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Background Repeated surgery excisions could induce obvious irregular astigmatism in patients with recurrent pterygium. Our study is aimed to illustrate the effect of adjunct excimer laser phototherapeutic keratectomy (PTK) in limbal-conjunctival autograft transplantation on visual quality for patients with recurrent pterygium. Methods Retrospective case-control study. Eyes that underwent pterygium excision with (PTK group) or without (control group) PTK from 2006 to 2017 were retrospectively included. Recurrence rate, preoperative and postoperative surface regularity index (SRI), surface asymmetry index (SAI), cylinder and LogMAR vision were collected. Postoperative anterior segment optical coherence topography and in vivo confocal microscopy were performed to monitor the cornea epithelium healing and cellular recovery process respectively. Results A total of 99 eyes of 99 patients were collected, of which 39 were treated with PTK and 60 without PTK. The mean follow-up time was 50.4 ± 38.1 months. The recurrence rate was 10.3% (4 eyes) in the PTK group and 13.3% (8 eyes) in the control group (p = 0.759). The SRI decreased 0.53 (range: − 0.88, 2.81), SAI decreased 0.53 (range: − 0.64, 2.94), and the cylinder decreased 2.08 (range:-0.16, 9.40) D in the PTK group, and the corresponding values were 0.48 (range:-0.45, 2.27), 0.27 (range:-1.06, 2.21) and 0.71 (range:-1.75, 3.55) D in the control group, respectively (Z = 1.76, 2.15, and 3.97, p = 0.005, 0.016, and 0.000 respectively). LogMAR vision improved in both groups after surgery, with an improvement of 0.18 (range: 0.00, 0.70) in the PTK group and 0.06 (range: − 0.12, 0.50) in the control group (Z = 4.08, p = 0.000). Besides, the eyes treated with PTK showed faster re-epithelization and better cellular recovery. Conclusions For recurrent pterygium, surgical excision with adjunct PTK might be a better option with improved corneal surface and vision outcomes.

Secreted protein acidic and rich in cysteine (SPARC) shows a specific colocalization with limbal epithelial stem cells (LESCs) in vivo; however, the inherent relationship between SPARC and LESCs is still unclear. This study investigated the effects of SPARC on the maintenance of LESC stemness and corneal wound healing. To test the influence of different concentration of exogenous SPARC on the proliferation of LESCs, cell counting kit-8 assay and 5-ethynyl-2'-deoxyuridine staining were performed and the results indicated that 1 μg/mL SPARC was the optimum concentration for enhanced LESC proliferation. Compared with a control group, SPARC-treated group showed a higher expression of LESC-positive markers p63α, ABCG-2, and Bmi-1, and a lower level of differentiation marker cytokeratin-3 (CK3), thereby suggesting that SPARC could maintain LESC characteristic phenotype and suppress spontaneous epithelial differentiation in vitro. In vivo, exogenous SPARC accelerated the wound-healing process by both the enhancement of LESC proliferation and promoting the migration of the proliferating cells. However, the intact epithelium impaired this function of SPARC by contact inhibition.

Concerns regarding animal welfare have led to the need for alternatives to animal eye irritation tests. The reconstructed human cornea-like epithelium (RhCE) test is described in the OECD TG 492 as an alternative to animal eye irritation tests. However, the accuracy and labor investment of this method can be improved if the results can be predicted before the experiment. In this study, we evaluated whether Hansen solubility parameter (HSP) values can be used to predict the results of RhCE method using the LabCyte CORNEA-MODEL for 65 test substances. We found that HSP values can predict the RhCE method with high correlation (accuracy 84.6% (55/65), false-negative rate of 16.2% (7/43), and false-positive rate of 13.6% (3/22). These results indicate that HSP values can be used to predict the results RhCE method using LabCyte CORNEA-MODEL with high reproducibility, and thus are useful for evaluating the safety of substances.

PURPOSE: To assess the diagnostic values of corneal epithelial and stromal thickness distribution characteristics in forme fruste keratoconus (FFKC) and subclinical keratoconus (KC). METHODS: This cross-sectional study was conducted at VISSUM Innovation and Miguel Hernandez University, Alicante, Spain. Twenty-seven eyes (27 subjects) with FFKC, 50 eyes (50 subjects) with subclinical KC with a best spectacle corrected distance visual acuity ?20/20 (Snellen) (grade zero KC according to the Red Temática de Investigación Cooperativa en Salud classification), and 66 control eyes (66 subjects) were included. Epithelial and stromal thicknesses and epithelium/stroma (E/S) thickness ratio at center, thinnest point, 5-, and 8-mm circles obtained from the MS-39 device (CSO, Firenze, Italy) were compared among the control, FFKC, and subclinical KC groups. RESULTS: The FFKC group had thinner 8-mm superior-nasal epithelium and higher central E/S ratio compared with the control group (P < 0.05). In the subclinical KC group, the E/S ratios in the 5-mm temporal and superior zones were higher than those in the control group (P < 0.05). The FFKC and subclinical KC groups had thinner stroma compared with the control group (P < 0.05). A two-parameter formula correctly classified 94% of the eyes with subclinical KC and 98.5% of the normals, whereas another three-parameter model had 75% sensitivity and 94.3% specificity for discriminating FFKC from normals. CONCLUSIONS: This study identified different epithelial distributional and behavioral patterns in eyes with FFKC and subclinical KC. Eyes with FFKC seem to have increased central E/S ratio and asymmetric superior-nasal epithelial thinning, whereas keratometric and volumetric alterations seem to be more prominent in subclinical KC.

