-8 and IL-6 in a p38 mitogen-activated protein kinase-independent manner. Collectively, our findings provide a novel role for galectin-3 as an amplifier of IL-1β responses during epithelial inflammation through an as yet unidentified mechanism.
Uchino Y, Woodward AM, Argüeso P. Differential effect of rebamipide on transmembrane mucin biosynthesis in stratified ocular surface epithelial cells. Exp Eye Res 2016;153:1-7.Abstract

Mucins are a group of highly glycosylated glycoproteins responsible for the protection of wet-surfaced epithelia. Recent data indicate that transmembrane mucins differ in their contribution to the protective function of the ocular surface, with MUC16 being the most effective barrier on the apical surface glycocalyx. Here, we investigated the role of the mucoprotective drug rebamipide in the regulation of transmembrane mucin biosynthesis using stratified cultures of human corneal and conjunctival epithelial cells. We find that the addition of rebamipide to corneal, but not conjunctival, epithelial cells increased MUC16 protein biosynthesis. Rebamipide did not affect the levels of MUC1, 4 and 20 compared to control. In these experiments, rebamipide had no effect on the expression levels of Notch intracellular domains, suggesting that the rebamipide-induced increase in MUC16 biosynthesis in differentiated corneal cultures is not regulated by Notch signaling. Overall these findings indicate that rebamipide induces the differential upregulation of MUC16 in stratified cultures of human corneal epithelial cells, which may have implications to the proper restoration of barrier function in ocular surface disease.

Uchiyama E, Faez S, Nasir H, Unizony SH, Plenge R, Papaliodis GN, Sobrin L. Accuracy of the International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) as a research tool for identification of patients with uveitis and scleritis. Ophthalmic Epidemiol 2015;22(2):139-41.Abstract

PURPOSE: To report on the accuracy of the International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) codes for identifying patients with polymyalgia rheumatica (PMR) and concurrent noninfectious inflammatory ocular conditions in a large healthcare organization database. METHODS: Queries for patients with PMR and uveitis or scleritis were executed in two general teaching hospitals' databases. Patients with ocular infections or other rheumatologic conditions were excluded. Patients with PMR and ocular inflammation were identified, and medical records were reviewed to confirm accuracy. RESULTS: The query identified 10,697 patients with the ICD-9-CM code for PMR and 4154 patients with the codes for noninfectious inflammatory ocular conditions. The number of patients with both PMR and noninfectious uveitis or scleritis by ICD-9-CM codes was 66. On detailed review of the charts of these 66 patients, 31 (47%) had a clinical diagnosis of PMR, 43 (65%) had noninfectious uveitis or scleritis, and only 20 (30%) had PMR with concurrent noninfectious uveitis or scleritis confirmed based on clinical notes. CONCLUSIONS: While the use of ICD-9-CM codes has been validated for medical research of common diseases, our results suggest that ICD-9-CM codes may be of limited value for epidemiological investigations of diseases which can be more difficult to diagnose. The ICD-9-CM codes for rarer diseases (PMR, uveitis and scleritis) did not reflect the true clinical problem in a large proportion of our patients. This is particularly true when coding is performed by physicians outside the area of specialty of the diagnosis.

