Cornea

PURPOSE: To compare the corneal stromal riboflavin concentration and distribution using 2 transepithelial corneal crosslinking (CXL) systems. SETTING: Absorption Systems, San Diego, California, USA. DESIGN: Experimental study. METHODS: The stromal riboflavin concentration of 2 transepithelial CXL systems was compared in rabbit eyes in vivo. The systems were the Paracel/Vibex Xtra, comprising riboflavin 0.25% solution containing TRIS and ethylenediaminetetraacetic acid and an isotonic solution of riboflavin 0.25%, (Group 1) and the CXLO system (Group 2). Manufacturers' Instructions For Use were followed. The intensity of riboflavin fluorescence by slitlamp observation 10, 15, and 20 minutes after instillation was graded on a scale of 0 to 5. The animals were humanely killed and the corneal stromal samples analyzed with liquid chromatography and mass spectrometry. RESULTS: The mean riboflavin fluorescence intensity grades in Group 1 (4 eyes) were 3.8, 4.8, and 4.8 at 10, 15, and 20 minutes, respectively. The mean grades in Group 2 (3 eyes) were 2.0, 2.3, and 2.0, respectively. The riboflavin distribution was uniform in Group 1 but not in Group 2. The mean riboflavin concentration by liquid chromatography and mass spectrometry was 27.0 μg/g stromal tissue in Group 1 and 6.7 μg/g in Group 2. A stromal riboflavin concentration theoretically adequate for CXL, 15 μg/g, was achieved in all eyes in Group 1 and no eyes in Group 2. Slitlamp grading correlated well with liquid chromatography and mass spectrometry concentration (R = 0.940). CONCLUSIONS: The system used in Group 1 produced corneal riboflavin concentrations that were theoretically adequate for effective transepithelial CXL (≥15 μg/g), while the system in Group 2 did not. Slitlamp grading successfully estimated the corneal riboflavin concentration and can be used to ensure an adequate concentration of riboflavin in the cornea for transepithelial CXL.

PURPOSE: To determine host and pathogen factors predictive of outcomes in a large clinical cohort with keratoconjunctivitis. DESIGN: Retrospective analyses of the clinical and molecular data from a randomized, controlled, masked trial for auricloscene for keratoconjunctivitis (NVC-422 phase IIB, NovaBay; clinicaltrials.gov identifier, NCT01877694). PARTICIPANTS: Five hundred participants from United States, India, Brazil, and Sri Lanka with clinical diagnosis of keratoconjunctivitis and positive rapid test results for adenovirus. METHODS: Clinical signs and symptoms and bilateral conjunctival swabs were obtained on days 1, 3, 6, 11, and 18. Polymerase chain reaction (PCR) analysis was performed to detect and quantify adenovirus in all samples. Regression models were used to evaluate the association of various variables with keratoconjunctivitis outcomes. Time to resolution of each symptom or sign was assessed by adenoviral species with Cox regression. MAIN OUTCOME MEASURES: The difference in composite scores of clinical signs between days 1 and 18, mean visual acuity change between days 1 and 18, and time to resolution of each symptom or sign. RESULTS: Of 500 participants, 390 (78%) showed evidence of adenovirus by PCR. Among adenovirus-positive participants, adenovirus D species was most common (63% of total cases), but a total of 4 species and 21 different types of adenovirus were detected. Adenovirus D was associated with more severe signs and symptoms, a higher rate of subepithelial infiltrate development, and a slower decline in viral load compared with all other adenovirus species. The clinical courses of all patients with non-adenovirus D species infection and adenovirus-negative keratoconjunctivitis were similar. Mean change in visual acuity between days 1 and 18 was a gain of 1.9 letters; worse visual outcome was associated with older age. CONCLUSIONS: A substantial proportion of keratoconjunctivitis is not associated with a detectable adenovirus. The clinical course of those with adenovirus D keratoconjunctivitis is significantly more severe than those with non-adenovirus D species infections or adenovirus-negative keratoconjunctivitis; high viral load at presentation and non-United States origin of participants is associated with poorer clinical outcome.

