Cornea

Woodward AM, Mauris J, Argüeso P. Binding of transmembrane mucins to galectin-3 limits herpesvirus 1 infection of human corneal keratinocytes. J Virol 2013;87(10):5841-7.Abstract
Epithelial cells lining mucosal surfaces impose multiple barriers to viral infection. At the ocular surface, the carbohydrate-binding protein galectin-3 maintains barrier function by cross-linking transmembrane mucins on the apical glycocalyx. Despite these defense mechanisms, many viruses have evolved to exploit fundamental cellular processes on host cells. Here, we use affinity assays to show that herpes simplex virus type 1 (HSV-1), but not HSV-2, binds human galectin-3. Knockdown of galectin-3 in human corneal keratinocytes by small interfering RNA significantly impaired HSV-1 infection, but not expression of nectin-1, indicating that galectin-3 is a herpesvirus entry mediator. Interestingly, exposure of epithelial cell cultures to transmembrane mucin isolates decreased viral infectivity. Moreover, HSV-1 failed to elute the biological counterreceptor MUC16 from galectin-3 affinity columns, suggesting that association of transmembrane mucins to galectin-3 provides protection against viral infection. Together, these results indicate that HSV-1 exploits galectin-3 to enhance virus attachment to host cells and support a protective role for transmembrane mucins under physiological conditions by masking viral entry mediators on the epithelial glycocalyx.
Yousuf MA, Zhou X, Mukherjee S, Chintakuntlawar AV, Lee JY, Ramke M, Chodosh J, Rajaiya J. Caveolin-1 associated adenovirus entry into human corneal cells. PLoS One 2013;8(10):e77462.Abstract
The cellular entry of viruses represents a critical area of study, not only for viral tropism, but also because viral entry dictates the nature of the immune response elicited upon infection. Epidemic keratoconjunctivitis (EKC), caused by viruses within human adenovirus species D (HAdV-D), is a severe, ocular surface infection associated with corneal inflammation. Clathrin-mediated endocytosis has previously been shown to play a critical role in entry of other HAdV species into many host cell types. However, HAdV-D endocytosis into corneal cells has not been extensively studied. Herein, we show an essential role for cholesterol rich, lipid raft microdomains and caveolin-1, in the entry of HAdV-D37 into primary human corneal fibroblasts. Cholesterol depletion using methyl-β-cyclodextrin (MβCD) profoundly reduced viral infection. When replenished with soluble cholesterol, the effect of MβCD was reversed, allowing productive viral infection. HAdV-D37 DNA was identified in caveolin-1 rich endosomal fractions after infection. Src kinase activity was also increased in caveolin-1 rich endosomal fractions after infection, and Src phosphorylation and CXCL1 induction were both decreased in caveolin-1-/- mice corneas compared to wild type mice. siRNA knock down of caveolin-1 in corneal cells reduced chemokine induction upon viral infection, and caveolin-1-/- mouse corneas showed reduced cellular entry of HAdV-D37. As a control, HAdV-C2, a non-corneal pathogen, appeared to utilize the caveolar pathway for entry into A549 cells, but failed to infect corneal cells entirely, indicating virus and cell specific tropism. Immuno-electron microscopy confirmed the presence of caveolin-1 in HAdV-D37-containing vesicles during the earliest stages of viral entry. Collectively, these experiments indicate for the first time that HAdV-D37 uses a lipid raft mediated caveolin-1 associated pathway for entry into corneal cells, and connects the processes of viral entry with downstream proinflammatory cell signaling.