Ung C, Laíns I, Silverman RF, Woods R, Lane AM, Papakostas TD, Husain D, Miller JW, Gragoudas ES, Kim IK, Miller JB. Evaluation of choroidal lesions with swept-source optical coherence tomography. Br J Ophthalmol 2018;Abstract
AIMS: The aim of our study was to image choroidal lesions with swept-source optical coherence tomography (SS-OCT) and to identify the morphological characteristics associated with optimal visualisation. METHODS: This was a prospective, cross-sectional study. Patients with choroidal melanocytic lesions <3 mm in thickness on B-scan ultrasonography were recruited. All participants underwent SS-OCT. On SS-OCT we evaluated qualitative (eg, lesion outline, detection of scleral-choroidal interface and quality of the image) and quantitative (measurement of maximum lesion thickness and the largest basal diameter) parameters. Probability of optimal image quality was examined using ordered logistic regression models. The main outcome measure was quality of the choroidal lesion images on SS-OCT, defined as: optimal, suboptimal or poor. RESULTS: We included 85 choroidal lesions of 82 patients. There were 24 choroidal lesions (29%) for which image quality was classified as optimal, 31 lesions (37%) as suboptimal and 30 lesions (36%) as poor. The factors associated with optimal image quality were distance closer to the fovea (OR 0.76, p<0.001), posterior pole location (OR 3.87, p=0.05), lower ultrasonography thickness (OR 0.44, p=0.04), lighter lesion pigmentation (OR 0.12, p=0.003) and smaller lesion diameter (OR 0.73, p<0.001). In the multivariable analysis, closer distance to the fovea (OR 0.81, p=0.005), lighter lesion pigmentation (OR 0.11, p=0.01) and smaller lesion diameter (OR 0.76, p=0.006) remained statistically significant. CONCLUSION: SS-OCT is useful in imaging most choroidal melanocytic lesions. Image quality is best when the choroidal lesion is closer to the fovea, has a smaller diameter and a lighter choroidal pigmentation.
Ung C, Sanchez AV, Shen L, Davoudi S, Ahmadi T, Navarro-Gomez D, Chen CJ, Hancock H, Penman A, Hoadley S, Consugar M, Restrepo C, Shah VA, Arboleda-Velasquez JF, Sobrin L, Gai X, Kim LA. Whole exome sequencing identification of novel candidate genes in patients with proliferative diabetic retinopathy. Vision Res 2017;Abstract
Rare or novel gene variants in patients with proliferative diabetic retinopathy may contribute to disease development. We performed whole exome sequencing (WES) on patients at the phenotypic extremes of diabetic retinal complications: 57 patients diagnosed with proliferative diabetic retinopathy (PDR) as cases and 13 patients with no diabetic retinopathy despite at least 10years of type 2 diabetes as controls. Thirty-one out of the 57 cases and all 13 controls were from the African American Proliferative Diabetic Retinopathy Study (AA). The rest of the cases were of mixed ethnicities (ME). WES identified 721 candidate genes with rare or novel non-synonymous variants found in at least one case with PDR and not present in any controls. After filtering for genes with null alleles in greater than two cases, 28 candidate genes were identified in our ME cases and 16 genes were identified in our AA cases. Our analysis showed rare and novel variants within these genes that could contribute to the development of PDR, including rare non-synonymous variants in FAM132A, SLC5A9, ZNF600, and TMEM217. We also found previously unidentified variants in VEGFB and APOB. We found that VEGFB, VPS13B, PHF21A, NAT1, ZNF600, PKHD1L1 expression was reduced in human retinal endothelial cells (HRECs) cultured under high glucose conditions. In an exome sequence analysis of patients with PDR, we identified variants in genes that could contribute to pathogenesis. Six of these genes were further validated and found to have reduced expression in HRECs under high glucose conditions, suggestive of an important role in the development of PDR.
Ursea R, De Castro D, Bowen TJ, Chan C-C. The role of conjunctival biopsy in the diagnosis of granulomatosis with polyangiitis. J Ophthalmic Inflamm Infect 2015;5(1):1.Abstract

BACKGROUND: The purpose of this study is to describe a patient who was diagnosed with granulomatosis with polyangiitis based on conjunctival biopsy. This study is a case report and review of the literature. FINDINGS: A 48-year-old Caucasian woman presented with a 2-week history of a left eye peripheral corneal ulcer with adjacent conjunctivitis and a 4-month history of a non-resolving productive cough. Given her elevated serum perinuclear antineutrophil cytoplasmic antibody (P-ANCA) and erythrocyte sedimentation rate (ESR) levels as well as a chest computed topography (CT) that showed bilateral patchy infiltrates, suspicion of limited granulomatosis with polyangiitis with lung and ocular involvement was high. Because bronchoalveolar lavage was nondiagnostic for granulomatous disease, conjunctival biopsy was initially attempted in order to avoid a more invasive lung biopsy. The conjunctival biopsy revealed mixed subacute inflammatory mediators and vasculitis consistent with granulomatosis with polyangiitis. CONCLUSIONS: Conjunctival biopsy may be a valuable, minimally invasive method for diagnosing systemic granulomatosis with polyangiitis.