PURPOSE: The purpose of this study was to examine the effect of corneal biomechanical properties on intraocular pressure (IOP) measurements obtained using a rebound self-tonometer (Icare HOME) compared with Goldmann applanation tonometry (GAT). METHODS: An observational study of 100 patients with glaucoma or ocular hypertension. All had a comprehensive ophthalmic examination and standard automated perimetry. IOP was assessed by GAT, Icare HOME and Ocular Response Analyzer, which was also used to assess corneal hysteresis (CH) and corneal resistance factor (CRF). Central corneal thickness (CCT) was recorded. RESULTS: Mean (±SD) IOP measurements were 14.3±3.9 and 11.7±4.7 mm Hg using GAT and Icare HOME, respectively. Average CCT, CRF, and CH were 534.5±37.3 μm, 9.0±1.7 mm Hg, and 9.4±1.5 mm Hg, respectively. The mean difference between Icare HOME and GAT was -2.66±3.13 mm Hg, with 95% limits of agreement of -8.80 to 3.48 mm Hg, however, there was evidence of proportional bias. There was negative correlation between IOP and CH [5.17 mm Hg higher Icare HOME IOP (P=0.041, R=0.029) and 7.23 mm Hg higher GAT IOP (P=0.008, R=0.080) for each 10 mm Hg lower CH], whereas thinner CCT was significantly associated with lower IOP (P<0.001, R=0.14 for Icare HOME and P<0.001, R=0.08 for GAT). In multivariable analysis, although CRF and CH remained associated with IOP measured using either GAT or Icare HOME, CCT was no longer significant. CONCLUSION: IOP measurements obtained using a self-tonometer, similar to GAT, were more influenced by overall corneal biomechanics than CCT.

Tissue adhesives are gaining popularity in ophthalmology, as they could potentially reduce the complications associated with current surgical methods. An ideal tissue adhesive should have superior tensile strength, be non-toxic and anti-inflammatory, improve efficiency and be cost-effective. Both synthetic and biological glues are available. The primary synthetic glues include cyanoacrylate and the recently introduced polyethylene glycol (PEG) derivatives, while most biological glues are composed of fibrin. Cyanoacrylate has a high tensile strength, but rapidly polymerises upon contact with any fluid and has been associated with histotoxicity. Fibrin induces less toxic and inflammatory reactions, and its polymerisation time can be controlled. Tensile strength studies have shown that fibrin is not as strong as cyanoacrylate. While more research is needed, PEG variants currently appear to have the most promise. These glues are non-toxic, strong and time-effective. Through MEDLINE and internet searches, this paper presents a systematic review of the current applications of surgical adhesives to corneal, glaucoma, retinal, cataract and strabismus surgeries. Our review suggests that surgical adhesives have promise to reduce problems in current ophthalmic surgical procedures.

PURPOSE: To compare the response of dry eye disease (DED) to treatment with topical steroid in patients with and without graft-vs-host disease (GVHD). DESIGN: Post hoc analysis of a double-masked, randomized clinical trial. METHODS: This single-center study included 42 patients with moderate-to-severe DED associated with (n = 21) or without (n = 21) chronic GVHD. In each group, patients received either loteprednol etabonate 0.5% ophthalmic suspension or artificial tears twice daily for 4 weeks. Clinical data, including Ocular Surface Disease Index (OSDI) questionnaire, corneal fluorescein staining (CFS), conjunctival lissamine green staining, tear break-up time (TBUT), and Schirmer test, were evaluated before and after treatment. RESULTS: There were no significant differences in signs and symptoms of DED between the groups at baseline. In non-GVHD patients receiving loteprednol treatment, the average OSDI score decreased by 34% from 49.5 ± 5.9 to 32.6 ± 4.8 (mean ± standard error of the mean, P = .001) and the average CFS score decreased by 41% from 5.6 ± 0.6 to 3.3 ± 0.9 (P = .02). On the other hand, loteprednol treatment in GVHD patients resulted in minimal change in OSDI (59.2 ± 6.7 to 61.1 ± 7.1, 3% increase, P = .66) and CFS (5.5 ± 0.5 to 5.3 ± 1.1, 4% decrease, P = .85) scores. Treatment with artificial tears resulted in 22% decrease of OSDI (P = .10) and 32% decrease of CFS (P = .02) scores in non-GVHD patients, and had minimal effect in patients with GVHD. CONCLUSIONS: DED patients with ocular GVHD have a less favorable response to a low-dose topical steroid regimen compared with those without ocular GVHD even with similar baseline disease severity.