Wang L, Shankarappa SA, Tong R, Ciolino JB, Tsui JH, Chiang HH, Kohane DS. Topical drug formulations for prolonged corneal anesthesia. Cornea 2013;32(7):1040-5.Abstract
PURPOSE: Ocular local anesthetics currently used in routine clinical practice for corneal anesthesia are short acting and their ability to delay corneal healing makes them unsuitable for long-term use. In this study, we examined the effect of the site 1 sodium channel blocker tetrodotoxin (TTX) on the duration of corneal anesthesia, applied with either proparacaine (PPC) or the chemical permeation enhancer octyl-trimethyl ammonium bromide (OTAB). The effect of test solutions on corneal healing was also studied. METHODS: Solutions of TTX, PPC, and OTAB, singly or in combination, were applied topically to the rat cornea. The blink response, an indirect measure of corneal sensitivity, was recorded using a Cochet-Bonnet esthesiometer, and the duration of corneal anesthesia was calculated. The effect of test compounds on the rate of corneal epithelialization was studied in vivo after corneal debridement. RESULTS: Combination of TTX and PPC resulted in corneal anesthesia that was 8 to 10 times longer in duration than that from either drug administered alone, whereas OTAB did not prolong anesthesia. The rate of corneal healing was moderately delayed after coadministration of TTX and PPC. CONCLUSIONS: Coadministration of TTX and PPC significantly prolonged corneal anesthesia, but in view of delayed corneal reepithelialization, caution is suggested in the use of the drug combination.
Schaumberg DA, Uchino M, Christen WG, Semba RD, Buring JE, Li JZ. Patient reported differences in dry eye disease between men and women: impact, management, and patient satisfaction. PLoS One 2013;8(9):e76121.Abstract
PURPOSE: Dry eye disease affects women twice as often as men, but there is little information on whether dry eye treatments, treatment satisfaction, or the impact of dry eye disease on patients' lives and vision might differ by sex. DESIGN: Questionnaire survey of 4000 participants in the Women's Health Study and the Physicians' Health Studies I and II with a prior report of a diagnosis of DED. METHODS: Among participants who re-confirmed a diagnosis of dry eye disease, we assessed symptoms, treatments, patient satisfaction and impact of dry eye disease, and analyzed differences between men and women using regression models. RESULTS: The final study population consisted of 1,518 women (mean age 70.7 years) and 581 men (mean age 76.7 years), with a mean reported duration of dry eye disease of 10.5 years and 10.1 years, respectively. The frequency and severity of dry eye disease symptoms were higher among women (each P<0.0001), and women reported a greater impact on everyday activities (P<0.0001). Women were more likely to use artificial tears (P<0.0001) use them more often (P<0.0001), and to use Restasis® (P<0.0001), omega-3 fatty acids (P=0.0006), and have punctal occlusion (P=0.005). Women spent more money per month on dry eye treatments (P<0.0001), but reported greater dissatisfaction with treatment side effects (P=0.001), and the amount of time before treatments started working (P=0.03). CONCLUSIONS: These data show that dry eye disease is generally experienced as being more severe among women, having a greater impact on their self-assessed well-being.
Palioura S, Chodosh J, Pineda R. A novel approach to the management of a progressive Descemet membrane tear in a patient with keratoglobus and acute hydrops. Cornea 2013;32(3):355-8.Abstract
PURPOSE: To report a case of corneal hydrops in a patient with keratoglobus that was managed with endothelial keratoplasty to achieve corneal stability and prevent a limbus-to-limbus tear in Descemet membrane. METHODS: A 30-year-old man with keratoglobus presented with corneal hydrops in his left eye resulting from a central vertical tear in Descemet membrane. His other eye had been previously treated with penetrating keratoplasty using a large graft (an 11-mm donor graft to a 10-mm recipient bed) because of a limbus-to-limbus tear in Descemet membrane without resolution of his edema. An attempt to approximate the edges of the Descemet tear in the left eye by an intracameral air injection failed, and the tear continued to progress peripherally. An endothelial keratoplasty button with anchoring sutures was placed over the Descemet tear because of excessive localized edema. RESULTS: One month after insertion of the sutured endothelial keratoplasty button, the edema had resolved, and 1 year later, the tear remains sealed. The patient's visual acuity improved from counting fingers at 1 foot to 20/100. CONCLUSIONS: Reconstitution of the posterior corneal surface in keratoglobus-induced hydrops can be achieved with endothelial keratoplasty over the Descemet tear. Preventing progression of a central Descemet tear is essential to bypass the need for a large-diameter penetrating keratoplasty graft and its complications in a young patient with a history of bilateral corneal hydrops.