The need for in vitro eye irritation test replacing in vivo is steadily increasing. The MCTT HCE™ eye irritation test (EIT) using 3D reconstructed human cornea-like epithelium, was developed to identify ocular irritants from non-irritants those that are not requiring classification and labelling for eye irritation. Here, we report the results of me-too validation study, which was conducted to evaluate the reliability and relevance of the MCTT HCETM EIT, according to performance standards (PS) of OECD TG 492. The optimal cutoff to determine irritation in the prediction model was preliminarily established at 45% with the receiver operation characteristics (ROC) curve for 141 reference substances. To demonstrate the reproducibility of within- and between-laboratory (WLR and BLR), a set of 30 PS reference chemicals were tested in three laboratories three times. The WLR and BLR concordance with the binary decision of whether non-irritant or irritant was estimated to be 90-100% and 90%, respectively, and both met the PS requirements. The predictive capacity of the respective laboratories for the 30 reference chemicals were evaluated based on three different estimation methods, and the results were comparable, with sensitivity ranging from 89.6 to 93.3%, the specificity ranging from 62.2 to 66.7%, and the accuracy ranging from 75.9 to 80.0%. Additional test with the new set of 30 PS substances in the revised OECD GD 216 yielded a performance of sensitivity ranging from 92.6-93.3%, the specificity 62.2-66.7% and the accuracy 77.4-80.0%. 95.0% sensitivity, 67.2% specificity, and 83.0% accuracy were obtained for 141 reference substances in total. Furthermore, separate cutoffs for liquids and solids, 35% and 60%, respectively, produced better predictivity, which was established as a final prediction model. Collectively, our study demonstrated that MCTT HCETM EIT meets the reproducibility and predictivity criteria stated in OECD TG 492 PS.

AIM: To observe the efficacy of different artificial eye drops on corneal epithelium healing in rabbit. METHODS: Thirty-five rabbits with 6 mm diameter central corneal epithelium removed were randomly assigned to six groups: 0.9% normal saline (NS) group, 0.1% hyaluronate (HA) group, 0.3% HA group, Tears Naturale Free® (TNF) group, 0.4% polyethylene glycol (PEG) group, 0.5% carboxymethyl cellulose (CMC) group and blank control group. Treatments were administered topically four times daily. Corneal epithelium healing was evaluated by the percentage reduction in wound area at 24, 36, 48, 60, and 72h after removal of the corneal epithelium. Cornea re-epithelialization was also assessed by histological analysis and electron microscopy. RESULTS: All corneal wounds completely re-epithelialized in less than 72h. The average re-epithelialization time was 47.61±4.25h in the 0.3% HA group and 49.72±1.05h in the 0.9% NS group, followed by 0.1% HA, TNF, 0.4% PEG, 0.5% CMC, and lastly by the control group. Compared to the control group, there were significant differences among 0.3% HA, 0.9% NS, PEG, and TNF (P

Purpose: Our aim was to examine the associations between the cornea morphological parameters, endothelial cell density, and anterior chamber measurements. Methods: This observational study included 50 eyes of 50 healthy volunteers. The thicknesses of corneal epithelium, Bowman’s layer, stroma, Descemet’s membrane–endothelium complex, and pre-ocular tear film were measured by means of anterior segment optical coherence tomography. The keratometry (K), corneal volume (CV), corneal optical densitometry (COD), iridocorneal angle (ICA), anterior chamber depth (ACD), and anterior chamber volume (ACV) measurements were taken by the Scheimpflug anterior segment imaging. The corneal endothelial cell density and percentage of hexagonality were measured by specular microscopy. Results: The COD was only correlated with the thickness of Bowman’s layer (r = -0.30, p = 0.035). The hexagonal endothelial cell percentage was correlated with the thickness of the pre-ocular tear film (r = -0.50, p

Purpose We have observed noticably weak epithelial attachment in vitamin D receptor knockout mice (VDR KO) undergoing epithelial debridement. We hypothesized that VDR KO negatively affects corneal epithelial cell desmosomes and/or hemidesmosomes. Methods Transcript levels of desmosome and hemidesmosome proteins in VDR KO corneas were assessed by qPCR. Western blotting and immunochemistry were used to detect proteins in cultured cells exposed to 1,25(OH)2D3 and 24R,25(OH)2D3. Results VDR KO resulted in decreased corneal desmosomal desmoglein 1 (DSG1) and desmocollin 2 (DSC2) mRNA, and hemidesmosomal plectin mRNA. DSG1 and plectin protein expression were reduced in VDR KO corneas. DSG1 protein expression increased in VDR wild types (VDR WT) and VDR KO mouse primary epithelial cells (MPCEC) treated with 1,25(OH)2D3 and 24R,25(OH)2D3. 24R,25(OH)2D3 treatment resulted in increased plectin and integrin β4 levels in VDR WT MPCEC, and decreased levels in VDR KO MPCEC. Treatment of human corneal epithelial cells (HCEC) with 1,25(OH)2D3 and 24R,25(OH)2D3 resulted in increased DSC2 and DSG1 protein expression. Plectin and integrin β4 were only increased in 24R,25(OH)2D3 treated HCEC. Conclusions VDR KO results in reduced desmosomal and hemidesmosomal mRNA and protein levels. 1,25(OH)2D3 and 24R,25(OH)2D3 increased DSG1 protein in all cells tested. For hemidesmosome proteins, 24R,25(OH)2D3 increased plectin and integrin β4 protein expression in VDR WT and HCEC, with decreased expression in VDR KO MPCEC. Thus, vitamin D3 is involved in desmosome and hemidesmosome junction formation/regulation, and their decreased expression likely contributes to the loosely adherent corneal epithelium in VDR KO mice. Our data indicate the presence of a VDR-independent pathway.