Utheim TP, Islam R, Fostad IG, Eidet JR, Sehic A, Olstad OK, Dartt DA, Messelt EB, Griffith M, Pasovic L. Storage Temperature Alters the Expression of Differentiation-Related Genes in Cultured Oral Keratinocytes. PLoS One 2016;11(3):e0152526.Abstract
PURPOSE: Storage of cultured human oral keratinocytes (HOK) allows for transportation of cultured transplants to eye clinics worldwide. In a previous study, one-week storage of cultured HOK was found to be superior with regard to viability and morphology at 12°C compared to 4°C and 37°C. To understand more of how storage temperature affects cell phenotype, gene expression of HOK before and after storage at 4°C, 12°C, and 37°C was assessed. MATERIALS AND METHODS: Cultured HOK were stored in HEPES- and sodium bicarbonate-buffered Minimum Essential Medium at 4°C, 12°C, and 37°C for one week. Total RNA was isolated and the gene expression profile was determined using DNA microarrays and analyzed with Partek Genomics Suite software and Ingenuity Pathway Analysis. Differentially expressed genes (fold change > 1.5 and P < 0.05) were identified by one-way ANOVA. Key genes were validated using qPCR. RESULTS: Gene expression of cultures stored at 4°C and 12°C clustered close to the unstored control cultures. Cultures stored at 37°C displayed substantial change in gene expression compared to the other groups. In comparison with 12°C, 2,981 genes were differentially expressed at 37°C. In contrast, only 67 genes were differentially expressed between the unstored control and the cells stored at 12°C. The 12°C and 37°C culture groups differed most significantly with regard to the expression of differentiation markers. The Hedgehog signaling pathway was significantly downregulated at 37°C compared to 12°C. CONCLUSION: HOK cultures stored at 37°C showed considerably larger changes in gene expression compared to unstored cells than cultured HOK stored at 4°C and 12°C. The changes observed at 37°C consisted of differentiation of the cells towards a squamous epithelium-specific phenotype. Storing cultured ocular surface transplants at 37°C is therefore not recommended. This is particularly interesting as 37°C is the standard incubation temperature used for cell culture.
Utheim O, Islam R, Lyberg T, Roald B, Eidet JR, de la Paz MF, Dartt DA, Raeder S, Utheim TP. Serum-free and xenobiotic-free preservation of cultured human limbal epithelial cells. PLoS One 2015;10(3):e0118517.Abstract

AIM/PURPOSE OF THE STUDY: To develop a one-week storage method, without serum and xenobiotics, that would maintain cell viability, morphology, and phenotype of cultured human limbal epithelial sheets. MATERIALS AND METHODS: Human limbal explants were cultured on intact human amniotic membranes for two weeks. The sheets were stored in a hermetically sealed container at 23°C in either a serum-free medium with selected animal serum-derived compounds (Quantum 286) or a xenobiotic-free medium (Minimal Essential Medium) for 4 and 7 days. Stored and non-stored cultures were analyzed for cell viability, amniotic membrane and epithelial sheet thickness, and a panel of immunohistochemical markers for immature cells (ΔNp63α, p63, Bmi-1, C/EBP∂, ABCG2 and K19), differentiated cells (K3 and Cx43), proliferation (PCNA), and apoptosis (Caspase-3). RESULTS: The cell viability of the cultures was 98 ± 1% and remained high after storage. Mean central thickness of non-stored limbal epithelial sheets was 23 ± 3 μm, and no substantial loss of cells was observed after storage. The non-stored epithelial sheets expressed a predominantly immature phenotype with ΔNp63α positivity of more than 3% in 9 of 13 cultures. After storage, the expression of ABCG2 and C/EBP∂ was reduced for the 7 day Quantum 286-storage group; (P = 0.04), and Bmi-1 was reduced after 4 day Quantum 286-storage; (P = 0.02). No other markers varied significantly. The expression of differentiation markers was unrelated to the thickness of the epithelia and amniotic membrane, apart from ABCG2, which correlated negatively with thickness of limbal epithelia (R = -0.69, P = 0.01) and ΔNp63α, which correlated negatively with amniotic membrane thickness (R = -0.59, P = 0.03). CONCLUSION: Limbal epithelial cells cultured from explants on amniotic membrane can be stored at 23°C in both serum-free and xenobiotic-free media, with sustained cell viability, ultrastructure, and ΔNp63α-positivity after both 4 and 7 days.