PURPOSE: To evaluate occurrence of subtarsal fibrosis in patients with graft-vs-host disease (GVHD) and to determine its association with ocular surface epitheliopathy. DESIGN: Cross-sectional study. METHODS: We enrolled 40 patients with moderate or severe dry eye disease, including 20 patients with chronic ocular GVHD and 20 patients without (as the control group). All patients had a comprehensive ophthalmic assessment including evaluation for subtarsal fibrosis, corneal and conjunctival staining, tear break-up time (TBUT), and Schirmer test. Furthermore, meibomian gland drop-out area and densities of epithelial and stromal immune cells were measured using meibography and in vivo confocal microscopy, respectively. RESULTS: Subtarsal fibrosis was not seen in any eye of the non-GVHD group. However, 16 eyes (40%) of 10 patients (50%) in the GVHD group had subtarsal fibrosis (P < .001) with an average involvement of 28.9% ± 13.7% of the tarsal area. Fibrosis was more frequent in the upper lids (35%) than in the lower lids (5%). Regression analyses showed that corneal fluorescein staining was significantly associated with the extent of fibrosis (P < .001, β = 0.14) and TBUT (P < .001, β = -0.53) but not with other clinical or imaging parameters. Conjunctival lissamine green staining also had a statistically significant association with the extent of fibrosis (P = .04, β = 0.12) but not other clinical or imaging parameters. Eyes with subtarsal fibrosis had a more severe ocular surface epitheliopathy compared with eyes without fibrosis. CONCLUSIONS: Subtarsal fibrosis is present in a significant percentage of patients with chronic ocular GVHD, likely contributing to the ocular surface damage in these patients.

Glycosylation is a major form of enzymatic modification of organic molecules responsible for multiple biological processes in an organism. The biosynthesis of glycans is controlled by a series of glycosyltransferases, glycosidases and glycan-modifying enzymes that collectively assemble and process monosaccharide moieties into a diverse array of structures. Many studies have provided insight into various pathways of glycosylation at the ocular surface, such as those related to the biosynthesis of mucin-type -glycans and -glycans on proteins, but many others still remain largely unknown. This review provides an overview of the different classes of glycans described at the ocular surface focusing on their biosynthetic pathways and biological relevance. A precise understanding of these pathways under physiological and pathological conditions could help identify biomarkers and novel targets for therapeutic intervention.