Keeping the integrity and transparency of the cornea is the most important issue to ensure normal vision. There are more than 10 million patients going blind due to the cornea diseases worldwide. One of the effective ways to cure corneal diseases is corneal transplantation. Currently, donations are the main source of corneas for transplantation, but immune rejection and a shortage of donor corneas are still serious problems. Graft rejection could cause transplanted cornea opacity to fail. Therefore, bioengineer-based corneas become a new source for corneal transplantation. Limbal stem cells (LSCs) are located at the basal layer in the epithelial palisades of Vogt, which serve a homeostatic function for the cornea epithelium and repair the damaged cornea. LSC-based transplantation is one of the hot topics currently. Clinical data showed that the ratio of LSCs to total candidate cells for a transplantation has a significant impact on the effectiveness of the transplantation. It indicates that it is very important to accurately identify the LSCs. To date, several putative biomarkers of LSCs have been widely reported, whereas their specificity is controversial. As reported, the identification of LSCs is based on the characteristics of stem cells, such as a nuclear-to-cytoplasm ratio (N/C) ≥ 0.7, label-retaining, and side population (SP) phenotype. Here, we review recently published data to provide an insight into the circumstances in the study of LSC biomarkers. The particularities of limbus anatomy and histochemistry, the limits of the current technology level for LSC isolation, the heterogeneity of LSCs and the influence of enzyme digestion are discussed. Practical approaches are proposed in order to overcome the difficulties in basic and applied research for LSC-specific biomarkers.

Background To study basal epithelial cell (BEC), sub-basal nerve plexus (SBN) and Langerhans cell (LC) density in patients with type 2 diabetes mellitus (T2DM) with corneal punctate epitheliopathy (CPE) and to assess their association with time to healing of CPE. Methods Retrospective study of in vivo confocal microscopy (IVCM) in 160 eyes from 160 patients with T2DM diagnosed with CPE due to a single cause. Key exclusion criteria included multiple-causes for CPE or treatment with autologous serum. A total of 149 eyes from 149 gender- age- and aetiolgy-matched patients with CPE without T2DM comprised the control group. Electronic records were reviewed for demographic features, history of T2DM and aetiology of CPE. Density of BEC, SBN and LC were compared between the two groups. Results The healing time in days for CPE with different aetiologies in the T2DM and control groups were as follows: dry eye (21.56 ± 2.41; 7.00 ± 2.19; P = 0.001); meibomian gland dysfunction (26.42 ± 6.04; 9.21 ± 2.55; P = 0.001); cataract extraction (38.00 ± 19.62; 25.83 ± 11.49; P = 0.043); drug induced (53.19 ± 18.83; 41.86 ± 23.87; P = 0.018) and exposure (38.25 ± 14.13; 29.00 ± 13.67; P = 0.026). LC density was 38.70 ± 9.65 cells/mm2 in the T2DM group comparedwith 25.53 ± 3.54 cells/mm2 in the controls (P = 0.001). SBN density was 11.76 ± 1.69 mm/mm2 in the T2DM group compared with 20.92 ± 1.43 mm/mm2 in the controls (P = 0.001). BEC density in the T2DM group was 4982 ± 1178 cells/mm2 compared with 5739 ± 394 cells/mm2 in the control group (P = 0.018). Age and duration of T2DM had no relationship with healing time (multiple linear regression, P = 0.618; P = 0.787). The density of LC in the T2DM group showed a negative correlation with SBN density (r = 0.350; R2 = 0.1225; P = 0.034). The density of SBN in the T2DM group showed a positive correlation with BEC density (r = 0.427; R2 = 0.1823; P = 0.008). The density of BEC in the T2DM group showed a negative correlation with healing time (r = 0.931; R2 = 0.8668; P = 0.001). Conclusions Utilising IVCM, we have demonstrated increased LC and decreased SBN in patients with T2DM and CPE. Both may be related to lower BEC density and nuclei enhanced reflection. Furthermore, decreased BEC density may lead to delay in cornea epithelium healing in the T2DM group comparedwith controls. An immune-mediated response may play a role in delayed wound closure in patients with T2DM.

PubMed: 18035754, We investigated whether the addition of Gingko Biloba extract (EGb 761) to rabbit corneal epithelial medium before cell freezing improved cell viability after freezing then thawing. After removal of corneas, they were treated with enzymes and the corneal epithelium was prepared as a single cell suspension in freezing media with or without EGb 761. After freezing for two weeks then thawing, a higher cell viability was found in the cornea cell suspensions which had been frozen pretreated with EGb 761 in the media. The improvement with corneal cell viability with EGb 761 pretreatment is postulated to be based on the antioxidant capacity of the plant extract.

Descemet’s stripping endothelial keratoplasty is an emerging technique finalized to treat endothelial dysfunction replacing only the pathological portion of cornea. The advent of any new technique puts us in front of new complications. The epithelial ingrowth is a well-known complication already studied in case of ocular trauma and more recently in refractive surgery. This job analyzed the potential etiopathogenesis of epithelial ingrowth after DSAEK, reviewing the cases described in literature, and suggests the potential therapy.