Utheim OA, Lyberg T, Eidet JR, Raeder S, Sehic A, Roald B, Messelt E, de la Paz MF, Dartt DA, Utheim TP. Effect of Transportation on Cultured Limbal Epithelial Sheets for Worldwide Treatment of Limbal Stem Cell Deficiency. Sci Rep 2018;8(1):10502.Abstract
Limbal stem cell deficiency can be treated with transplantation of cultured human limbal epithelial cells (LEC). It can be advantageous to produce LEC in centralized labs and thereafter ship them to eye clinics. The present study used transport simulations of LEC to determine if vigorous shaking during transport altered the viability, morphology and phenotype during a 4 day-long storage of LEC with a previously described serum-free storage method. Inserts with LEC cultured on amniotic membranes were sutured to caps inside air-tight containers with generous amounts of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)-buffered minimal essential medium (MEM). The containers were distributed among the following testing conditions: 6 hours with full containers, 36 hours with full containers, 36 hours with container three quarters full of medium, and 36 hours with container full of medium containing a shear-protecting agent (Pluronic-F68). Compared to stored, but non-transported controls, no statistically significant changes in viability and immunohistochemical staining were observed. The epithelial sheets remained intact. However, an air-liquid interface in the containers reduced the number of desmosomes and hemi-desmosomes compared to the controls. In conclusion, cultured LEC sheets appear to endure vigorous shaking for at least 36 hours if the container is full.
Utochkin IS, Wolfe JM. Visual search for changes in scenes creates long-term, incidental memory traces. Atten Percept Psychophys 2018;80(4):829-843.Abstract
Humans are very good at remembering large numbers of scenes over substantial periods of time. But how good are they at remembering changes to scenes? In this study, we tested scene memory and change detection two weeks after initial scene learning. In Experiments 1-3, scenes were learned incidentally during visual search for change. In Experiment 4, observers explicitly memorized scenes. At test, after two weeks observers were asked to discriminate old from new scenes, to recall a change that they had detected in the study phase, or to detect a newly introduced change in the memorization experiment. Next, they performed a change detection task, usually looking for the same change as in the study period. Scene recognition memory was found to be similar in all experiments, regardless of the study task. In Experiment 1, more difficult change detection produced better scene memory. Experiments 2 and 3 supported a "depth-of-processing" account for the effects of initial search and change detection on incidental memory for scenes. Of most interest, change detection was faster during the test phase than during the study phase, even when the observer had no explicit memory of having found that change previously. This result was replicated in two of our three change detection experiments. We conclude that scenes can be encoded incidentally as well as explicitly and that changes in those scenes can leave measurable traces even if they are not explicitly recalled.
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Vajaranant TS, Ray RM, Pasquale LR, Mares JA, Ritch R, Gower EW, Haan MN, Jackson RD, Maki PM. Racial Differences in the Effects of Hormone Therapy on Incident Open-Angle Glaucoma in a Randomized Trial. Am J Ophthalmol 2018;Abstract
PURPOSE: We conducted a secondary analysis of a randomized, placebo-controlled trial to test if hormone therapy (HT) altered the risk of open-angle glaucoma (OAG), and if the risk reduction varied by race. DESIGN: Secondary analysis of randomized controlled trial data Methods: We linked Medicare claims data to 25,535 women in the Women's Health Initiative. Women without a uterus were randomized to receive either oral conjugated equine estrogens (CEE 0.625mg/day) or placebo, and women with a uterus received oral CEE and medroxyprogesterone acetate (CEE 0.625mg/day+MPA 2.5mg/day) or placebo. We used Cox proportional hazards models to calculate hazard ratios (HR) and 95% confidence interval. RESULTS: After excluding women with prevalent glaucoma or without claims for eye-care provider visits, the final analysis included 8,102 women (mean age=68.5±4.8 years). The OAG incidence was 7.6% (mean follow-up=11.5±5.2 years; mean HT duration=4.4±2.3 years). Increased age (p-trend=0.01) and African-American race (HR=2.69, 95%CI=2.13 to 3.42; Caucasian as a reference) were significant risk factors for incident OAG. We found no overall benefit of HT in reducing incident OAG (HR=1.01, 95%CI=0.79-1.29 in the CEE trial, and HR=1.05, 95%CI=0.85-1.29 in the CEE+MPA trial). However, race modified the relationship between CEE use and OAG risk (p-interaction=0.01), and risk was reduced in African-American women treated with CEE (HR=0.49, 95%CI=0.27-0.88), compared to placebo. Race did not modify the relation between CEE+MPA use and OAG risk (p-interaction=0.68). CONCLUSIONS: Analysis suggests that HT containing estrogen, but not a combination of estrogen and progesterone, reduces the risk of incident OAG among African-American women. Further investigation is needed.
Valdez CN, Arboleda-Velasquez JF, Amarnani DS, Kim LA, D'Amore PA. Retinal microangiopathy in a mouse model of inducible mural cell loss. Am J Pathol 2014;184(10):2618-26.Abstract