Qazi Y, Hamrah P. Gene therapy in corneal transplantation. Semin Ophthalmol 2013;28(5-6):287-300.Abstract
Corneal transplantation is the most commonly performed organ transplantation. Immune privilege of the cornea is widely recognized, partly because of the relatively favorable outcome of corneal grafts. The first-time recipient of corneal allografts in an avascular, low-risk setting can expect a 90% success rate without systemic immunosuppressive agents and histocompatibility matching. However, immunologic rejection remains the major cause of graft failure, particularly in patients with a high risk for rejection. Corticosteroids remain the first-line therapy for the prevention and treatment of immune rejection. However, current pharmacological measures are limited in their side-effect profiles, repeated application, lack of targeted response, and short duration of action. Experimental ocular gene therapy may thus present new horizons in immunomodulation. From efficient viral vectors to sustainable alternative splicing, we discuss the progress of gene therapy in promoting graft survival and postulate further avenues for gene-mediated prevention of allogeneic graft rejection.
Sayegh RR, Dohlman CH. Wide-angle fundus imaging through the Boston keratoprosthesis. Retina 2013;33(6):1188-92.Abstract
PURPOSE: To explore the feasibility and compare the outcomes of three wide-angle fundus cameras for imaging the peripheral retina through the Type 1 Boston keratoprosthesis. METHODS: The noncontact Optos and the contact RetCam and Panoret wide-angle imaging systems were used to image the retina of eyes implanted with a keratoprosthesis. The failure-to-image rate, ease of acquisition, and quality of the images were noted, and the field of view was compared. Limitations and complications were recorded. Optos was then performed on patients referred for ultrasound B-scan evaluation, and the imaging findings were correlated. RESULTS: Retinal images with all three cameras were obtained on four eyes. Optos could be performed on all four eyes, RetCam on three, and Panoret on two. The field of view was comparable between the three different cameras. The best quality images were obtained with Optos. The external illumination of the Panoret made it impossible to image the only darkly pigmented individual in the series. Both contact devices failed to image another patient who was too agitated. Two patients had some ocular irritation from the coupling agent that resolved with replacement of the contact lens. Optos images were obtained on an additional six eyes, and findings correlated well with those on B-scan. Optos was superior to B-scan in an eye with silicone oil filling. CONCLUSION: Wide-angle fundus imaging through the keratoprosthesis is possible, and all three cameras performed similarly. The good quality of pictures obtained with the noncontact Optos, as well as its ease of use, comfort, and safety make it a preferred choice. Optos complements B-scan in the examination of the peripheral retina through the keratoprosthesis, and it may even be superior in certain settings.
Nassiri N, Eslani M, Panahi N, Mehravaran S, Ziaei A, Djalilian AR. Ocular graft versus host disease following allogeneic stem cell transplantation: a review of current knowledge and recommendations. J Ophthalmic Vis Res 2013;8(4):351-8.Abstract
Graft versus host disease (GVHD) is a common complication of allogeneic stem cell transplantation (allo-SCT). Ocular GVHD develops in approximately 40-60% of patients following allo-SCT and its most common clinical manifestations include keratoconjunctivitis sicca and cicatricial conjunctivitis. Ocular GVHD may lead to severe ocular surface disease, which can significantly diminish quality of life and restrict daily activities. It is thus important to monitor the condition closely since with timely diagnosis, irreversible damage can be avoided. The current review will focus on updated information regarding ocular GVHD.