Biomedical engineering is a unique area that allows fusion between two distinct fields of engineering and medicine. The integration of efforts from both fields promises progress through acquisition of information from tissues, cells, and organs through non-invasive methods of assessment. Here we investigate the ability of a hyperspectral device in extracting data from tissues through the wavelength spectrum, in foreseeing its potential in clinical diagnostics by simplifying methods of examination by clinicians in detecting corneal injuries. Hyperspectral imaging using 400 to 1000nm visible wavelength was used to scan five porcine eyes (injured and non-injured). Images were saved in three dimensional images of rows, columns, and depth slices at 1200 to 1300×804×604 and were processed. All laboratory works were performed in accordance with the general risk assessment of University of Strathclyde. In our results, analysis of the images reveals significant cue between 500 to 800nm bands in differentiating between injured and non-injured parts of the eye.

Background/Aim. Pterygium internum presents a significant epidemiological problem. Its etiopathogenesis is still unclear. After surgical removal it shows great tendency to recidives. The aim of this paper was to investigate connection between histopathological characteristics (inflammation presence, vascularization degree and fibrinoid changes) of pterygium and its clinical characteristics (duration, size, pterygium grade, presence of Fuchs' spots and recidives) and determine indicators for recidives prediction. Metods. We analyzed the material taken from 55 patients operated by Arlthi's method. The samples were colored by using standard histochemical methods: hematoxylin-eosin (HE), trichrome Masson painting, Gomora reticulin and PAS technique. Results. Pterygium was mostly covered by epithelium resembling conjunctival, while in the frontal cap region it showed the morphology of plate-layer cornea epithelium. The structural basis of the epithelium was composed of continuous basal lamina and continuous adhesive fibres underneath. This adhesive basis showed fibrinoid changes in the form of oval islands of various size, parallel to convexity of pterygium, or in the form of unified focus. The number, caliber and type of blood vessels showed a pronounced variability. Duration of pterygium and its size were in negative correlation with intensity of inflammation, without statistical significance. Pterygium with a longer period of develepment had negative correlation with the degree of vascularization (p > 0.05). The presence of Fuchs' spots showed a statitistically significant positive correlation with the degree of vascularization (p < 0.05). The intensity of fibrinoid changes was higher in pterygium with Fuchs' spots and in those which recurred (p > 0.05). Conclusion. Histopathological analysis of pterygium morphological shapes is a good basis for the prediction of recidives, which present the greatest problem in the treatment of this widely spread disease.

Rabbit cornea epithelium was exposed to UV-radiation for 15 min from a distance of 50 cm, and specimens were studied after 1/2, 1, and 3 h, respectively. The following changes occurred: 1) Reduced number of the large impressions or the full-thickness holes of the superficial cells. 2) Partly rejected cells. 3) Numerous small plasma membrane defects. This investigation shows plasma membrane defects at an early stage after the exposure, and this may be the locus for the initial damage.

The keratoprosthesis (KPro; “artificial cornea”) offers patients with cornea blindness, the hope of visual recovery. The first KPro designs entailed placing an optically clear material through an opaque cornea. Although introduced over 200 years ago, the basic concept is retained in many of the modern KPro designs. The Boston KPro, employs a polymethylmethacrylate (PMMA) (inert plastic) optic stabilized between a front and back plate. With over 2000 implanted world-wide to date, the Boston KPro has an excellent retention rate in patients who have failed prior penetrating keratoplasties, but lack chemical injuries or auto-immune cicatricial diseases, such as Stevens-Johnson syndrome or pemphigoid. From the Boston KPro multicenter study, the patient population who had a primary diagnosis of graft rejection retained the KPro in 97% of the eyes with a mean follow-up of 8.5 months.1 However, patients with cicatricial autoimmune disease had a retention rate of 83% and those with chemical injuries as a primary diagnosis had an 89% retention rate. Over a longer follow-up period, the retention rate decreases further for patients with autoimmune diseases and chemical injuries.2,3 For this subset of desperate patients, we continue to search for methods of increasing the retention rate of a KPro. Some KPro models differ from early designs by surrounding a central optic with a carrier or haptic that is intended to “biointegrate” with the cornea. In theory, the KPro could potentially integrate with the cornea epithelium, stroma, endothelium, or a combination of the corneal layers. Most of the “biointegratable” KPro research has focused on integration with either the cornea epithelium or the stomal keratocytes. Integration with the cornea epithelium may offer the advantage of a barrier to infection, but it would offer little structural support. Given that the stroma comprises about 90% of the cornea thickness, a type of stromal adhesion may increase the structural integrity of a KPro. Bio-integratable KPro designs can be separated into autologous biologic models and other models derived from biologic materials. These models include biologic haptics, coatings, and scaffolds. This paper will review both the clinical use of and the research on biologic KPro models.

Purpose Given the refractive importance of the human cornea, surprisingly little attention has been directed to the study of local variation in corneal refractive-index. This in vitro and in vivo study measures the refractive-index of different portions of the bovine and human cornea. Methods Fifty fresh bovine corneas (obtained from an abattoir) and 10 human subjects were used for the study. The refractive-index of the central, nasal, and temporal corneal epithelium was measured with a bench-top Abbe refractometer in the case of bovine corneas and with a hand-held refractometer with humans. Results The mean (+/-standard deviation) refractive-indices of the central, nasal, and temporal bovine corneal epithelium were 1.3760 (+/-0.003), 1.3757 (+/-0.002), and 1.3746 (+/-0.002), respectively. Refractive-indices of the anterior and posterior bovine corneal stroma were 1.3731 (+/-0.002) and 1.3708 (+/-0.004), respectively. The mean (+/-standard deviation) refractive-index in the central, nasal, and temporal periphery of the human cornea epithelium were 1.3970 (+/-0.001), 1.3946 (+/-0.001), and 1.3940 (+/-0.001), respectively. Conclusion There are small local differences in the refractive-index of the bovine and human corneal epithelium and the refractive-index of the epithelium is higher than that of the anterior and posterior stroma of the bovine cornea.