Diabetes can lead to vision loss because of progressive degeneration of the neurovascular unit in the retina, a condition known as diabetic retinopathy. In its early stages, the pathology is characterized by microangiopathies, including microaneurysms, microhemorrhages, and nerve layer infarcts known as cotton-wool spots. Analyses of postmortem human retinal tissue and retinas from animal models indicate that degeneration of the pericytes, which constitute the outer layer of capillaries, is an early event in diabetic retinopathy; however, the relative contribution of specific cellular components to the pathobiology of diabetic retinopathy remains to be defined. We investigated the phenotypic consequences of pericyte death on retinal microvascular integrity by using nondiabetic mice conditionally expressing a diphtheria toxin receptor in mural cells. Five days after administering diphtheria toxin in these adult mice, changes were observed in the retinal vasculature that were similar to those observed in diabetes, including microaneurysms and increased vascular permeability, suggesting that pericyte cell loss is sufficient to trigger retinal microvascular degeneration. Therapies aimed at preventing or delaying pericyte dropout may avoid or attenuate the retinal microangiopathy associated with diabetes.

Valentino MD, McGuire AM, Rosch JW, Bispo PJM, Burnham C, Sanfilippo CM, Carter RA, Zegans ME, Beall B, Earl AM, Tuomanen EI, Morris TW, Haas W, Gilmore MS. Unencapsulated Streptococcus pneumoniae from conjunctivitis encode variant traits and belong to a distinct phylogenetic cluster. Nat Commun 2014;5:5411.Abstract

Streptococcus pneumoniae, an inhabitant of the upper respiratory mucosa, causes respiratory and invasive infections as well as conjunctivitis. Strains that lack the capsule, a main virulence factor and the target of current vaccines, are often isolated from conjunctivitis cases. Here we perform a comparative genomic analysis of 271 strains of conjunctivitis-causing S. pneumoniae from 72 postal codes in the United States. We find that the vast majority of conjunctivitis strains are members of a distinct cluster of closely related unencapsulated strains. These strains possess divergent forms of pneumococcal virulence factors (such as CbpA and neuraminidases) that are not shared with other unencapsulated nasopharyngeal S. pneumoniae. They also possess putative adhesins that have not been described in encapsulated pneumococci. These findings suggest that the unencapsulated strains capable of causing conjunctivitis utilize a pathogenesis strategy substantially different from that described for S. pneumoniae at other infection sites.

Van Tyne D, Ciolino JB, Wang J, Durand ML, Gilmore MS. Novel Phagocytosis-Resistant Extended-Spectrum β-Lactamase-Producing Escherichia coli From Keratitis. JAMA Ophthalmol 2016;134(11):1306-1309.Abstract

Importance: Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli are highly antibiotic resistant, and primary ocular infection by ESBL E coli has rarely been reported. A novel mutation conferring phagocytosis resistance would position a strain well to infect the cornea. Observations: A woman with recurrent keratitis presented with a corneal ulcer, which was culture positive for ESBL E coli. Resistant to nearly all other antimicrobials, the infection was treated with amikacin and polymyxin B-trimethoprim, and the ulcer resolved over 3 weeks. Analysis of the E coli genome showed it to belong to multilocus sequence type 131 (ST131). This isolate was found to possess a novel deletion in yrfF, an essential regulator of bacterial capsule synthesis. Disruption of yrfF, which confers mucoidy and increased virulence, has not been previously observed in ESBL E coli from any infection site. This novel variant was experimentally proven to cause the mucoid phenotype, and corresponding resistance to phagocytic killing. Conclusions and Relevance: Increased resistance to immune clearance in an ESBL E coli lineage already known for its virulence is an unsettling development. This phenotype, which likely positioned it as an unusual cause of corneal ulcer, can be easily recognized in the laboratory, which should help limit its spread.

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