Schrems-Hoesl LM, Schrems WA, Cruzat A, Shahatit BM, Bayhan HA, Jurkunas UV, Hamrah P. Cellular and subbasal nerve alterations in early stage Fuchs' endothelial corneal dystrophy: an in vivo confocal microscopy study. Eye (Lond) 2013;27(1):42-9.Abstract
PURPOSE: To analyze the morphology and density of corneal epithelial cells, keratocytes, and subbasal nerves, in patients with early stage Fuchs' endothelial corneal dystrophy (FECD) by in vivo confocal microscopy (IVCM). METHODS: IVCM (Confoscan 4, Nidek, Inc.) of the central cornea was performed in 30 corneas of 30 patients with early stage FECD and 13 corneas of 13 normal controls. Images were analyzed for morphology and density of the superficial and basal epithelial cells, keratocyte density, endothelial cell density (ECD), as well as subbasal corneal nerve parameters. Central corneal thickness (CCT) was measured in all patients and normals by ultrasound pachymetry. RESULTS: The ECD was significantly lower (-45.5%, P<0.001) in FECD patients as compared with controls. Total number of nerves and main nerve trunks were significantly reduced (-46.3%, P<0.001; -39.7%, P<0.001) in patients with FECD. Posterior keratocyte density was significantly higher in FECD patients (P<0.001). Significant inverse correlations were found between CCT and total number of nerves (r=-0.69, P<0.001), CCT and main nerve trunks (-0.47, P=0.016), as well as CCT and total nerve length (r=-0.62, P=0.006). Significant correlation was found between ECD and total number of nerves (r=0.44, P=0.012) as well as between ECD and main nerve trunks (r=0.65, P<0.001). CONCLUSIONS: IVCM demonstrates alterations in corneal innervation in patients with early stage FECD, suggesting a potential role of corneal nerves in the pathogenesis of FECD. Additional studies are required to investigate whether subbasal nerve alterations are caused by nonspecific corneal edema, from FECD-induced decrease in ECD, or potentially leading to loss of endothelial cells.
Palioura S, Kim B, Dohlman CH, Chodosh J. The Boston keratoprosthesis type I in mucous membrane pemphigoid. Cornea 2013;32(7):956-61.Abstract
PURPOSE: To evaluate the use of the Boston keratoprosthesis type I implantation in patients with mucous membrane pemphigoid (MMP). METHODS: Retrospective review of 8 eyes of 8 patients with severe ocular surface disease and corneal blindness as a result of MMP who underwent Boston keratoprosthesis type I implantation at the Massachusetts Eye and Ear Infirmary from January 1, 2000, through December 31, 2009. The main outcome measures were best-corrected visual acuity, keratoprosthesis retention, and postoperative complications. RESULTS: The mean age of patients was 71.3 years (range, 55-94 years), and the mean duration of their disease preoperatively was 6.1 years (range, 1.7-11.4 years). Visual acuity after the surgery improved to 20/200 or better in 6 eyes (75%) and to 20/40 or better in 3 eyes (37.5%). Only 1 of 6 eyes (16.7%) was able to maintain visual acuity of 20/200 or better over a mean follow-up period of 3.2 years. Five of the 8 Boston keratoprosthesis type I devices (62.5%) extruded or had to be replaced during a mean follow-up time of 1.7 ± 1.7 years. Loss of vision to worse than 20/200 during the follow-up period occurred because of keratoprosthesis type I extrusion, end-stage glaucoma, and retinal or choroidal detachment. CONCLUSIONS: The clinical outcomes of Boston keratoprosthesis type I implantation in MMP are guarded and, as judged from the literature, less favorable than those with the Boston keratoprosthesis type II for the same disease.