Purpose: The aim of this study was to compare the thickness of preocular tear film, corneal layers, and anterior sclera in patients with moderate to high myopia and emmetropia with anterior segment spectral domain optical coherence tomography. Methods: This cross-sectional comparative study included 31 patients with high myopia and 31 emmetropic healthy controls. Patients with myopia had axial lengths ranging from 24 to 29 mm, whereas age-matched controls had axial lengths from 21 to 23.9 mm. Patients with myopia had refractive errors from 24.00 to 211.00 diopters spherical equivalent. Preocular tear film, corneal epithelium, Bowman layer, stroma, and Descemet membrane-endothelium complex were measured in the central cornea. Anterior scleral measurements were made at distances of 1, 3, and 5 mm from the limbus. Corneal-scleral thickness was measured with the anterior segment module of spectral domain optical coherence tomography. Results: No statistically significant differences (P > 0.05) between patients with myopia and controls emerged regarding the thickness of preocular tear film, corneal epithelium, Bowman layer, corneal stroma, and Descemet membrane-endothelium complex. The anterior scleral thickness values of 1, 3, and 5 mm from the limbus were similar in both patients with myopia and control participants (P > 0.05). There were no significant correlations between central cornea stromal thickness and anterior scleral thickness in myopic participants (P > 0.05). Conclusions: The thickness of anterior wall structures and preocular tear film of patients with moderate to high myopia are not statistically different compared with those of healthy controls. Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.

AIM:To investigate the effect of amniotic membrane covering (AMC) on the healing of cornea epithelium and visual acuity for fungal keratitis after debridement.METHODS:Twenty fungal keratitis patients were divided into two groups randomly, the AMC group and the control group, ten patients each group. Both debridement of the infected cornea tissue and standard anti-fungus drugs treatments were given to every patients, monolayer amniotic membrane were sutured to the surface of the entire cornea and bulbar conjunctiva with 10-0 nylon suture for patients in the AMC group. The diameter of the ulcer was determined with slit lamp microscope and the depth of the infiltration was determined with anterior segment optical coherence tomography. Uncorrected visual acuity (UCVA) was tested before surgery and three month after healing of the epithelial layer. The healing time of the cornea epithelium, visual acuity (VA) was compared between the two groups using t-test.RESULTS:There was no statistical difference of the diameter of the ulcer, depth of the infiltration, height of the hypopyon and VA between the two groups before surgery (P＞0.05). The average healing time of the AMC group was 6.89±2.98d, which was statistically shorter than that of the control group (10.23±2.78d) (P＜0.05). The average UCVA of the AMC group was 0.138±0.083, which was statistically better than that of the control group (0.053±0.068) (P＜0.05).CONCLUSION:AMC surgery could promote healing of cornea epithelium after debridement for fungal keratitis and lead to better VA outcome.

L'application d'une concentration de 10 -6 M de melittine sur le cote lacrymal de la cornee de grenouille a augmente la conductance transepitheliale. Cet effet a ete attribue a l'ouverture d'une conductance Na + membranaire apicale. Toutefois, la puissance du venin a cette concentration (la resistance fractionnaire (fR o ) de la membrane apicale a diminue a pres de zero) a masque cet effet ainsi que d'autres. L'application de 3 x 10 -7 M de melittine du cote lacrymal a induit des effets notables (fR o > 0) et reversibles. Dans ce cas, on a pu directement observer les effets de la melittine sur les parametres de transport de Na + , Cl - et K + . On a insere des microelectrodes dans les cellules epitheliales de cornees intactes de ouaouarons dans une preparation in vitro. Dans des conditions de courant de court-circuit (I cc ), 20 min apres l'application de melittine, les effets suivants ont ete observes: I cc augmente de 8,2 par 3,1 μA/cm 2 ; fR o a chute de 51% par 18%; le potentiel intracellulaire, V o , s'est depolarise de -56,5 par 19,4 mV; la conductance transepitheliale, g t , a augmente de 0,29 par 0,57 mS/cm 2 . Des diminutions d'un facteur 10 des concentrations de Cl - ou de Na + lacrymales ont modifie les parametres de transport, ce qui est en accord avec la formation d'une conductance Na + et une augmentation de la conductance Cl - membranaire apicale induites par le venin. Les faibles effets de la melittine dans des solutions sans Cl - et, particulierement, sans Na + , sont venus etayer ces observations. Des variations de la concentration de K + n'ont eu aucun effet sur les parametres de transport. Ces resultats indiquent que l'effet de la melittine a cette faible concentration est du a une proteine des canaux Na + et non pas a des conductances aspecifiques.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:28:49Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:48:19Z : No. of bitstreams: 1 2-s2.0-84876059594.pdf: 325470 bytes, checksum: 793db00f101b8990f7ce3b874b80b0b7 (MD5) Made available in DSpace on 2014-05-27T11:28:49Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-04-01 A exemplo do que ocorre com outros fármacos, a biodisponibilidade da anfotericina B (AmB) através da via ocular repreesenta um grande desafio para a pesquisa científica. Na verdade, a baixa biodisponibilidade apresentada por tal molécula é devido, principalmente, a presença das barreiras oculares que impedem a absorção deste fármaco para os tecidos internos. A toxicidade associada a este fármaco, assim como a sua baixa capacidade em permear a córnea íntegra também consiste em um problema associado ao seu uso clínico. Novos veículos seguros e eficazes para a liberação ocular da AmB são, portanto, necessários. As microemulsões (MEs) aparecem como um interessante sistema devido à sua transparência, estabilidade termodinâmica e favorável viscosidade. O conhecimento do processo de formação das MEs contendo AmB, assim como a compreensão de toda a sua caracterização físico- química correspondem a aspectos importantes para se determinar as condições favoráveis para que estes sistemas venham a ser empregados como colírios. Esta revisão tem como objetivo, portanto, abordar este tema enfatizando os principais aspectos relacionados com a utilização de MEs como sistemas de liberação para a AmB através da via tópica oftálmica. As occurs with a number of drugs, the bioavailability of amphotericin B (AmB) used to treat fungal infections by the ocular route remains a great challenge to research scientists. In fact, the poor bioavailability of AmB is due mainly to the corneal barrier, which leads to a precorneal loss and consequent decrease in the absorption of this drug into the intraocular tissues. The toxicity associated with this molecule, together with its poor ability to penetrate the intact corneal epithelium, also represents a major drawback to its clinical use. New effective and safe drug vehicles for ocular delivery of AmB are therefore urgently needed. Microemulsions (MEs) seem to be an interesting system, owing to their transparent appearance, thermodynamic stability and favorable viscosity. Knowledge of the process of formation of AmB-containing MEs, as well as a good understanding of the physical chemistry of such systems, would provide reliable information on the best conditions for the use of these systems as eye drops. The goal of this research was thus to make an approach to this subject by reviewing the main studies on the use of MEs as delivery systems for AmB in topical eye treatment. Centro de Ciências da Saúde (CCS) Programa de Pós-Graduação em Ciências da Saúde (PPGCSa) Laboratório de Sistemas Dispersos (LaSiD), Natal-RN Centro de Ciências Biológicas e da Saúde (CCBS) Departamento de Farmácia, Campina Grande-PB Grupo de Sistemas Biomiméticos Fármacos Universidade Estadual Paulista (UNESP), Araraquara-SP Centro de Ciências da Saúde (CCS) Programa de Pós-Graduação em Ciências da Saúde (PPGCSa) Laboratorio de Sistemas Dispersos, (LASID), Universidade Federal do Rio Grande do Norte (UFRN), Natal-RN Departamento de Medicamentos Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ Grupo de Sistemas Biomiméticos Fármacos Universidade Estadual Paulista (UNESP), Araraquara-SP