Fuchs endothelial corneal dystrophy (FECD) is a genetic and oxidative stress disorder of post-mitotic human corneal endothelial cells (HCEnCs), which normally exhibit hexagonal shape and form a compact monolayer compatible with normal corneal functioning and clear vision. FECD is associated with increased DNA damage, which in turn leads to HCEnC loss, resulting in the formation rosettes and aberrant extracellular matrix (ECM) deposition in the form of pro-fibrotic guttae. Since the mechanism of ECM deposition in FECD is currently unknown, we aimed to investigate the role of endothelial-mesenchymal transition (EMT) in FECD using a previously established cellular in vitro model that recapitulates the characteristic rosette formation, by employing menadione (MN)-induced oxidative stress. We demonstrate that MN treatment alone, or a combination of MN and TGF-β1 induces reactive oxygen species (ROS), cell death, and EMT in HCEnCs during rosette formation, resulting in upregulation of EMT- and FECD-associated markers such as Snail1, N-cadherin, ZEB1, and transforming growth factor-beta-induced (TGFβI), respectively. Additionally, FECD ex vivo specimens displayed a loss of organized junctional staining of plasma membrane-bound N-cadherin, with corresponding increase in fibronectin and Snail1 compared to ex vivo controls. Addition of N-acetylcysteine (NAC) downregulated all EMT markers and abolished rosette formation. Loss of NQO1, a metabolizing enzyme of MN, led to greater increase in intracellular ROS levels as well as a significant upregulation of Snail1, fibronectin, and N-cadherin compared to normal cells, indicating that NQO1 regulates Snail1-mediated EMT. This study provides first line evidence that MN-induced oxidative stress leads to EMT in corneal endothelial cells, and the effect of which is further potentiated when redox cycling activity of MN is enhanced by the absence of NQO1. Given that NAC inhibits Snail-mediated EMT, this may be a potential therapeutic intervention for FECD.

Cosmetic products, such as mascara, eye shadow, eyeliner and eye makeup remover are used extensively to highlight the eyes or clean the eyelids, and typically contain preservatives to prevent microbial growth. These preservatives include benzalkonium chloride (BAK) and formaldehyde (FA)-releasing preservatives. We hypothesize that these preservatives, at concentrations (BAK = 1 mg/ml; FA = 0.74 mg/ml) approved for consumer use, are toxic to human ocular surface and adnexal cells. Accordingly, we tested the influence of BAK and FA on the morphology, survival, and proliferation and signaling ability of immortalized human meibomian gland (iHMGECs), corneal (iHCECs) and conjunctival (iHConjECs) epithelial cells. iHMGECs, iHCECs and iHConjECs were cultured with different concentrations of BAK (5 μg/ml to 0.005 μg/ml) or FA (1 mg/ml to 1 μg/ml) under basal, proliferating or differentiating conditions up to 7 days. We used low BAK levels, because we found that 0.5 mg/ml and 50 μg/ml BAK killed iHMGECs within 1 day after a 15 min exposure. Experimental procedures included analyses of cell appearance, cell number, and neutral lipid content (LipidTox), lysosome accumulation (LysoTracker) and AKT signaling in all 3 cell types. Our results demonstrate that BAK and FA cause dose-dependent changes in the morphology, survival, proliferation and AKT signaling of iHMGECs, iHCECs and iHConjECs. Many of the concentrations tested induced cell atrophy, poor adherence, decreased proliferation and death, after 5 days of exposure. Cellular signaling, as indicated by AKT phosphorylation after 15 (FA) or 30 (BAK) minutes of treatment, was also reduced in a dose-dependent fashion in all 3 cell types, irrespective of whether cells had been cultured under proliferating or differentiating conditions. Our results support our hypothesis and demonstrate that the cosmetic preservatives, BAK and FA, exert many toxic effects on cells of the ocular surface and adnexa.

The purpose of this study is to determine neural, vascular, protein secretion, and cellular signaling changes with disease progression in lacrimal glands of the thrombospondin-1 (TSP-1) mouse model of dry eye compared to C57BL/6 wild-type (WT) mice. Neural innervation was reduced in TSP-1 lacrimal glands compared to WT controls, whereas the number of blood vessels was increased. Intracellular Ca stores and the amount of lysosomes, mitochondria, and secretory granules, but not the endoplasmic reticulum, were reduced in TSP-1 compared to WT acini at 12 weeks of age. Ex vivo high KCl-evoked secretion was decreased in TSP-1 compared to WT lacrimal gland tissue pieces. The α-adrenergic agonist-stimulated response was increased in TSP-1 at 4 and 24 weeks but decreased at 12 weeks, and the ATP and MeSATP-stimulated peak [Ca] responses were decreased at 24 weeks. These changes were observed prior to the appearance of mononuclear infiltrates. We conclude that in the lacrimal gland the absence of TSP-1: injures peripheral nerves; blocks efferent nerve activation; decreases protein secretion; and alters intracellular Ca stores. Through these effects the absence of TSP-1 leads to disruption of ocular surface homeostasis and development of dry eye.