Qazi Y, Hamrah P. Corneal Allograft Rejection: Immunopathogenesis to Therapeutics. J Clin Cell Immunol 2013;2013(Suppl 9)Abstract
Corneal transplantation is among the most successful solid organ transplants. However, despite low rejection rates of grafts in the 'low-risk' setting, rejection can be as high as 70% when grafted into 'high-risk' recipient beds. Under normal homeostatic conditions, the avascular cornea provides a unique environment that facilitates immune and angiogenic privilege. An imbalance in pro-inflammatory, angiogenic and lymphangiogenic mediators leads to a breakdown in corneal immune privilege with a consequent host response against the donor graft. Recent developments in lamellar and endothelial keratoplasties have reduced the rates of graft rejection even more, while providing improved visual outcomes. The corneal layer against which an immune response is initiated, largely determines reversibility of the acute episode. While epithelial and stromal graft rejection may be treated with topical corticosteroids with higher success, acute endothelial rejection mandates a more aggressive approach to therapy due to the lack of regenerative capacity of this layer. However, current immunosuppressive regimens come with the caveat of ocular and systemic side effects, making prolonged aggressive treatment undesirable. With the advent of biologics, efficacious therapies with a superior side effect profile are on the horizon. In our review we discuss the mediators of ocular immune privilege, the roles of cellular and molecular immune players in graft rejection, with a focus on human leukocyte antigen and antigen presenting cells. Furthermore, we discuss the clinical risk factors for graft rejection and compare rates of rejection in lamellar and endothelial keratoplasties to traditional penetrating keratoplasty. Lastly, we present the current and upcoming measures of therapeutic strategies to manage and treat graft rejection, including an overview of biologics and small molecule therapy.
Li D, Shatos MA, Hodges RR, Dartt DA. Role of PKCα activation of Src, PI-3K/AKT, and ERK in EGF-stimulated proliferation of rat and human conjunctival goblet cells. Invest Ophthalmol Vis Sci 2013;54(8):5661-74.Abstract
PURPOSE: To determine the order and components of the signaling pathway utilized by epidermal growth factor (EGF) to stimulate conjunctival goblet cell proliferation. METHODS: Goblet cells from rat bulbar and forniceal conjunctiva and human bulbar conjunctiva were grown in organ culture. Goblet cells (GCs) were serum starved for 24 hours and preincubated with inhibitors for 30 minutes or small interfering RNA (siRNA) for 48 hours prior to addition of EGF. Proliferation was then measured or Western blot analysis was performed using antibodies against phosphorylated protein kinase B (AKT), extracellular signal-regulated kinase 1/2 (ERK1/2), or the non-receptor tyrosine kinase Src. Rat GCs were also incubated with adenoviruses expressing dominant negative protein kinase Cα (DNPKCα) or constitutively activated protein kinase Cα (myrPKCα), and activation of AKT and ERK1/2 was determined by Western blot analysis. RESULTS: Inhibitors of phosphoinositol-3 kinase (PI-3K)/AKT pathway blocked EGF-stimulated ERK1/2 activation and GC proliferation. Inhibitors of EGF-stimulated ERK1/2 activity did not inhibit AKT activation but blocked proliferation. DNPKCα blocked EGF-stimulated activation of AKT and ERK1/2 while myrPKCα increased activation of these kinases. Inhibitors of PI-3K, ERK1/2, and protein kinase C (PKC) blocked myrPKCα-stimulated GC proliferation. EGF and myrPKCα increased phosphorylation of Src, and inhibition of Src with the chemical inhibitor PP1 or siRNA inhibited EGF-stimulated GC proliferation. CONCLUSIONS: We found that EGF activates a major pathway to stimulate goblet cell proliferation. This pathway consists of induction of phospholipase C (PLC)γ to activate PKCα. Active PKCα phosphorylates Src to induce PI-3K to phosphorylate AKT that subsequently activates the ERK1/2 cascade to stimulate goblet cell proliferation.