Amac : Bu calismada, sicanlarda dogumdan baslayarak yasa kosut gelisimin kritik evrelerde kornea ve retinada olaylanan yapisal degisikliklerin histokimyasal yontemler ile degerlendirilmesi ve ince yapi duzeyinde karsilastirmali olarak incelenmesi amaclandi. Yontemler : Calismamizda dogumdan sonra 1. ve 22. gununde, 10. hafta ve 22. ayinda sican gozleri alinarak yasa bagli olarak kornea ve retinadaki degisiklikler PAS-Alsian mavisi boyama yontemi kullanilarak incelendi. Yapilan olcumler istatistiksel olarak degerlendirildi. Ayrica kornea ince yapisi elektron mikroskop duzeyinde resimlendirildi. Bulgular : Histokimyasal olarak yapilan boyamalarda yasa kosut olarak Descement membrani, stroma, epitel kalinligi ve genel kornea kalinliginda artis saptandi. Artan yasa kosut kornea stromasinda acilma, kollagen liflerde dalgali uzanim ayirt edildi. Retinada yapilan incelemelerde, 22 gunluk gruptan itibaren sinir lif katmaninin belirginlestigi dikkati cekti. 10 haftalik grupta bazi alanlarda yaygin intrasitoplazmik odem izlendi. Yirmi iki aylik grupta ganglion hucrelerindeki basiklasma ve sinir lif katmaninin kalinlasmasi belirgindi. Ince yapi duzeyinde kornea incelendiginde kornea epiteli, Descement membrani ve stromanin 10 haftalik eriskin yapisini kazandigi, 22 ay grubunda ise tum katmanlarda dejeneratif degisikliklerin oldugu belirlendi. Istatistiksel degerlendirmelerde; yasin ilerlemesine kosut Descement membrani, stroma, epitel kalinligi ve kornea kalinliginda artis saptandi. Sonuc : Ozellikle kritik gelisim donemlerinde hem histokimyasal hem de ince yapi duzeyinde yapilan incelemelerde gozle ilgili yapilarin da gelisim gosterdigi ancak yaslanmaya kosut bu yapilarda dejeneratif degisikliklerin gerceklestigi belirlendi. En belirgin dejenerasyon korneada saptandi. Anahtar Kelimeler: Goz, yaslanma, histokimya, ultrastruktur Investigation of Age-Related Changes on Eye by Using Histochemical and Ultrastructural Methods Objective : The purpose of this study is to evaluate structural changes in the cornea and retina of rats during the critical periods of development in parallel with aging from birth onwards through the histochemical methods and to make comparative examination at the ultrastructural level. Methods : In our study the changes in the cornea and retina in parallel with aging were examined through PAS-Alsian blue staining method by taking the eyes of rats on the 1st and 22nd day, and in the 10th week and 22nd month. Measurements made were evaluated statistically. Also the ultrastructure of the cornea was illustrated at electron microscopy level. Results : In the histochemical stainings, an increase was determined in the descement membrane, stroma, epithelial thickness and general cornea thickness in parallel with aging. An opening in the cornea stroma and fluctuant extention in the collagen fibres in parallel with aging were distinguished. In the examinations made in the retina, evidence of nerve fibre layers as from the 22nd-day group was striking. An intracytoplasmic edema was observed in some areas in the 10-week group. In the 22-month group it was clear that ganglion cells became flat and nerve fibre layer thickened. When the cornea was examined at ultrastructural level, it was determined that the cornea epithelium, descement membrane and stroma gained the structure of the 10-week adult, there were degenerative changes in all layers in the 22-month group. According the statistical analysis, increased thickness in Descement membrane, stroma and epithelial layer were observed in the cornea. Conclusion : In the examinations made at histochemical and ultrastructural level, especially during the critical development periods, it was determined that the stuctures related to the eye developed but there were degenerative changes in these structures in parallel with aging. The most distinct degeneration was determined in the cornea. Key Words: Eye, aging, histochemistry, ultrastructure