PURPOSE: To determine the demographic features of patients affected by limbal stem cell deficiency (LSCD), and to identify the underlying causes of LSCD DESIGN: Retrospective, multi-center case series SETTING: Two large tertiary care ophthalmology hospitals SUBJECTS: Patients with a diagnosis of LSCD presenting from January 1, 2005 to December 31, 2014 METHODS: Records of patients with a clinical diagnosis of LSCD were reviewed. Demographic details and clinical features at presentation, as well as the underlying cause of LSCD (if identified) were noted. Descriptive statistical analysis and chart preparation were done. MAIN OUTCOME MEASURES: Type of LSCD (unilateral or bilateral), age and sex of patients, extent of LSCD (clock hours of limbus involved) and underlying cause of LSCD RESULTS: We found 1331 patients with LSCD in the ten year period under study. Unilateral LSCD was more common (791 patients) than bilateral LSCD (540 patients). Out of 1331 patients, 875 (65.74%) were male. The median age of patients was 24 years. Extent of LSCD could be determined in 1849 eyes, of which 1239 eyes (67%) had total LSCD. The underlying cause of LSCD could be identified in 1512 eyes. In cases of unilateral LSCD, ocular surface burns was the commonest identifiable cause ( 83.73%). The leading identifiable causes of bilateral LSCD were ocular surface burns (29.95%), allergic conjunctivitis (29.48%), Stevens Johnson syndrome (SJS) or toxic epidermal necrolysis (TEN) (23.11%), aniridia (9.43%) and mucous membrane pemphigoid (3.54%). Lime ("chuna") injury was responsible for ocular surface burns in 352 (62.08%) out of 567 cases in which the agent was identified. CONCLUSIONS: In our study, unilateral LSCD was more common than bilateral LSCD. Young males were commonly affected, with a majority of eyes suffering from total LSCD. Overall, ocular surface burns are the leading cause of LSCD.Unilateral and bilateral LSCD had a markedly different distribution of causes, necessitating different approaches to management.

PURPOSE: To assess the relationship of forward and backward scattering and corneal higher-order aberrations (HOAs) with corrected distance visual acuity (CDVA) after penetrating keratoplasty (PK). METHODS: This retrospective study comprised 25 eyes of 25 consecutive patients who underwent PK using the VisuMax femtosecond laser system and age-matched 25 eyes of 25 healthy subjects. We quantitatively assessed objective scattering index (OSI) using the double-pass instrument (OQAS II, Visiometrics), corneal densitometry (CD) and corneal HOAs with the Scheimpflug rotating camera (Pentacam HR, Oculus) 1 year postoperatively. RESULTS: The OSI, CD, and corneal HOAs were significantly larger in the PK group than those in the control group (p ≤ 0.011). We found significant correlations of logMAR CDVA with the OSI (r = 0.477, p = 0.016), and with the anterior, posterior, and total corneal HOAs of the central 4-mm zone (anterior: r = 0.573, p = 0.003, posterior: r = 0.596, p = 0.002, total: r = 0.472, p = 0.017), but no significant association with the CD of the 0-2 mm zone at any layers (anterior: r = 0.236, p = 0.257, center: r = 0.139, p = 0.506, posterior: r = 0.073, p = 0.728, total: r = 0.212, p = 0.308). Similar results were obtained when the analysis was repeated with corneal HOAs of the central 6-mm zone and CDs in 2-6 mm zone. CONCLUSIONS: Our pilot study demonstrated that the postoperative CDVA was significantly correlated with OSI and corneal HOAs, but not with backward scattering in post-PK eyes, suggesting that OSI as well as corneal HOAs plays an essential role in postoperative visual performance after PK.

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.