Hamrah P, Cruzat A, Dastjerdi MH, Prüss H, Zheng L, Shahatit BM, Bayhan HA, Dana R, Pavan-Langston D. Unilateral herpes zoster ophthalmicus results in bilateral corneal nerve alteration: an in vivo confocal microscopy study. Ophthalmology 2013;120(1):40-7.Abstract
PURPOSE: Herpes zoster ophthalmicus (HZO), thought to be a unilateral disease, results in loss of corneal sensation, leading to neurotrophic keratopathy. This study aimed to analyze bilateral corneal nerve changes in patients with HZO by in vivo confocal microscopy (IVCM) and their correlation with corneal sensation as a measure of nerve function. DESIGN: Prospective, cross-sectional, controlled, single-center study. PARTICIPANTS: Twenty-seven eyes with the diagnosis of HZO and their contralateral clinically unaffected eyes were studied and compared with normal controls (n = 15). METHODS: In vivo confocal microscopy (Confoscan 4; Nidek Technologies, Gamagori, Japan) and corneal esthesiometry (Cochet-Bonnet; Luneau Ophthalmologie, Chartres, France) of the central cornea were performed bilaterally in all patients and controls. Patients were grouped into normal (>5.5 cm), mild (>2.5-5.5 cm), and severe (<2.5 cm) loss of sensation. MAIN OUTCOME MEASURES: Changes in corneal nerve density, total nerve number, main nerve trunks, branching, and tortuosity were evaluated after IVCM and were correlated to corneal sensation, disease duration, and number of recurrences. RESULTS: Eyes with herpes zoster ophthalmicus had a significant (P<0.001) decrease in total nerve length (595.8±358.1 vs. 2258.4±989.0 μm/frame), total number of nerves (5.4±2.8 vs. 13.1±3.8), number of main nerve trunks (2.3±1.1 vs. 4.7±1.2), and number of nerve branches (3.2±2.3 vs. 8.4±3.7) as compared with controls. In the contralateral clinically unaffected eyes, total nerve length (1053.1±441.4 μm/frame), total number of nerves (8.3±2.9), and main nerve trunks (3.1±1.0) also were decreased significantly as compared with controls (P<0.01). Reduced nerve density, total nerve count, main trunks, and tortuosity was correlated significantly with corneal sensation across all subgroups (P<0.001). CONCLUSIONS: Patients with unilateral HZO demonstrated a profound and significant bilateral loss of the corneal nerve plexus as compared with controls, demonstrating bilateral changes in a clinically unilateral disease. Loss of corneal sensation strongly correlated with subbasal nerve plexus alterations as shown by IVCM. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.
Marko CK, Menon BB, Chen G, Whitsett JA, Clevers H, Gipson IK. Spdef null mice lack conjunctival goblet cells and provide a model of dry eye. Am J Pathol 2013;183(1):35-48.Abstract
Goblet cell numbers decrease within the conjunctival epithelium in drying and cicatrizing ocular surface diseases. Factors regulating goblet cell differentiation in conjunctival epithelium are unknown. Recent data indicate that the transcription factor SAM-pointed domain epithelial-specific transcription factor (Spdef) is essential for goblet cell differentiation in tracheobronchial and gastrointestinal epithelium of mice. Using Spdef(-/-) mice, we determined that Spdef is required for conjunctival goblet cell differentiation and that Spdef(-/-) mice, which lack conjunctival goblet cells, have significantly increased corneal surface fluorescein staining and tear volume, a phenotype consistent with dry eye. Microarray analysis of conjunctival epithelium in Spdef(-/-) mice revealed down-regulation of goblet cell-specific genes (Muc5ac, Tff1, Gcnt3). Up-regulated genes included epithelial cell differentiation/keratinization genes (Sprr2h, Tgm1) and proinflammatory genes (Il1-α, Il-1β, Tnf-α), all of which are up-regulated in dry eye. Interestingly, four Wnt pathway genes were down-regulated. SPDEF expression was significantly decreased in the conjunctival epithelium of Sjögren syndrome patients with dry eye and decreased goblet cell mucin expression. These data demonstrate that Spdef is required for conjunctival goblet cell differentiation and down-regulation of SPDEF may play a role in human dry eye with goblet cell loss. Spdef(-/-) mice have an ocular surface phenotype similar to that in moderate dry eye, providing a new, more convenient model for the disease.

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