We present results of OCT and polarization-OCT applied to various ex vivo tissue samples and discuss related issues of image contrast, comparing in vivo and ex vivo preparations. Time-domain and frequency-domain OCT at 835nm and 1300nm have been applied to ex vivo skin and rabbit cornea. We can distinguish rabbit cornea epithelium for up to a month after excision. However, the skin loses all contrast upon excision and despite numerous experiments we cannot distinguish epidermis, which is clearly visible in vivo. Using a time-domain system, birefringence is clearly visible for decalcified tissue but can also be detected more weakly on fully mineralised tissue. Analysis suggests that demineralization increases the birefringence value.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:22:27Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:43:54Z : No. of bitstreams: 1 2-s2.0-34248579599.pdf: 604626 bytes, checksum: 1957b4bfbf6c652cb41fa86d4a6aa46d (MD5) Made available in DSpace on 2014-05-27T11:22:27Z (GMT). No. of bitstreams: 0 Previous issue date: 2007-04-01 • Aim: To observe the effects of topical application of 5-fluorouracil (5-FU) on the rabbits corneal limbus. • Methods: 5-FU was applied topically to the corneoscleral region of rabbits eyes. The animals were sacrificed immediately or 7, 15, 30 and 60 days later, and tissues were submitted to histological examination. • Results: All animals presented corneoscleral deepithelization close to the site of application during the immediate post-operative period, whereas on the 4th postoperative day ulceration was no longer present. Histological examination showed absence of epithelium and slight edema. After one week (G2), 2 animals presented epithelial defects, thickened epithelium with larger basal cells and loose chromatin, and slight subepithelial edema. The remaining groups showed no alterations. • Conclusion: The 5-FU topically on the corneoscleral limbus postpone the epitelization. Department of Ophthalmology Faculdade de Medicina de Botucatu, 18618-000-Botucatu-Sao, Paulo State Department of Pathology Faculdade de Medicina de Botucatu, 18618-000-Botucatu-Sao, Paulo State Department of Ophthalmology Faculdade de Medicina de Botucatu, 18618-000-Botucatu-Sao, Paulo State Department of Pathology Faculdade de Medicina de Botucatu, 18618-000-Botucatu-Sao, Paulo State

The corneal endothelium is essential for the maintenance of the corneal transparency. The aim of this study was to examine the morphology of the endothelial surface and perform morphometric analysis of the normal corneal endothelial cells of the Magellanic penguin (Spheniscus magellanicus) using scanning electron microscopy. The present work demonstrates that the corneal endothelium of the Magellanic penguin is similar to those described in other vertebrates.

Introduction The cornea is the anterior, transparent part of the collagenous wall of the eyeball. It is the window of the eye to the outer world. Its properties allow for the formation of an optical image on the light‐sensitive retina in the back of the eye. This requires transparency and regularity, but it also demands that the gross dimensions of the eye be kept constant. A regulated hydrostatic pressure within a relatively stiff eyeball accomplishes this. The aim of this chapter is to describe the human cornea. Regarding dimensions, there are of course large species variations. Functional aspects have often been studied in animal corneas and usually extended to all other species. Unless otherwise stated the text applies to the “standard human cornea”.

Purpose To describe the effect of coupling solutions used during laser photocoagulation on the ocular surface of patients with noninsulin-dependent diabetes mellitus (NIDDM). Design A prospective case-controlled study. Methods Ninety-two eyes of 46 NIDDM patients with clinically significant macular edema, poor metabolic control of diabetes, and peripheral neuropathy and 100 eyes of 50 normal control subjects were studied. The patients' eyes were assigned to argon green focal/grid laser photocoagulation using an applanation contact lens and one of the coupling fluids; 2% methocel, Thilo-Tears Gel, 1.4% sodium hyaluronate, or 0.9% simple saline. The control subjects received time-matched three-mirror contact lens fundus examinations. All subjects underwent corneal sensitivity measurements, Schirmer test, tear film breakup time, and corneal fluorescein staining before as well as 3 and 8 days after the laser procedures and contact lens examinations. Patients with corneal problems persisting after 8 days were followed longer. Results Diabetic eyes assigned to 2% methocel and 1.4% sodium hyaluronate had significantly lower mean corneal sensitivities and break-up time values as well as significantly higher mean fluorescein staining scores at all examination points after laser photocoagulation. All diabetic eyes with aqueous deficiency assigned to 2% methocel and 1.4% sodium hyaluronate developed delayed corneal epithelial healing. Conclusion The use of viscous coupling solutions during applanation contact lens–aided laser procedures may be detrimental for the corneal epithelium in poorly controlled NIDDM patients with peripheral neuropathy and coexisting aqueous deficiency.

Bu çalışma, 22 Mayıs 2003 tarihleri arasında Miami[Florida]’da düzenlenen 8. Annual Meeting of the International-Ocular-Surface-Society’da bildiri olarak sunulmuştur. Objective: To describe the ocular surface and tear function findings in premature and term babies. Design: Prospective, case-control study. Participants: Forty-eight eyes of 24 premature babies seen at the Department of Ophthalmology of Uludag University School of Medicine, Bursa, Turkey, from March 2002 through September 2002 and 50 eyes of 25 healthy term babies were studied. Intervention: The subjects underwent routine ophthalmic examinations; corneal sensitivity measurements; Schirmer test with anesthesia, with and without nasal stimulation; primary Jones test; fluorescein staining of the ocular surface; and conjunctival impression cytology. Main Outcome Measures: Premature and term babies were compared for corneal sensitivity, lacrimal drainage system patency, tear function and ocular surface staining parameters, goblet cell density, and squamous metaplasia grade. The relation of these parameters to the status of the ocular surface was also investigated. Results: Mean corneal sensitivity scores were 45 +/- 5.0 mm and 55 +/- 4.5 mm in the premature and term babies, respectively (P

In summary, we found that AMT can be helpful for the treatment of epithelial defects and stromal ulcers. We came to learn that cases with total limbal deficiency or extensive ocular surface inflammation may not be cured with AMT alone. AMT provides improvement of corneal sensitivity, tear film stability and impression cytology parameters. However, the entire mechanism of the healing effect of amniotic membrane is still unknown. A recent experimental study revealed that amniotic membrane transplantation vs tarsorraphy in murine HSV-1 stromal keratitis resulted in improvements in substance-P-related corneal nerve fibres in eyes treated with amniotic membrane, but not in eyes treated with tarsorraphy. We think that prospective, controlled, comparative studies of corneal ulcers treated by, for instance, tarsorraphy or other modalities should be performed to investigate whether the beneficial changes of amniotic membrane transplantation are greater or lesser than the changes that might have been observed by an alternative treatment. In this study, we tried to point to the effects of amniotic membrane transplantation on the ocular surface together with the problematic and unresolved issues. Thus, it is our sincere belief that further studies should be carried out along these lines.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:29:40Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:43:14Z : No. of bitstreams: 1 2-s2.0-84879942078.pdf: 528802 bytes, checksum: 81a8810f0e9a6d7b82ac2c651e0fc456 (MD5) Made available in DSpace on 2014-05-27T11:29:40Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-06-13 Corneal collagen cross-linking (CXL) has been described as a promising therapy for keratoconus. According to standard CXL protocol, epithelium should be debrided before treatment to allow penetration of riboflavin into the corneal stroma. However, removal of the epithelium can increase procedure risks. In this study we aim to evaluate stromal penetration of a biocompatible riboflavin-based nanoemulsion system (riboflavin-5-phosphate and riboflavin-base) in rabbit corneas with intact epithelium. Two riboflavin nanoemulsions were developed. Transmittance and absorption coefficient were measured on corneas with intact epithelia after 30, 60, 120, 180, and 240 minutes following exposure to either the nanoemulsions or standard 0.1% or 1% riboflavin-dextran solutions. For the nanoemulsions, the epithelium was removed after measurements to assure that the riboflavin had passed through the hydrophobic epithelium and retained within the stroma. Results were compared to de-epithelialized corneas exposed to 0.1% riboflavin solution and to the same riboflavin nanoemulsions for 30 minutes (standard protocol). Mean transmittance and absorption measured in epithelialized corneas receiving the standard 0.1% riboflavin solution did not reach the levels found on the debrided corneas using the standard technique. Neither increasing the time of exposure nor the concentration of the riboflavin solution from 0.1% to 1% improved riboflavin penetration through the epithelium. When using riboflavin-5-phosphate nanoemulsion for 240 minutes, we found no difference between the mean absorption coefficients to the standard cross-linking protocol (p = 0.54). Riboflavin nanoemulsion was able to penetrate the corneal epithelium, achieving, after 240 minutes, greater stromal concentration when compared to debrided corneas with the standard protocol (p = 0.002). The riboflavin-5-phosphate nanoemulsion diffused better into the stroma than the riboflavin-base nanoemulsion. © 2013 Bottos et al. Department of Ophthalmology Paulista School of Medicine Federal University of Sao Paulo - UNIFESP, Sao Paulo, SP Department of Pharmaceutics School of Pharmaceutical Sciences Sao Paulo State University, UNESP, Araraquara, SP Department of Informatics in Health Paulista School of Medicine Federal University of Sao Paulo - UNIFESP, Sao Paulo, SP Department of Pharmaceutics School of Pharmaceutical Sciences Sao Paulo State University, UNESP, Araraquara, SP

Multiple forms of lactic dehydrogenase (LDH) in beef cornea epithelial extract were studied by use of diethylaminoethyl (DEAE) cellulose anion-exchange chromatography. After passage through DEAE cellulose anion-exchange column, two forms of diphosphopyridine nucleotide (DPNH) linked LDH were detected. Both forms used triphosphopyridine nucleotide for the reduction of pyruvate to lactate but at a much lower rate than DPNH.

Seven human corneas were studied, using the enzyme-linked immunosorbent assay (ELISA) technique, for expression of human leukocyte antigens (HLA) class-I and class-II antigens on the epithelial cells of the central 8-mm corneal button and the remaining corneal periphery and limbus. The antigenicity was compared with that of Raji and Daudi cell lines, which served as controls. It was demonstrated that the epithelium of the two corneal parts shows a high expression of HLA class-I antigens, albeit not as strong as that found in the control cell lines. On the other hand, HLA class-II antigens were found only in the limbal-peripheral area. Expression of the HLA class-I antigens was weaker in the central portion of the cornea. It is proposed that the absence of HLA class-II antigens expression in the central portion of the cornea offers an additional explanation for the greater success of small corneal grafts, since the recognition step necessary for the immunological response is lacking.